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1. Fat Graft Retention: Adipose Tissue, Adipose-Derived Stem Cells, and Aging

Author

Trotzier, C., Sequeira, I., Auxenfans, C., Mojallal, A. A., CarMeN, laboratoire, Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), L'Oréal Recherche France (L'Oréal Recherche), L'OREAL, Queen Mary University of London (QMUL), Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL), Banque de tissus et de cellules [CHU - HCL] (Hôpital Edouard Herriot), Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Hôpital de la Croix-Rousse [CHU - HCL], and Université de Lyon

Subjects
[SDV.BA] Life Sciences [q-bio]/Animal biology, [SDV.BA]Life Sciences [q-bio]/Animal biology, Surgery
Abstract

Over the past 30 years, there has been a dramatic increase in the use of autologous fat grafting for soft-tissue augmentation and to improve facial skin quality. Several studies have highlighted the impact of aging on adipose tissue, leading to a decrease of adipose tissue volume and preadipocytes proliferation and increase of fibrosis. Recently, there has been a rising interest in adipose tissue components, including Adipose-derived Stem/Stromal Cells (ASCs) due to their regenerative potential, including inflammation, fibrosis and vascularization modulation. Due to their differentiation potential and paracrine function, ASC has been largely used for fat grafting procedures as they are described to be a key component in fat graft survival. However, many parameters as surgical procedures of adipose tissue biology could change clinical outcomes. Variation on fat grafting methods lead to numerous inconsistent clinical outcomes. Donor-to-donor variation could also be imputed to ASCs, tissue inflammatory state or tissue origin. In this review, we aim to analyze (1) the parameters involved on the graft survival, and (2) the effect of aging on adipose tissue components, especially ASCs, that could lead to a decrease of skin regeneration and fat graft retention.

Published
2022
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2. Practical approaches and optimization of a heterogeneous fluorescent tumoroid generation of ductal breast carcinoma in situ to study the impact of the adipose and inflammatory microenvironment

Author

Habanjar, Ola, Maurin, Anne-Catherine, Vituret, Cyrielle, Longechamp, Lucie, Garnier, Cecile, Decombat, Caroline, Bourgne, Céline, Auxenfans, C., Diab-Assaf, Mona, Caldefie Chezet, Florence, Delort, Laetitia, Unité de Nutrition Humaine (UNH), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Clermont Auvergne (UCA), Role of intra-Clonal Heterogeneity and Leukemic environment in ThErapy Resistance of chronic leukemias (CHELTER), Université Clermont Auvergne (UCA), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), and Université Libanaise

Subjects
[SDV.CAN]Life Sciences [q-bio]/Cancer
Abstract

International audience

Published
2022

3. Impact du microenvironnement inflammatoire et de l'obésité dans la transition du cancer canalaire in situ (CCIS) en carcinome invasif (CCI) via le développement d’un modèle 3D de tumoroïdes fluorescents hétérogènes

Author

Habanjar, Ola, Maurin, Anne-Catherine, Vituret, Cyrielle, Longechamp, Lucie, Garnier, Cecile, Decombat, Caroline, Bourgne, Céline, Auxenfans, C., Diab-Assaf, Mona, Caldefie Chezet, Florence, Delort, Laetitia, Unité de Nutrition Humaine (UNH), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Clermont Auvergne (UCA), Université Clermont Auvergne (UCA), Role of intra-Clonal Heterogeneity and Leukemic environment in ThErapy Resistance of chronic leukemias (CHELTER), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), and Université Libanaise

Subjects
[SDV.CAN]Life Sciences [q-bio]/Cancer
Abstract

International audience

Published
2022

4. A preliminary study for an intraoperative 3D bioprinting treatment of severe burn injuries

Author

Christophe, Marquette, Perrinet, Clément, Courtial, Edwin‐Joffrey, Colly, Arthur, Marquette, Christophe, Fulchiron, René, Albouy, Marion, Desanlis, Adeline, Brosset, Sophie, Auxenfans, Celine, Courtial, Edwin-Joffrey, Eli, Kyle, Cambron, Scott, Palmer, Justin, Vidal, Luciano, Thépot, Amélie, Santos, Morgan Dos, Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Ingénierie des Matériaux Polymères (IMP), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet [Saint-Étienne] (UJM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), LabSkin Creations, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, Hôpital de la Croix-Rousse [CHU - HCL], Hospices Civils de Lyon (HCL), Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Sarcomes osseux et remodelage des tissus calcifiés - Phy-Os [Nantes - INSERM U1238] (Phy-Os), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), and Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université Bretagne Loire (UBL)

Subjects
medicine.medical_specialty, 3D bioprinting, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, business.industry, law, Medicine, Severe burn, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], business, [SDV.IB.BIO]Life Sciences [q-bio]/Bioengineering/Biomaterials, Surgery, law.invention
Abstract

Background: Intraoperative three-dimensional (3D) fabrication of living tissues could be the next biomedical revolution in patient treatment. Approach: We developed a surgery-ready robotic 3D bioprinter and demonstrated that a bioprinting procedure using medical grade hydrogel could be performed using a 6-axis robotic arm in vivo for treating burn injuries.Results: We conducted a pilot swine animal study on a deep third-degree severe burn model. We observed that the use of cell-laden bioink as treatment substantially affects skin regeneration, producing in situ fibroblast growth factor (FGF) and vascular endothelial growth factor (VEGF), necessary for tissue regeneration and re-epidermalization of the wound. Innovation and Conclusion: We described an animal study of intraoperative 3D bioprinting living tissue. This emerging technology brings the first proof of in vivo skin printing feasibility using a surgery-ready robotic arm-based bioprinter. Our positive outcome in skin regeneration, joined with this procedure's feasibility, allow us to envision the possibility of using this innovative approach in a human clinical trial in the near future.

Published
2021

5. The Adipose Microenvironment Dysregulates the Mammary Myoepithelial Cells and Could Participate to the Progression of Breast Cancer

Author

François Fenaille, Caroline Decombat, Marion Vermerie, Florence Castelli, Charles Dumontet, Adrien Rossary, Laetitia Delort, Juliette Cholet, Céline Auxenfans, Florence Caldefie-Chezet, Unité de Nutrition Humaine (UNH), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Clermont Auvergne (UCA), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'Etude du Métabolisme des Médicaments (LEMM), Service de Pharmacologie et Immunoanalyse (SPI), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Médicaments et Technologies pour la Santé (MTS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut des Sciences du Vivant Frédéric JOLIOT (JOLIOT), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), and Rossary, Adrien

Subjects
0301 basic medicine, obesity, Cell type, mature adipocytes, Adipose tissue, [SDV.CAN]Life Sciences [q-bio]/Cancer, Inflammation, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, myoepithelial cells, human adipose stem cells, Cell and Developmental Biology, 03 medical and health sciences, 0302 clinical medicine, Breast cancer, [SDV.CAN] Life Sciences [q-bio]/Cancer, [SDV.BDD] Life Sciences [q-bio]/Development Biology, medicine, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, lcsh:QH301-705.5, [SDV.BDD]Life Sciences [q-bio]/Development Biology, Original Research, miRNA, Myoepithelial cell, Cancer, Cell Biology, medicine.disease, metabolomics, 3. Good health, breast cancer progression, 030104 developmental biology, lcsh:Biology (General), 030220 oncology & carcinogenesis, Cancer cell, Cancer research, medicine.symptom, Stem cell, Developmental Biology
Abstract

Breast cancer is the most common cancer among women worldwide. Overweight and obesity are now recognized as established risk factors for this pathology in postmenopausal women. These conditions are also believed to be responsible for higher recurrence and mortality rates. Reciprocal interactions have been described between adipose and cancer cells. An adipose microenvironment favors a greater proliferation of cancer cells, their invasion and even resistance to anti-cancer treatments. In addition, the chronic low-grade inflammation observed in obese individuals is believed to amplify these processes. Among the cell types present in the breast, myoepithelial cells (MECs), located at the interface of the epithelial cells and the stroma, are considered “tumor suppressor” cells. During the transition from ductal carcinomain situto invasive cancer, disorganization or even the disappearance of MECs is observed, thereby enhancing the ability of the cancer cells to migrate. As the adipose microenvironment is now considered as a central actor in the progression of breast cancer, our objective was to evaluate if it could be involved in MEC functional modifications, leading to the transition ofin situto invasive carcinoma, particularly in obese patients. Through a co-culture model, we investigated the impact of human adipose stem cells from women of normal weight and obese women, differentiated or not into mature adipocytes, on the functionality of the MECs by measuring changes in viability, apoptosis, gene, and miRNA expressions. We found that adipose cells (precursors and differentiated adipocytes) could decrease the viability of the MECs, regardless of the original BMI. The adipose cells could also disrupt the expression of the genes involved in the maintenance of the extracellular matrix and to amplify the expression of leptin and inflammatory markers. miR-122-5p and miR-132-3p could also be considered as targets for adipose cells. The metabolite analyses revealed specific profiles that may be involved in the growth of neoplastic cells. All of these perturbations could thus be responsible for the loss of tumor suppressor status of MECs and promote the transition fromin situto invasive carcinoma.

Published
2021
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6. Spen modulates lipid droplet content in adult Drosophila glial cells and protects against paraquat toxicity

Author

Girard, Victor, Goubard, Valérie, Querenet, Matthieu, Seugnet, Laurent, Pays, Laurent, Nataf, Serge, Dufourd, Eloïse, Cluet, David, Mollereau, Bertrand, Davoust, Nathalie, Laboratoire de biologie et modélisation de la cellule (LBMC UMR 5239), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche en neurosciences de Lyon - Lyon Neuroscience Research Center (CRNL), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Banque de tissus et de cellules [CHU - HCL] (Hôpital Edouard Herriot), Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), SFR Biosciences, Institut Universitaire de France (IUF), Ministère de l'Education nationale, de l’Enseignement supérieur et de la Recherche (M.E.N.E.S.R.), SFR Biosciences Projets Developpement Technologique Fondation Servier ENS Fond Recherche Fondation pour la Recherche Medicale French Ministry of Research and Education and Association France Parkinson, ANR-12-BSV1-0019,LipidinRetina,Dérégulation du métabolisme des lipides et conséquences sur la fonction et la survie des photorécépteurs: le rôle des gènes FATP chez la Drosophile et la souris(2012), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Centre de recherche en neurosciences de Lyon (CRNL), Université de Lyon-Université de Lyon-Université Jean Monnet [Saint-Étienne] (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), ROSSI, Sabine, and BLANC - Dérégulation du métabolisme des lipides et conséquences sur la fonction et la survie des photorécépteurs: le rôle des gènes FATP chez la Drosophile et la souris - - LipidinRetina2012 - ANR-12-BSV1-0019 - BLANC - VALID

Subjects
Homeodomain Proteins, Male, Paraquat, Herbicides, Disease model, Parkinson's disease, [SDV]Life Sciences [q-bio], lcsh:R, lcsh:Medicine, Gene Expression Regulation, Developmental, RNA-Binding Proteins, Parkinson Disease, Lipid Droplets, Article, [SDV] Life Sciences [q-bio], Drosophila melanogaster, nervous system, Animals, Drosophila Proteins, lcsh:Q, lcsh:Science, Astrocyte, Neuroglia
Abstract

International audience; Glial cells are early sensors of neuronal injury and can store lipids in lipid droplets under oxidative stress conditions. Here, we investigated the functions of the RNA-binding protein, SPEN/SHARP, in the context of Parkinson's disease (PD). Using a data-mining approach, we found that SPEN/SHARP is one of many astrocyte-expressed genes that are significantly differentially expressed in the substantia nigra of PD patients compared with control subjects. Interestingly, the differentially expressed genes are enriched in lipid metabolism-associated genes. In a Drosophila model of PD, we observed that flies carrying a loss-of-function allele of the ortholog split-ends (spen) or with glial cell-specific, but not neuronal-specific, spen knockdown were more sensitive to paraquat intoxication, indicating a protective role for Spen in glial cells. We also found that Spen is a positive regulator of Notch signaling in adult Drosophila glial cells. Moreover, Spen was required to limit abnormal accumulation of lipid droplets in glial cells in a manner independent of its regulation of Notch signaling. Taken together, our results demonstrate that Spen regulates lipid metabolism and storage in glial cells and contributes to glial cell-mediated neuroprotection.

Published
2020
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7. Molecular Mapping of Hydrogen Sulfide Targets in Normal Human Keratinocytes

Author

Olivia Gross-Amat, Marine Guillen, Jean-Pascal Gimeno, Michel Salzet, Nicolas Lebonvallet, Laurent Misery, Céline Auxenfans, Serge Nataf, Laboratoire sur les interactions Epithéliums Neurones (LIEN), Université de Brest (UBO), Neuroimmunologie des annélides (NA), Université de Lille, Sciences et Technologies-Centre National de la Recherche Scientifique (CNRS), Hôpital de la Cavale Blanche - CHRU Brest (CHU - BREST ), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), French association for research on spa waters (AFRETH: 'Association Francaise pour la Recherche Thermale'), INSERM, Université de Lille, Cardiovasculaire, métabolisme, diabétologie et nutrition [CarMeN], Protéomique, Réponse Inflammatoire, Spectrométrie de Masse (PRISM) - U1192, Université Claude Bernard Lyon 1 [UCBL], Laboratoire de Neurosciences de Brest [LNB], Laboratoire sur les interactions Epithéliums Neurones [LIEN], and Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 [LBTI]

Subjects
Inflammation, Keratinocytes, human keratinocyte, H2S, Inflammasomes, Superoxide Dismutase, [SDV]Life Sciences [q-bio], Apoptosis, RNAseq, proteomics, Cullin Proteins, Article, lcsh:Chemistry, lcsh:Biology (General), lcsh:QD1-999, NAD(P)H Dehydrogenase (Quinone), Humans, Hydrogen Sulfide, lcsh:QH301-705.5
Abstract

Although sulfur-rich thermal waters have ancestrally been used in the context of dermatological conditions, a global mapping of the molecular effects exerted by H2S on human keratinocytes is still lacking. To fill this knowledge gap, we subjected cultured human keratinocytes to distinct amounts of the non-gaseous hydrogen sulfur donor NaHS. We first checked that H2S accumulated in the cytoplasm of keratinocytes under our experimental conditions andused a combination of proteomics, genomics and biochemical approaches to unravel functionally relevant H2S targets in human keratinocytes. We found that the identified targets fall into two main categories: (i) the oxidative stress response molecules superoxide dismutase 2 (SOD2), NAD(P)H quinone dehydrogenase 1 (NQO1) and culin 3 (CUL3) and (ii) the chemokines interleukin-8 (IL-8) and CXCL2. Interestingly, NaHS also stimulated the caspase-1 inflammasome pathway, leading to increased secretion of the pro-inflammatory molecule interleukin-18 (IL-18). Interestingly, the secretion of interleukin-1 beta (IL-1β) was only modestly impacted by NaHS exposure despite a significant accumulation of IL-1β pro-form. Finally, we observed that NaHS significantly hampered the growth of human keratinocyte progenitors and stem cells cultured under clonogenic conditions or as epidermal cell sheets. We conclude that H2S exerts specific molecular effects on normal human keratinocytes.

Published
2020
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8. Validation of an implantable bioink using mechanical extraction of human skin cells: First steps to a 3D bioprinting treatment of deep second degree burn

Author

Patricia Rousselle, Amélie Thépot, Céline Auxenfans, Marion Albouy, Christophe A. Marquette, Morgan Dos Santos, Adeline Desanlis, Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), LabSkin Creations, Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), and Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS)

Subjects
Keratinocytes, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, 0206 medical engineering, Biomedical Engineering, Medicine (miscellaneous), Mice, Nude, Wound healing, Human skin, 02 engineering and technology, law.invention, Biomaterials, 03 medical and health sciences, Mice, Dermis, Tissue engineering, law, medicine, Animals, Humans, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Non-cultured skin cells, [SDV.IB.BIO]Life Sciences [q-bio]/Bioengineering/Biomaterials, 030304 developmental biology, 0303 health sciences, 3D bioprinting, Mice, Inbred BALB C, Tissue Scaffolds, business.industry, On-site therapy, Regeneration (biology), Bioprinting, Skin cells therapy, Cells, Immobilized, Fibroblasts, 020601 biomedical engineering, medicine.anatomical_structure, Printing, Three-Dimensional, Heterografts, Epidermis, business, Burns, Total body surface area, Biomedical engineering
Abstract

International audience; Clinical grade cultured epithelial autograft (CEA) are routinely used to treat burns covering more than 60% of the total body surface area. However, although the epidermis may be efficiently repaired by CEA, the dermal layer, which is not spared in deep burns, requires additional treatment strategies. Our aim is to develop an innovative method of skin regeneration based on in situ 3D bioprinting of freshly isolated autologous skin cells. We describe herein bioink formulation and cell preparation steps together with experimental data validating a straightforward enzyme-free protocol of skin cell extraction. This procedure complies with both the specific needs of 3D bioprinting process and the stringent rules of good manufacturing practices. This mechanical extraction protocol, starting from human skin biopsies, allows harvesting a sufficient amount of both viable and growing keratinocytes and fibroblasts. We demonstrated that a dermis may be reconstituted in vitro starting from a medical grade bioink and mechanically-extracted skin cells. In these experiments, proliferation of the extracted cells can be observed over the first 21 days period after 3D bioprinting and the analysis of type I collagen exhibited a de novo production of extracellular matrix proteins. Finally, in vivo experiments in a murine model of severe burn provided evidences that a topical application of our medical grade bioink was feasible and well-tolerated. Overall, these results represent a valuable groundwork for the design of future 3D bioprinting tissue engineering strategies aimed at treating, in a single intraoperative step, patients suffering from extended severe burns.

Published
2020
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9. Keratinocyte stem cells are more resistant to UVA radiation than their direct progeny

Author

Nicolas Bechetoille, Frédéric Demarne, Walid Rachidi, Elodie Metral, Odile Damour, Banque de Tissus et de Cellules, Laboratoire Lésions des Acides Nucléiques (LAN), Service de Chimie Inorganique et Biologique (SCIB - UMR E3), Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), SYstèmes Moléculaires et nanoMatériaux pour l’Energie et la Santé (SYMMES), Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Chimie Interface Biologie pour l’Environnement, la Santé et la Toxicologie (CIBEST ), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)

Subjects
Adult, Keratinocytes, 0301 basic medicine, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, DNA Repair, Cell division, Ultraviolet Rays, Photoaging, Primary Cell Culture, lcsh:Medicine, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Radiation Tolerance, 03 medical and health sciences, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], medicine, Humans, DNA Breaks, Single-Stranded, Progenitor cell, Clonogenic assay, lcsh:Science, ComputingMilieux_MISCELLANEOUS, Skin, Multidisciplinary, Epidermis (botany), Chemistry, Stem Cells, lcsh:R, Cell Differentiation, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Dermis, medicine.disease, Cell biology, Comet assay, 030104 developmental biology, medicine.anatomical_structure, Epidermal Cells, Pyrimidine Dimers, Female, lcsh:Q, Comet Assay, Epidermis, Stem cell, Keratinocyte, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology, DNA Damage
Abstract

The epidermis undergoes constant renewal during its lifetime. This is possible due to a special population of keratinocyte stem cells (KSCs) located at the basal layer. These cells are surrounded by their direct progeny, keratinocyte progenitors or transient amplifying cells (TAs), which arise from cell division. Skin is exposed every day to sun radiation; in particular, UVA radiation penetrates through the epidermis and induces damage to KSCs and TAs. Although keratinocytes in the basal layer are the most likely skin carcinomas and/or photoaging cells of origin, surprisingly few studies have addressed the specific responses of these cells to UV radiation. In this study, we showed for the first time that keratinocyte stem cells were more resistant to UVA irradiation than their direct progeny, transient amplifying cells. Using both the MTT assay and clonogenic assay, we found that KSCs were more photo-resistant compared to TAs after exposure to different doses of UVA (from 0 to 50 J/cm2). Moreover, KSCs had a greater ability to reconstruct human epidermis (RHE) after UVA exposure compared with TAs. Finally, investigations of DNA repair using the comet assay showed that DNA single-strand breaks and thymine dimers were repaired quicker and more efficiently in KSCs compared with TAs. In a previous work, we showed that the same stem cell population was more resistant to ionizing radiation, another carcinogenic agent. Collectively, our results combined with other observations demonstrate that keratinocyte stem cells, which are responsible for epidermal renewal throughout life, are equipped with an efficient arsenal against several genotoxic agents. Our future work will try to identify the factors or signaling pathways that are responsible for this differential photo-sensitivity and DNA repair capacity between KSCs and TAs.

Published
2018

10. Paclitaxel Treatment and Proprotein Convertase 1/3 (PC1/3) Knockdown in Macrophages is a Promising Antiglioma Strategy as Revealed by Proteomics and Cytotoxicity Studies

Author

Duhamel, Marie, Rose, Mélanie, Rodet, Franck, Murgoci, Adriana Natalia, Zografidou, Lea, Régnier-Vigouroux, Anne, Abeele, Fabien Vanden, Kobeissy, Firas, Nataf, Serge, Pays, Laurent, Wisztorski, Maxence, Cizkova, Dasa, Fournier, Isabelle, Salzet, Michel, Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Institut National de la Recherche Agronomique (INRA)

Subjects
Biochemistry & Molecular Biology, Calcium Signaling, Paclitaxel, Cell Survival, growth, [SDV]Life Sciences [q-bio], human gliomas, in-vitro, Exosomes, Cell Line, proteomics, glioma, Animals, proprotein convertase 1, breast-cancer, Secretome, taxol, Immunology, Brain Neoplasms, tumor-associated macrophages, Research, Antineoplastic Agents, Phytogenic, Coculture Techniques, peptide, quantification, Rats, macrophages, Cytokines, cells, nanoparticles, Cancer therapeutics
Abstract

International audience; High grade gliomas are the most common brain tumors in adult. These tumors are characterized by a high infiltration in microglial cells and macrophages. The immunosuppressive tumor environment is known to orient immune cells toward a pro-tumoral and anti-inflammatory phenotype. Therefore, the current challenge for cancer therapy is to find a way to reorient macrophages toward an antitumoral phenotype. Previously, we demonstrated that macrophages secreted antitumoral factors when they were invalidated for the proprotein converstase 1/3 (PC1/3) and treated with LPS. However, achieving an activation of macrophages via LPS/TLR4/Myd88-dependent pathway appears yet unfeasible in cancer patients. On the contrary, the antitumor drug Paclitaxel is also known to activate the TLR4 MyD88-dependent signaling pathway and mimics LPS action. Therefore, we evaluated if PC1/3 knock-down (KD) macrophages could be activated by Paclitaxel and efficient against glioma. We report here that such a treatment of PC1/3 KD macrophages drove to the overexpression of proteins mainly involved in cytoskeleton rearrangement. In support of this finding, we found that these cells exhibited a Ca2+ increase after Paclitaxel treatment. This is indicative of a possible depolymerization of microtubules and may therefore reflect an activation of inflammatory pathways in macrophages. In such a way, we found that PC1/3 KD macrophages displayed a repression of the anti-inflammatory pathway STAT3 and secreted more pro-inflammatory cytokines. Extracellular vesicles isolated from these PC1/3 KD cells inhibited glioma growth. Finally, the supernatant collected from the coculture between glioma cells and PC1/3 KD macrophages contained more antitumoral factors. These findings unravel the potential value of a new therapeutic strategy combining Paclitaxel and PC1/3 inhibition to switch macrophages toward an antitumoral immunophenotype.

Published
2018
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11. Long-term Genoprotection Effect of Sechium edule Fruit Extract Against UVA Irradiation in Keratinocytes

Author

Metral, Elodie, Rachidi, Walid, Damour, Odile, Demarne, Frédéric, Bechetoille, Nicolas, Laboratoire Lésions des Acides Nucléiques (LAN), Service de Chimie Inorganique et Biologique (SCIB - UMR E3), Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), SYstèmes Moléculaires et nanoMatériaux pour l’Energie et la Santé (SYMMES), Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Chimie Interface Biologie pour l’Environnement, la Santé et la Toxicologie (CIBEST ), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Banque de Tissus et de Cellules, Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Rachidi, Walid

Subjects
[SDV.BIO]Life Sciences [q-bio]/Biotechnology, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], [SDV.MHEP.DERM] Life Sciences [q-bio]/Human health and pathology/Dermatology, [SDV.BC.IC] Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BIO] Life Sciences [q-bio]/Biotechnology, [SDV.CAN] Life Sciences [q-bio]/Cancer, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], [SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], ComputingMilieux_MISCELLANEOUS, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology
Abstract

International audience

Published
2018

12. Preserving Basement Membranes during Detachment of Cultivated Oral Mucosal Epithelial Cell Sheets for the Treatment of Total Bilateral Limbal Stem Cell Deficiency

Author

Rovere, Marie-Rose, Rousselle, Patricia, Haftek, Marek, Charleux, Bruce, Kocaba, Viridiana, Auxenfans, Céline, Nataf, Serge, Damour, Odile, Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Hôpital Edouard Herriot, Laboratoire des Substituts Cutanés, Hospices Civils de Lyon (HCL), Fonctions Normales et Pathologiques de la Barrière Cutanée [Hôpital Edouard Herriot - HCL], Université de Lyon-Université de Lyon-Hôpital Edouard Herriot [CHU - HCL], Banque de tissus et cellules, Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Institut National de la Recherche Agronomique (INRA)

Subjects
épithélium, limbal stem cell deficiency, cultivated oral mucosa epithelium, enzymatic detachment, ex vivo cornea model, stem cell therapy, Swine, culture surface, [SDV]Life Sciences [q-bio], lcsh:Medicine, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Limbus Corneae, Research & Experimental Medicine, Basement Membrane, Corneal Diseases, Microscopy, Electron, Transmission, expanded ex-vivo, muqueuse buccale, cellule souche, Animals, Humans, ocular surface reconstruction, disorders, ComputingMilieux_MISCELLANEOUS, Cells, Cultured, combination, disease, Transplantation, Stem Cells, lcsh:R, cellule épithéliale, Mouth Mucosa, association, temperature, Original Articles, Cell Biology, traitement enzymatique, collagenase, ex vivo, [SDV.IB]Life Sciences [q-bio]/Bioengineering, thérapie, Stem Cell Transplantation
Abstract

International audience; Total bilateral limbal stem cell deficiency leading to loss of corneal clarity, potential vision loss, pain, photophobia, and keratoplasty failure cannot be treated by autologous limbal transplantation, and allogeneic limbal transplantation requires subsequent immunosuppressive treatment. Cultured autologous oral mucosal epithelial cells have been shown to be safe and effective alternatives. These cells can be transplanted on supports or without support after detachment from the culture dishes. Dispase, known for epidermal sheet detachment, is reported as not usable for oral mucosa. The objective was to find an optimized detachment method providing a sufficiently resistant and adhesive cultured oral mucosal epithelium (COME), which can be grafted without sutures. Enzymatic treatments (dispase or collagenase at different concentrations) were compared to enzyme-free mechanical detachment. Histological immunofluorescence (IF) and Western blotting (WB) were used to examine the impact on adhesion markers (laminin-332, 1-integrin, and type VII collagen) and junctional markers (E-cadherin, P-cadherin). Finally, the COME ability to adhere to the cornea and produce a differentiated epithelium 15 d after grafting onto an ex vivo porcine stroma model were investigated by histology, IF, and transmission electron microscopy. Collagenase at 0.5 mg/mL and dispase at 5 mg/mL were selected for comparative study on adhesive expression marker by IF and WB showed that levels of basement membrane proteins and cell-cell and cell-matrix junction proteins were not significantly different between the 3 detachment methods. Collagenase 0.5 mg/mL was selected for the next step validation because of the better reproducibility, 100% success (vs. 33% with dispase 5 mg/mL). Grafted onto porcine de-epithelialized corneal stroma, collagenase 0.5 mg/mL detached COME were found to adhere, stratify, and continue to ensure renewal of the epithelium. For COME, collagenase 0.5 mg/mL enzymatic detachment was selected and validated on its resistance and adhesive marker expression as well as their anchorage onto our new ex vivo de-epithelialized stroma model.

Published
2018
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13. Cancer du sein et obésité: impact du microenvironnement adipeux sur la fonctionnalité des cellules myoépithéliales

Author

Delort, Laetitia, Montagnon, Valentin, Bernard-Vermerie, Marion, Decombat, Caroline, Mojallal, Ali, Auxenfans, Céline, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Service de chirurgie plastique, reconstructrice et esthétique, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, and Fondation ARC pour la recherche sur le cancer

Subjects
[SDV.AEN]Life Sciences [q-bio]/Food and Nutrition, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract

National audience; Introduction et but de l’étude: L'obésité est reconnue comme un facteur de risque de cancer du sein chez les femmes ménopausées, également responsable de taux plus élevés de récidive et de mortalité. Parmi l’ensemble des types cellulaires présents au niveau du sein, les cellules myoépithéliales (CME), peu étudiées à ce jour, forment une monocouche séparant les cellules épithéliales de la membrane basale et sont considérées comme des "suppresseurs de tumeurs" qui favorisent l'inhibition de la croissance tumorale, de l'invasion ou de l'angiogenèse. Au cours de la transition d’un carcinome canalaire in situ (CCIS) en carcinome canalaire invasif (CCI), une désorganisation voire une disparition complète des CME est observée, ce qui augmente la capacité de migration des cellules cancéreuses. Lee microenvironnement adipeux étant actuellement reconnu comme un élément favorisant la carcinogenèse mammaire, nous avons souhaité caractériser son rôle sur la disparition et la fonctionnalité des cellules myoépithéliales, et ce, plus particulièrement, dans un contexte d'obésité. Matériel et méthodes: Pour cela, des CME (Hs 578Bst) ont été co-cultivées en présence de lignées de cellules souches adipeuses (ASC) (hMAD, 3T3F442a) ou d’ASC obtenues à partir de patientes minces ou obèses, et différenciées par la suite en adipocytes matures (AM). Après 72h de co-culture, la prolifération des CME a été mesurée par un test fluorescent à la résazurine (Fluoroskan Ascent FL®) et l'apoptose évaluée (annexine V-FITC/PI). En parallèle, un modèle de culture tridimensionnelle de sphéroïdes a été développé afin d’évaluer l'influence du secrétome adipocytaire sur la structure et la fonctionnalité des CME. Résultats et Analyse statistique : En présence de hMAD différenciées ou non en AM, une diminution significative de la viabilité des CME (-22%, n=4, p=0,05 ; -11%, n=3, p=0,01 respectivement) associée à une tendance à l’augmentation de l’apoptose a été montrée. Une diminution non significative de la prolifération des CME est observée en présence des cellules adipeuses 3T3-F442A alors qu’aucun effet n’est observé en présence des ASC prélevées sur des femmes minces et obèses. Conclusion: Ces résultats préliminaires montrent que les cellules adipeuses et leurs sécrétions influencent le comportement des CME. Les modèles de co-culture et de sphéroïdes développés avec les CME permettront d’avoir une meilleure compréhension de l’influence du surpoids et de l’obésité sur la fonctionnalité de ces cellules, et afin, à terme, d’identifier de nouvelles cibles thérapeutiques et préventives du cancer du sein. Remerciements: Projet financé par la Fondation ARC pour la recherche sur le cancer

Published
2017

14. Selenium preserves keratinocyte stemness and delays senescence by maintaining epidermal adhesion

Author

David Béal, Eric Blouin, Anne-Marie Roussel, Lara Jobeili, Walid Rachidi, Patricia Rousselle, Odile Damour, Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Environnement Ville Société (EVS), École normale supérieure - Lyon (ENS Lyon)-École des Mines de Saint-Étienne (Mines Saint-Étienne MSE), Institut Mines-Télécom [Paris] (IMT)-Institut Mines-Télécom [Paris] (IMT)-Université Lumière - Lyon 2 (UL2)-Université Jean Moulin - Lyon 3 (UJML), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet [Saint-Étienne] (UJM)-École Nationale des Travaux Publics de l'État (ENTPE)-École nationale supérieure d'architecture de Lyon (ENSAL)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de bioénergétique fondamentale et appliquée (LBFA), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes (UGA), Laboratoire Lésions des Acides Nucléiques (LAN), Service de Chimie Inorganique et Biologique (SCIB - UMR E3), Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes (UGA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Université Grenoble Alpes (UGA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Environnement, Ville, Société (EVS), École normale supérieure de Lyon (ENS de Lyon)-École des Mines de Saint-Étienne (Mines Saint-Étienne MSE), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet - Saint-Étienne (UJM)-École Nationale des Travaux Publics de l'État (ENTPE)-École nationale supérieure d'architecture de Lyon (ENSAL)-Centre National de la Recherche Scientifique (CNRS), Laboratory of Fundamental and Applied Bioenergetics = Laboratoire de bioénergétique fondamentale et appliquée (LBFA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS), Labcatal Pharmaceuticals, ANR [ANR-13-RPIB-0003-01], Biologie et physiopathologie cutanées : expression génique, signalisation et thérapie, Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015 - 2019) (COMUE UCA)-IFR50-Institut National de la Santé et de la Recherche Médicale (INSERM), SYstèmes Moléculaires et nanoMatériaux pour l’Energie et la Santé (SYMMES), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Banque de Tissus et de Cellules, Chimie Interface Biologie pour l’Environnement, la Santé et la Toxicologie (CIBEST ), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National de la Recherche Agronomique (INRA), and Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet [Saint-Étienne] (UJM)-École Nationale des Travaux Publics de l'État (ENTPE)-École nationale supérieure d'architecture de Lyon (ENSAL)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Keratinocytes, Aging, Antioxidant, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, Time Factors, medicine.medical_treatment, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Antioxidants, Basement Membrane, Skin Aging, 0302 clinical medicine, complémentation, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], selenium, Geriatry and gerontology, Cells, Cultured, Cellular Senescence, ComputingMilieux_MISCELLANEOUS, kératinocyte, integumentary system, Stem Cells, food and beverages, Cell Differentiation, Environmental exposure, vieillissement, 3. Good health, Cell biology, adhesion, medicine.anatomical_structure, Phenotype, replicative life span, skin aging, keratinocytes stem cells, Keratinocyte, Research Paper, Senescence, Cell Survival, chemistry.chemical_element, sénescence, [SDV.CAN]Life Sciences [q-bio]/Cancer, 03 medical and health sciences, Sodium Selenite, antioxydant, adhésion cellulaire, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], medicine, Cell Adhesion, Skin equivalent, Humans, Cell Proliferation, différenciation cellulaire, Dose-Response Relationship, Drug, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Cell Biology, medicine.disease, 030104 developmental biology, chemistry, peau, Gériatrie et gérontologie, Skin cancer, Epidermis, 030217 neurology & neurosurgery, Selenium, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology
Abstract

International audience; Skin is constantly exposed to environmental factors such as pollutants, chemicals and ultra violet radiation (UV), which can induce premature skin aging and increase the risk of skin cancer. One strategy to reduce the effect of oxidative stress produced by environmental exposure is the application of antioxidant molecules. Among the endogenous antioxidants, selenoproteins play a key role in antioxidant defense and in maintaining a reduced cellular environment. Selenium, essential for the activity of selenoproteins, is a trace element that is not synthesized by organisms and must be supplied by diet or supplementation. The aim of this study is to evaluate the effect of Selenium supplementation on skin aging, especially on keratinocytes, the main cells of the epidermis. Our results demonstrate for the first time to our knowledge, the major role of Selenium on the replicative life span of keratinocytes and on aging skin. Selenium protects keratinocyte stem cells (KSCs) against senescence via preservation of their stemness phenotype through adhesion to the basement membrane. Additionally, Selenium supplementation maintains the homeostasis of skin during chronological aging in our senescent skin equivalent model. Controlled supplementation with Selenium could be a new strategy to protect skin against aging.

Published
2017
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15. Evaluation of Three Devices for the Isolation of the Stromal Vascular Fraction from Adipose Tissue and for ASC Culture: A Comparative Study

Author

Ali Mojallal, F. Boucher, Jonathan Rodriguez, Cyrille Debard, Hugo Fabre, Odile Damour, Nacira Abbassi, Anne-Sophie Pratta, Fanny Rodriguez, Frédéric Mallein-Gerin, Jacqueline Adobati, Céline Auxenfans, Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), Service Chirurgie Plast Esthétique et Reconstructrice, Hospices Civils de Lyon (HCL), Institut de biologie et chimie des protéines [Lyon] (IBCP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), chirurgie plastique, reconstructrice et esthétique, Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Institut National de la Recherche Agronomique (INRA)

Subjects
0301 basic medicine, lcsh:Internal medicine, Pathology, medicine.medical_specialty, Stromal cell, Article Subject, Population, Adipose tissue, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Biology, Flow cytometry, 03 medical and health sciences, medicine, lcsh:RC31-1245, education, Molecular Biology, ComputingMilieux_MISCELLANEOUS, education.field_of_study, medicine.diagnostic_test, Cell Biology, Stromal vascular fraction, Chondrogenesis, Phenotype, Cell biology, 030104 developmental biology, Adipogenesis, Research Article
Abstract

Adipose-derived stem/stromal cells (ASCs) reside in the stromal vascular fraction (SVF) of adipose tissue (AT) and can be easily isolated. However, extraction of the SVF from lipoaspirate is a critical step in generating ASC, and semiautomated devices have been developed to enhance the efficacy and reproducibility of the outcomes and to decrease manipulation and contamination. In this study, we compared the reference method used in our lab for SVF isolation from lipoaspirate, with three medical devices: GID SVF-1™, Puregraft™, and Stem.pras®. Cell yield and their viability were evaluated as well as their phenotype with flow cytometry. Further on, we determined their proliferative potential using population doublings (PD), PD time (PDT), and clonogenicity assay (CFU-F). Finally, we checked their genetic stability using RT-qPCR for TERT mRNA assay and karyotyping as well as their multilineage potential including adipogenic, chondrogenic, and osteogenic differentiation. Our results demonstrate that all the devices allow the production of SVF cells with consistent yield and viability, in less time than the reference method. Expanded cells from the four methods showed no significant differences in terms of phenotype, proliferation capabilities, differentiation abilities, and genetic stability.

Published
2017
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16. α6 Integrin (α6high)/Transferrin Receptor (CD71)low Keratinocyte Stem Cells Are More Potent for Generating Reconstructed Skin Epidermis Than Rapid Adherent Cells

Author

Nicolas Bechetoille, Odile Damour, W. Rachidi, Elodie Metral, Frédéric Demarne, Rachidi, Walid, Laboratoire Lésions des Acides Nucléiques (LAN), Service de Chimie Inorganique et Biologique (SCIB - UMR E3), Institut Nanosciences et Cryogénie (INAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut Nanosciences et Cryogénie (INAC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS), SYstèmes Moléculaires et nanoMatériaux pour l’Energie et la Santé (SYMMES), Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Chimie Interface Biologie pour l’Environnement, la Santé et la Toxicologie (CIBEST ), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Banque de Tissus et de Cellules, Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)

Subjects
0301 basic medicine, Keratinocytes, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Cell Separation, [SDV.BC.IC] Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Integrin alpha6, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, lcsh:Chemistry, 0302 clinical medicine, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], lcsh:QH301-705.5, Spectroscopy, ComputingMilieux_MISCELLANEOUS, education.field_of_study, medicine.diagnostic_test, keratinocyte stem cells, Stem Cells, Temperature, skin, epidermis, basal keratinocytes, adhesion, α6 integrin, CD71, General Medicine, Cell sorting, Flow Cytometry, Computer Science Applications, Cell biology, medicine.anatomical_structure, Phenotype, 030220 oncology & carcinogenesis, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Stem cell, Keratinocyte, Population, Transferrin receptor, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, Catalysis, Article, Collagen Type I, Flow cytometry, Inorganic Chemistry, Colony-Forming Units Assay, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, Antigens, CD, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Receptors, Transferrin, medicine, [SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Cell Adhesion, Humans, Regeneration, Physical and Theoretical Chemistry, education, Molecular Biology, Epidermis (botany), Organic Chemistry, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.MHEP.DERM] Life Sciences [q-bio]/Human health and pathology/Dermatology, Epithelium, [SDV.BIO] Life Sciences [q-bio]/Biotechnology, Clone Cells, 030104 developmental biology, lcsh:Biology (General), lcsh:QD1-999, Epidermal Cells, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology
Abstract

The epidermis basal layer is composed of two keratinocyte populations: Keratinocyte Stem cells (KSC) and Transitory Amplifying (TA) cells that arise from KSC division. Unfortunately, no specific marker exists to differ between KSC and TA cells. Here, we aimed at comparing two different methods that pretended to isolate these two populations: (i) the rapid adhesion method on coated substrate and (ii) the flow cytometry method, which is based on the difference in cell surface expressions of the α6 integrin and transferrin receptor (CD71). Then, we compared different parameters that are known to discriminate KSC and TA populations. Interestingly, we showed that both methods allow enrichment in stem cells. However, cell sorting by flow cytometry (α6high/CD71low) phenotype leads to a better enrichment of KSC since the colony forming efficiency is five times increased versus total cell suspension, whereas it is only 1.4 times for the adhesion method. Moreover, α6high/CD71low cells give rise to a thicker pluristratified epithelium with lower seeding density and display a low Ki67 positive cells number, showing that they have reached the balance between proliferation and differentiation. We clearly demonstrated that cells isolated by a rapid adherent method are not the same population as KSC isolated by flow cytometry following α6high/CD71low phenotype.

Published
2017

17. A unique TGFB1-driven genomic program links Astrocytosis, low-grade inflammation and partial demyelination in spinal cord periplaques from progressive multiple sclerosis patients

Author

Nataf, Serge, Barritault, Marc, Pays, Laurent, Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN), Institut National de la Recherche Agronomique (INRA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre de Recherche en Cancérologie de Lyon (CRCL), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Groupement Hospitalier Est, Hospices Civils de Lyon, Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), INSA lyon, Direction de la Recherche, and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
chimiokine, Multiple Sclerosis, approche transcriptomique, [SDV]Life Sciences [q-bio], Cell Cycle Proteins, Biotechnologies, Interleukin-12 Subunit p35, Article, identification de gènes, neuroinflammation, lcsh:Chemistry, Transforming Growth Factor beta1, myéline, Glial Fibrillary Acidic Protein, Autre (Chimie), cytokine, Humans, sclérose en plaques, lcsh:QH301-705.5, bioinformatique, ComputingMilieux_MISCELLANEOUS, Myelin Sheath, Inflammation, Platelet-Derived Growth Factor, Lymphokines, Interleukin-17, Intracellular Signaling Peptides and Proteins, astrocytes, Computational Biology, bioinformatics, Interleukin-33, Interleukin-12, [SDV] Life Sciences [q-bio], Oligodendroglia, myelin, lcsh:Biology (General), lcsh:QD1-999, Spinal Cord, Connexin 43, Other, multiple sclerosis
Abstract

International audience; We previously reported that, in multiple sclerosis (MS) patients with a progressive form of the disease, spinal cord periplaques extend distance away from plaque borders and are characterized by the co-occurrence of partial demyelination, astrocytosis and low-grade inflammation. However, transcriptomic analyses did not allow providing a comprehensive view of molecular events in astrocytes vs. oligodendrocytes. Here, we re-assessed our transcriptomic data and performed co-expression analyses to characterize astrocyte vs. oligodendrocyte molecular signatures in periplaques. We identified an astrocytosis-related co-expression module whose central hub was the astrocyte gene Cx43/GJA1 (connexin-43, also named gap junction protein α-1). Such a module comprised GFAP (glial fibrillary acidic protein) and a unique set of transcripts forming a TGFB/SMAD1/SMAD2 (transforming growth factor β/SMAD family member 1/SMAD family member 2) genomic signature. Partial demyelination was characterized by a co-expression network whose central hub was the oligodendrocyte gene NDRG1 (N-myc downstream regulated 1), a gene previously shown to be specifically silenced in the normal-appearing white matter (NAWM) of MS patients. Surprisingly, besides myelin genes, the NDRG1 co-expression module comprised a highly significant number of translation/elongation-related genes. To identify a putative cause of NDRG1 downregulation in periplaques, we then sought to identify the cytokine/chemokine genes whose mRNA levels inversely correlated with those of NDRG1. Following this approach, we found five candidate immune-related genes whose upregulation associated with NDRG1 downregulation: TGFB1(transforming growth factor β 1), PDGFC (platelet derived growth factor C), IL17D (interleukin 17D), IL33 (interleukin 33), and IL12A (interleukin 12A). From these results, we propose that, in the spinal cord periplaques of progressive MS patients, TGFB1 may limit acute inflammation but concurrently induce astrocytosis and an alteration of the translation/elongation of myelin genes in oligodendrocytes.

Published
2017
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18. Supernatants of Adipocytes From Obese Versus Normal Weight Women and Breast Cancer Cells: In Vitro Impact on Angiogenesis

Author

Lauriane, Bougaret, Laetitia, Delort, Hermine, Billard, Charlotte, Lequeux, Nicolas, Goncalves-Mendes, Ali, Mojallal, Odile, Damour, Marie-Paule, Vasson, Florence, Caldefie-Chezet, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Faculté de Pharmacie, Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Banque de tissus et de cellules, Chirurgie Plastique, Reconstructrice et Esthétique, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Cancéropôle Lyon Auvergne Rhône-Alpes, Contract grant sponsor: The French Anti-Cancer League office (Ligue Contre le Cancer CD63), Contract grant sponsor: Canceropole Lyon-Auvergne-Rhone-Alpes (CLARA ONCOSTARTER), Contract grant sponsor: INCA (PL BIO projet Mammadipo No 6666), Contract grant sponsor: The French Ministry of Research and Technology, Unité de Nutrition Humaine - Clermont Auvergne (UNH), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne (UCA), Université Clermont Auvergne (UCA), and Centre Jean Perrin - Centre Régional de Lutte contre le Cancer d'Auvergne

Subjects
patiente obèse, Breast Neoplasms, [SDV.CAN]Life Sciences [q-bio]/Cancer, mammary malignant tumor, adipocyte, Body Mass Index, stratégie préventive, Cell Movement, Adipocytes, Human Umbilical Vein Endothelial Cells, Humans, Obesity, Cell Proliferation, Neovascularization, Pathologic, Staining and Labeling, cancer du sein, Body Weight, Cell Differentiation, Antibodies, Neutralizing, Vascular Endothelial Growth Factor Receptor-2, Adipose Tissue, Gene Expression Regulation, Culture Media, Conditioned, femme menopausée, MCF-7 Cells, Receptors, Leptin, Female, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, lipocytes, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Subcellular Fractions
Abstract

Breast cancer is correlated with a higher risk of metastasis in obese postmenopausal women. Adipokines, whose plasma concentrations are modulated in obese subjects and adipocytes surround mammary cells, suggesting that adipocyte secretome affect mammary tumorogenesis. We hypothesize that mature adipocyte secretions from obese women conditioned or not by breast neoplasic cells, increase changes on the angiogenesis stages. Supernatants of human mature adipocytes, differentiated from stem cells of either adipose tissue of normal weight (MA20) or obese (MA30) women or obtained from co-cultures between MA20 and MA30 and breast cancer cell line MCF-7, were collected. The impact of these supernatants was investigated on proliferation, migration, and tube formation by endothelial cells (HUVEC). MA20 and MA30 showed a preservation of their "metabolic memory" (increase of Leptin, ObR, VEGF, CYP19A1, and a decrease of Adiponectin expression in MA30 compared to MA20). Supernatants from obese-adipocytes increased HUVEC proliferation, migration, and sprouting like with supernatants obtained from co-cultures of MA/MCF-7 regardless the women's BMI. Additional analyses such as the use of neutralizing antibodies, analysis of supernatants (Milliplex®) and variations in gene expression (qRT-PCR), strongly suggest an implication of IL-6, or a synergistic action among adipokines, probably associated with that of VEGF or IL-6. As a conclusion, supernatants from co-cultures of MA30 and MCF-7 cells increase proliferation, migration, and sprouting of HUVEC cells. These results provide insights into the interaction between adipocytes and epithelial cancer cells, particularly in case of obesity. The identification of synergistic action of adipokines would therefore be a great interest in developing preventive strategies. J. Cell. Physiol. 232: 1808-1816, 2017. © 2016 Wiley Periodicals, Inc.

Published
2017
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19. Impact du secrétome adipocytaire sur l’hormonothérapie dans le cancer du sein en situation de surpoids/obésité

Author

Delort, Laetitia, Decombat, Caroline, Bernard-Vermerie, M., VASSON, Marie-Paule, Mojallal, A., Auxenfans, C., Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA), Faculté de Pharmacie, Service de Nutrition, Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER-UNICANCER, Centre Hospitalier Universitaire de Lyon, Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), and Centre de Recherche en Nutrition Humaine (CRNH). Clermont-Ferrand, FRA.

Subjects
[SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition
Abstract

National audience; L’obésité, facteur de risque établi de cancer du sein chez les femmes ménopausées, est aussi responsable de plus forts taux de récidives et de mortalité. Dans ce contexte, nos travaux évaluent l’impact du secrétome adipocytaire (SA) dans la moindre réponse à l’hormonothérapie. Pour cela, 1/ des cellules cancéreuses mammaires ont été soit co-cultivées avec des cellules souches adipeuses (CSAd) humaines (hMAD) différenciées en adipocytes matures (AM) soit cultivées en présence de milieux conditionnés (MC), puis traitées avec un anti-estrogène (anti-E ; Tamoxifène Tx ou Fulvestrant Fv). La prolifération cellulaire a été mesurée par fluorescence à la résazurine (Fluoroskan Ascent FL®, n=3) et suivie en temps réel par impédancemétrie (iCELLigence, n=3). 2/ L’impact du surpoids a été évalué en utilisant des AM différenciés à partir de CSAd de femmes minces ou obèses cultivées en présence de cellules MCF-7 et de Fv. Dans nos différents modèles, les SA sont capables de majorer la prolifération des cellules MCF-7, récepteur aux estrogènes positives (RE+), et d’inhiber totalement l’effet antiprolifératif du Tx et du Fv. L’utilisation de cellules MDA-MB-231 RE- a montré que le SA majore la prolifération cellulaire et contrecarre l’effet antiprolifératif du Tx, suggérant que les effets observés ne passeraient pas exclusivement par la voie du RE dans notre modèle. Par ailleurs, en utilisant des AM de femmes de poids variables, seuls les SA correspondant aux femmes d’IMC>30 amoindrissent l’efficacité du Fv, cet effet s’accompagnant d’une augmentation de l’expression du gène OB-R. Ainsi, nos résultats préliminaires suggèrent que le SA limiterait l’efficacité de l’hormonothérapie et ce, de façon plus prononcée en cas de surpoids, ce qui pourrait contribuer au processus d’échappement tumoral

Published
2016

20. Perlecan expression influences the keratin 15‐positive cell population fate in the epidermis of aging skin

Author

Patricia Rousselle, Sophie Boulesteix, Morgan Dos Santos, Christine Guicher, Odile Damour, Guila Dayan, John M. Whitelock, Anna Michopoulou, V. André-Frei, Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), BASF BCS France, University of New South Wales [Sydney] (UNSW), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], and Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL)

Subjects
0301 basic medicine, Keratinocytes, Male, Aging, Cellular differentiation, [SDV]Life Sciences [q-bio], Extracellular matrix, 0302 clinical medicine, Keratin, chemistry.chemical_classification, education.field_of_study, biology, integumentary system, Cell Differentiation, Middle Aged, Cell biology, Extracellular Matrix, medicine.anatomical_structure, Biochemistry, Female, Keratinocyte, Research Paper, Adult, keratin 15, Materials science, Population, keratinocyte, Perlecan, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 03 medical and health sciences, Young Adult, medicine, Humans, education, [SDV.IB.BIO]Life Sciences [q-bio]/Bioengineering/Biomaterials, skin aging, Aged, Basement membrane, Epidermis (botany), Keratin-15, Cell Biology, basement membrane, carbohydrates (lipids), perlecan, stem cell, 030104 developmental biology, chemistry, Epidermal Cells, biology.protein, Epidermis, 030217 neurology & neurosurgery, Heparan Sulfate Proteoglycans
Abstract

International audience; The epidermis is continuously renewed by stem cell proliferation and differentiation. Basal keratinocytes append the dermal‐epidermal junction, a cell surface‐associated, extracellular matrix that provides structural support and influences their behaviour. It consists of laminins, type IV collagen, nidogens, and perlecan, which are necessary for tissue organization and structural integrity. Perlecan is a heparan sulfate proteoglycan known to be involved in keratinocyte survival and differentiation. Aging affects the dermal epidermal junction resulting in decreased contact with keratinocytes,thus impacting epidermal renewal and homeostasis. We found that perlecan expression decreased during chronological skin aging. Our in vitro studies revealed reduced perlecan transcript levels in aged keratinocytes. The production of in vitro skin models revealed that aged keratinocytes formed a thin and poorly organized epidermis. Supplementing these models with purified perlecan reversed the phenomenon allowing restoration of a well‐differentiated multi‐layered epithelium. Perlecan down‐regulation in cultured keratinocytes caused depletion of the cell population that expressed keratin 15. This phenomenon depended on the perlecan heparan sulphate moieties, which suggested the involvement of a growth factor.Finally, we found defects in keratin 15 expression in the epidermis of aging skin. This study highlighted a new role for perlecan in maintaining the self‐renewal capacity of basal keratinocytes.

Published
2016
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21. Cartilage-Characteristic Matrix Reconstruction by Sequential Addition of Soluble Factors During Expansion of Human Articular Chondrocytes and Their Cultivation in Collagen Sponges

Author

Stéphanie Claus, Muriel Piperno, Elisabeth Aubert-Foucher, Odile Damour, Nathalie Mayer, Emeline Perrier-Groult, Frédéric Mallein-Gerin, Jérôme E. Lafont, Hanane Chajra, Institut de biologie et chimie des protéines [Lyon] (IBCP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biologie Tissulaire et d'ingénierie Thérapeutique UMR 5305 (LBTI), Service de rhumatologie, Hospices Civils de Lyon (HCL)-Affiliation nécessaire..., Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon

Subjects
Cartilage, Articular, [SDV]Life Sciences [q-bio], Biomedical Engineering, Bone Morphogenetic Protein 2, Medicine (miscellaneous), Bioengineering, Bone morphogenetic protein, Organ culture, Fibroblast growth factor, Mice, 03 medical and health sciences, Chondrocytes, 0302 clinical medicine, Nude mouse, Tissue engineering, Gene expression, medicine, Animals, Humans, Insulin, Cells, Cultured, ComputingMilieux_MISCELLANEOUS, Aged, Cell Proliferation, 030304 developmental biology, 0303 health sciences, Tissue Engineering, Tissue Scaffolds, biology, Chemistry, Cartilage, Middle Aged, biology.organism_classification, Immunohistochemistry, In vitro, Extracellular Matrix, Cell biology, medicine.anatomical_structure, Gene Expression Regulation, Solubility, Protein Biosynthesis, 030220 oncology & carcinogenesis, Intercellular Signaling Peptides and Proteins, Triiodothyronine, Cattle, Fibroblast Growth Factor 2, Collagen, Chondrogenesis, Biomedical engineering
Abstract

Articular cartilage has a poor capacity for spontaneous repair. Tissue engineering approaches using biomaterials and chondrocytes offer hope for treatments. Our goal was to test whether collagen sponges could be used as scaffolds for reconstruction of cartilage with human articular chondrocytes. We investigated the effects on the nature and abundance of cartilage matrix produced of sequential addition of chosen soluble factors during cell amplification on plastic and cultivation in collagen scaffolds.Isolated human articular chondrocytes were amplified for two passages with or without a cocktail of fibroblast growth factor (FGF)-2 and insulin (FI). The cells were then cultured in collagen sponges with or without a cocktail of bone morphogenetic protein (BMP)-2, insulin, and triiodothyronine (BIT). The constructs were cultivated for 36 days in vitro or for another 6-week period in a nude mouse-based contained-defect organ culture model. Gene expression was analyzed using polymerase chain reaction, and protein production was analyzed using Western-blotting and immunohistochemistry.Dedifferentiation of chondrocytes occurred during cell expansion on plastic, and FI stimulated this dedifferentiation. We found that addition of BIT could trigger chondrocyte redifferentiation and cartilage-characteristic matrix production in the collagen sponges. The presence of FI during cell expansion increased the chondrocyte responsiveness to BIT.

Published
2012
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22. Rôle du secretome adipocytaire dans la resistance au traitement par hormonotherapie et sur les cellules souches cancereuses : cas du cancer mammaire en situation d’obésité

Author

Delort, Laetitia, Bougaret, Laurianne, Mojallal, Ali, Damour, Odile, Auxenfans, Céline, Vasson, Marie-Paule, Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA), Université d'Auvergne - Clermont-Ferrand I (UdA), chirurgie plastique, reconstructrice et esthétique, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, Cancéropôle Lyon Auvergne Rhône-Alpes, Institut National de Recherche Agronomique (INRA). UMR Unité de Nutrition Humaine (1019)., laboratoire de recherche en Cardiovasculaire, Métabolisme, Diabétologie et Nutrition (CarMeN). Lyon, FRA., Université Joseph Fourier (Grenoble 1) (UJF). Grenoble, FRA., Centre de Recherche en Nutrition Humaine (CRNH). FRA., Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université, and Université d'Auvergne (Clermont Ferrand 1) (UdA)

Subjects
obésité, cancer du sein, Alimentation et Nutrition, femme menopausée, secrétion adipocytaire, Food and Nutrition, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition
Abstract

Cette année les assises sont en partenariat avec le Pôle de compétitivité Lyonbiopôle et le cluster Nutravita, des symposiums thématiques portant sur les interactions entre la recherche académique, l’innovation et l’industrie seront proposés; L’obésité est désormais reconnue comme un facteur de risque de développement de cancer du sein chez les femmes ménopausées, mais elle est également responsable d’une augmentation du risque métastatique, de récidives et du taux de mortalité. Nos travaux consistent à déterminer l’impact des sécrétions adipocytaires dans l’augmentation du risque de mortalité observée en situation d’obésité en nous focalisant sur le processus de résistance au traitement d’hormonothérapie par Tamoxifène (Tx) ainsi que sur le comportement des cellules souches cancéreuses (CSC) mammaires, dont le rôle dans la résistance aux traitements est désormais soulevé. Pour cela, i/ la prolifération des cellules cancéreuses mammaires (MCF-7) en présence de Tx et de milieux conditionnés provenant de la culture d’adipocytes matures (AM) issus de la différentiation in vitro de cellules souches adipocytaires prélevées sur des femmes normopondérées (AM20) ou obèses (AM30) a été évaluée. ii/ Un modèle original de culture 3D entre « MCF-7-fibroblastes-AM20 ou AM30 » a été mis au point en présence ou non de Tx. La mesure de l’expression de 42 gènes (impliqués dans l’angiogenèse, les voies cytokiniques et hormonales, la prolifération cellulaire) a été réalisée. iii/ Après avoir caractérisé et isolé les cellules souches cancéreuses (CSC), l’impact de traitements anti-cancéreux d’hormonothérapie ou de chimiothérapie et de deux adipokines a été évalué sur la prolifération des CSC mammaires MCF-7. Le secrétome pourrait intervenir dans la réponse à la thérapie puisqu’une diminution de l’efficacité du Tx est retrouvée en présence des milieux conditionnés d’AM30 dans nos différents modèles. Des profils d’expression des MCF7 différents ont pu être définis en fonction de leur exposition au Tx et/ou aux AM30. Concernant l’étude des CSC, les cellules isolées voient leur prolifération augmenter en présence des traitements anti-cancéreux mais également en présence de leptine et d’IL-6. Ainsi, le secrétome adipocytaire pourrait être directement impliqué dans l’augmentation de mortalité décrite en situation de surpoids et de cancer du sein en diminuant l’efficacité des traitements anticancéreux d’hormonothérapie tels que le tamoxifène. De plus, nos premiers travaux suggèrent que ces sécrétions pourraient aussi jouer un rôle dans le comportement des CSC, ce qui ouvre des perspectives préventives et/ou thérapeutiques intéressantes en termes de prise en charge des patientes en surcharge pondérale

Published
2015

23. Interactions between adipocyte secretions and breast cells: impact on tumor progression and resistance to therapy

Author

Bougaret, Laurianne, Delort, Laetitia, Billard, Hermine, Mojallal, Ali, Le Huede, Camille, Damour, Odile, VASSON, Marie-Paule, Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA), Service de chirurgie plastique reconstructrice et esthétique, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, CHU Clermont-Ferrand, Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Cancéropôle Lyon Auvergne Rhône-Alpes, and Federation of European Nutrition Societies (FENS).

Subjects
obésité, angiogenesis, therapy, adipokines, cancer du sein, Alimentation et Nutrition, femme menopausée, Food and Nutrition, [SDV.AEN]Life Sciences [q-bio]/Food and Nutrition, ComputingMilieux_MISCELLANEOUS
Abstract

National audience

Published
2015
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24. Les sécrétions adipocytaires induisent-elles une diminution de l’efficacité des traitements d’hormonothérapie en situation d’obésité ?

Author

Bougaret, Laurianne, Goiffon, Elise, Delort, Laetitia, Billard, Hermine, Metral, E., Mojallal, A., Damour, O., VASSON, Marie-Paule, Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université, Banque de tissus et de cellules, 3Service de chirurgie plastique, reconstructrice et esthétique, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre Jean Perrin - Centre Régional de Lutte contre le Cancer d'Auvergne, Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Société Francaise de Nutrition Clinique et Métabolisme (SFNEP). FRA., and Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA)

Subjects
[SDV]Life Sciences [q-bio]
Abstract

Introduction et but de l’étude L’obésité, facteur de risque de cancer du sein chez les femmes ménopausées, est associée à un pronostic défavorable avec survenue de récidives. Au cours de la chimiothérapie, les femmes obèses ou ayant une prise de poids >5% présentent une moindre réponse thérapeutique associée à un risque accru de rechutes et de mortalité. Ceci pourrait impliquer une interaction entre sécrétions adipocytaires et thérapie. Dans cette étude, l’objectif est de caractériser, in vitro, i/ l’impact d’une sécrétion adipocytaire spécifique, la leptine (LEP), sur l’efficacité de l’hormonothérapie : deux anti-œstrogènes (Tamoxifène (Tx) et Fulvestrant (Fv)) et un anti-aromatase (Létrozole (Lz)) ; ii/ l’influence du secrétât adipocytaire global de la femme mince et obèse sur la réponse au Tx. Matériel et méthodes Pour cela, i/ la prolifération des cellules mammaires néoplasiques (MCF-7) ou issues de dystrophie fibro-kystique (MCF10a) en présence de Tx, Fv ou Lz et de LEP (10-100-1000ng/mL) a été évaluée (fluorescence, rezasurine). ii/ Un modèle original (1) de culture tridimensionnelle entre « MCF-7-fibro-blastes-adipocytes matures issus de la différenciation de cellules souches adipocytaires de femmes minces (AM20) ou obèses (AM30) » a été utilisé en présence ou non de Tx. Des marquages immunohistologiques (marqueur de prolifération, Ki67), un suivi de variations d’expressions géniques et une analyse en composante principale (ACP) ont été réalisés dans ce modèle 3D. Résultats et Analyse statistique L’inhibition de prolifération des MCF-7 induite par le Tx et le Lz (−15 %, −12 % respectivement, p < 0,05, Tx ou Lz vs Contrôle (C)) n’est plus retrouvée en présence de LEP. À l’inverse, l’effet antiprolifératif du Fv (-20 %, p < 0,001, Fv vs C) n’est pas modifié par la LEP. La prolifération des MCF- 10a n’est pas modifiée par le Tx alors que le Fv et le Lz la diminuent (−8 %, p < 0,05 et −16 %, p < 0,001 respectivement, Fv ou Lz vs C). En culture 3D, le marquage Ki67 montre que l’effet antiprolifératif du Tx est amoindri en présence d’AM30. L’ACP, réalisée à partir d’un panel de 42 gènes (angiogenèse, cytokines/hormones, réponse aux traitements, prolifération cellulaire, stress oxydant) permet notamment de distinguer deux profils d’expression des MCF7 en fonction de leur exposition au Tx et/ou aux AM30. Ainsi, des gènes impliqués dans la réponse immune et dans la signalisation hormonale, tels que le TNFα, voient leur expression modifiée (TNFα : R =2,71, MCF-7/AM30 vs MCF-7/AM20, p < 0,01). Conclusion Ces résultats montrent que la LEP amoindrit l’efficacité de l’hormonothérapie (Tx et Lz), et dans une moindre proportion, celle du Fv. Le modèle 3D met en évidence que les sécrétions adipocytaires, notamment celles des AM30, contrecarrent l’effet du Tx. Cette étude permet de suggérer des liens étroits entre sécrétions adipocytaires et résistance à l’hormonothérapie chez les femmes en surpoids.

Published
2014
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25. Interaction entre cellules mammaires et adipocytaires en situation d’obésité : implication dans le cancer du sein

Author

Delort, Laetitia, Bougaret, Laurianne, Billard, Hermine, Mojallal, A., Damour, O., VASSON, Marie-Paule, Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA), Hôpital de la Croix-Rousse [CHU - HCL], Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Société Francaise de Nutrition Clinique et Métabolisme (SFNEP). FRA., and Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université

Subjects
[SDV.AEN]Life Sciences [q-bio]/Food and Nutrition
Abstract

International audience; Introduction et but de l’étude L’obésité est un facteur de risque d’apparition du cancer du sein chez les femmes ménopau-sées. Une influence du microenvironnement adipocytaire est aujourd’hui démontrée dans le développement de cette pathologie. Ainsi, l’objectif de notre étude est de caractériser, dans un contexte d’obésité, les interactions entre les cellules mammaires et leur microenvironnement adipeux et plus particulièrement les adipocytes matures (AM) et leurs progéniteurs (cellules souches adipocytaires [ASC] et préadipocytes [PA]). Matériel et méthodes Afin d’évaluer l’influence de l’obésité, des ASC extraites à partir de tissu adipeux sous-cutané abdominal de femmes minces et obèses (n =6) ont été mises à différencier en pré-adipocytes (PA20 et PA30) puis en adipocytes matures (AM20 et AM30) et des analyses par qRT-PCR effectuées. Ces deux types de cellules adipeuses ont ensuite été co-cultivées en présence de cellules mammaires cancéreuses (MCF-7 et MDA-MB-231) ou non cancéreuses (MCF10a) et leur prolifération a été mesurée (fluorescence à la résazurine, Fluoroskan Ascent FL®, n =6). Résultats et Analyse statistique Les résultats montrent que les AM provenant de femmes obèses comparés à ceux issus de femmes de poids normal expriment plus fortement la leptine (x8,6, p =0,03), son récepteur (x3,5, p =0,01), l’aromatase (x13,06, p =0,01) et le VEGF (x2, p =0,01) et plus faiblement l’adiponectine (x0,3, p =0,04), ce qui suggère que les AM gardent en mémoire leur environnement métabolique d’origine. Les capacités de prolifération des cellules cancéreuses mammaires MCF-7 positives pour le récepteur aux œstrogènes (RE +) sont augmentées lorsqu’elles sont co-cultivées en présence des PA30, AM20 et AM30 (+125%, p =0,03 ; +150%, p=0,01; +170%, p < 0,001 respectivement). Les cellules cancéreuses MDA-MB-231 (RE-) et non cancéreuses MCF10a semblent moins sensibles à la présence de cellules adipeuses. Le suivi des capacités de prolifération des cellules adipeuses a montré que la présence des cellules mammaires cancéreuses ou non étaient capables d’augmenter de façon considérable la prolifération des PA20 (+181 %, p=0,08 ; +149%, p=0,08 ; +164%, p =0,06) mais pas celle des PA30. La prolifération des AM20 n’est pas modifiée en présence de cellules mammaires alors que celle des AM30 parait augmentée en présence des cellules MCF-7 et MDA-MB-231 (+120%, p < 0,001 ; +113%, p =0,03), ceci pouvant s’expliquer par la présence résiduelle de cellules souches. Conclusion Ainsi, ces résultats montrent l’existence d’interactions réciproques entre cellules mammaires et adipeuses, se traduisant par des modifications du comportement des différents types cellulaires présents dans le microenvironnement et pouvant jouer un rôle clé dans la progression de la cancérogenèse mammaire.

Published
2014
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26. Dialogue entre adipocytes et cellules neoplasiques mammaires : Impact sur le processus d'angiogenèse tumorale et sur l'efficacité de l'hormonotherapie en situation d'obésité

Author

Bougaret, Laurianne, Delort, Laetitia, Billard, Hermine, Le Huédé, C., Mojallal, A., Damour, O., VASSON, Marie-Paule, Caldefie-Chezet, Florence, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université, Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Chirurgie plastique esthétique et reconstructrice, Hôpital Edouard Herriot, Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, European Center for Nutrition and Health (CENS). FRA., and Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA)

Subjects
[SDV.AEN]Life Sciences [q-bio]/Food and Nutrition, ComputingMilieux_MISCELLANEOUS
Abstract

National audience

Published
2014

27. Interspecies differences with in vitro and in vivo models of vascular tissue engineering

Author

Marlène Durand, Murielle Rémy, Jean François Ponsot, Laurence Bordenave, Jean-Claude Voegel, Patrick Menu, Marie Françoise Harmand, Xavier Berard, Bioingénierie tissulaire (BIOTIS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Bordeaux (UB), CHU Bordeaux [Bordeaux], Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de tectonique, mécanique de la lithosphère (LTML (UMR_7578)), IPG PARIS-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Banque de tissus et de cellules, Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Université Montesquieu - Bordeaux 4, Biomateriaux et Reparation Tissulaire, Université Bordeaux Segalen - Bordeaux 2-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de tectonique, mécanique de la lithosphère (LTML), and Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7)-IPG PARIS

Subjects
Materials science, Polymers, Cell, Biophysics, Bioengineering, 02 engineering and technology, 030204 cardiovascular system & hematology, Biomaterials, 03 medical and health sciences, 0302 clinical medicine, Tissue engineering, Blood vessel prosthesis, In vivo, medicine, Animals, Humans, Microscopy, Phase-Contrast, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Cells, Cultured, Cell Proliferation, Sheep, Tissue Engineering, Tissue Scaffolds, Cell growth, 021001 nanoscience & nanotechnology, In vitro, Blood Vessel Prosthesis, Endothelial stem cell, medicine.anatomical_structure, Mechanics of Materials, Cell culture, Ceramics and Composites, 0210 nano-technology, Shear Strength, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Biomedical engineering
Abstract

International audience; In arterial replacement there is a clear clinical need for a functional ă substitute possessing appropriate haemocompatible properties to be ă implanted as small diameter artery. Endothelial cell seeding constitutes ă an appreciated method to improve blood compatibility on the condition ă that cells firmly adhere to the support. Along this way, an innovative ă technique based on multilayered polyelectrolyte films (PEM) as cell ă adhesive substrate was previously validated in vitro and in vivo in a ă small-animal model. In this study, we extended the work on a larger ă animal (sheep) to validate furthermore the paradigm of PEM ă functionalization for vascular substitutes. We tested in vitro: the ă efficiency of PEM to induce endothelial progenitor differentiation in ă sheep endothelial cells; the ability of PEM to sustain cell ă proliferation and allow resistance to shear stress; the fate of ă PEM-coated de-endothelialized human saphenous veins under flow ă conditions, a prerequisite step before in vivo experiments. Despite in ă vitro differences we were encouraged by testing in vivo PEM-coated ă prosthesis as carotid replacement in sheep, but without success. In ă order to explain the implantation failure, an in vitro ă haemocompatibility evaluation was performed that highlighted ă interspecies differences able to explain, at least in part, the graft ă failure obtained. (C) 2013 Elsevier Ltd. All rights reserved.

Published
2013
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28. Reciprocal interactions between breast tumor and its adipose microenvironment based on a 3D adipose equivalent model

Author

Odile Damour, Florence Caldefie-Chezet, Alice Dubouloz, Virginie Dubois, Marie-Paule Vasson, Charlotte Lequeux, Hermine Billard, Laetitia Delort, Ali Mojallal, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université, Université d'Auvergne - Clermont-Ferrand I (UdA), Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL), Banque de tissus et de cellules, Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Ligue Nationale contre le Cancer, French Ministry of Research and Technology fellowship, and Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université-Institut National de la Recherche Agronomique (INRA)

Subjects
Keratinocytes, Anatomy and Physiology, Transcription, Genetic, Cellular differentiation, [SDV]Life Sciences [q-bio], Adipose tissue, lcsh:Medicine, 0302 clinical medicine, Molecular Cell Biology, Breast Tumors, Tumor Microenvironment, Child, lcsh:Science, 0303 health sciences, Mammary tumor, Multidisciplinary, Cancer Risk Factors, Cell Differentiation, Receptors, Adipokine, 3. Good health, Gene Expression Regulation, Neoplastic, Somatic Cells, Oncology, Adipose Tissue, 030220 oncology & carcinogenesis, Medicine, Synthetic Biology, Cellular Types, Research Article, medicine.medical_specialty, Cell Physiology, Clinical Research Design, Adipokine, Breast Neoplasms, Biology, Models, Biological, 03 medical and health sciences, Breast cancer, Adipokines, Internal medicine, Cell Line, Tumor, medicine, Humans, Obesity, 030304 developmental biology, Nutrition, Tumor microenvironment, Adiponectin, lcsh:R, Modeling, Cancer, Cancers and Neoplasms, Epithelial Cells, medicine.disease, Endocrinology, Cancer research, lcsh:Q
Abstract

International audience; Breast cancer has become the most common cancer among women in industrialized countries. Obesity is well established as a risk factor, in particular owing to the attendant secretion of the entities called adipokines; there is growing evidence for a role of cells and factors present in the mammary tumor microenvironment such as fibroblasts, preadipocytes, adipocytes and their secretions. To study how the microenvironment influences breast cancer growth, we developed a novel tridimensional adipose model epithelialized with normal human keratinocytes or with breast cancer cell lines. These mimicked a breast tumor in contact with an adipose microenvironment and allowed monitoring of the interactions between the cells. Leptin and adiponectin, two major adipokines, and their respective receptors, ObRt and AdipoR1, were expressed in the model, but not the second adiponectin receptor, AdipoR2. The differentiation of preadipocytes into adipocytes was greater when they were in contact with the breast cancer cell lines. The contact of breast cancer cell lines with the microenvironment completely modified their transcriptional programs by increasing the expression of genes involved in cell proliferation (cyclinD1, MAPK), angiogenesis (MMP9, VEGF) and hormonal pathways (ESR1, IL6). This tridimensional adipose model provides new insights into the interactions between breast cancer cells and their adipose microenvironment, and provides a tool to develop new drugs for the treatment of both cancer and obesity.

Published
2013
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29. Deciphering influence of donor age on adipose-derived stem cells: in vitro paracrine function and angiogenic potential.

Author

Trotzier C, Bellanger C, Abdessadeq H, Delannoy P, Mojallal A, and Auxenfans C

Subjects
Humans, Adult, Middle Aged, Aged, Female, Male, Stem Cells metabolism, Stem Cells cytology, Cells, Cultured, Mesenchymal Stem Cells metabolism, Mesenchymal Stem Cells cytology, Culture Media, Conditioned pharmacology, Adipocytes metabolism, Adipocytes cytology, Age Factors, Young Adult, Aging physiology, Tissue Donors, Adipose Tissue cytology, Adipose Tissue metabolism, Neovascularization, Physiologic, Paracrine Communication, Cell Differentiation, Endothelial Cells metabolism, Endothelial Cells cytology, Coculture Techniques
Abstract

Background: As fat grafting is commonly used as a filler, Adipose-derived stem/stromal cells (ASC) have been reported to be key player in retention rate. Paracrine and differentiation potential of those cells confer them strong pro-angiogenic capacities. However, a full characterization of the influence of aging on ASC has not been reported yet. Here we've investigated the effect of age on paracrine function, stemness and angiogenic potential of ASC., Methods: ASC were extracted from young and old adult donors. We assessed stromal vascular fraction cell populations repartition, ASC stemness potential, capability to differentiate into mesenchymal lineages as well as their secretome. Angiogenic potential was assessed using a sprouting assay, an indirect co-culture of ASC and dermal microvascular endothelial cells (EC). Total vascular sprout length was measured, and co-culture soluble factors were quantified. Pro-angiogenic factors alone or in combination as well as ASC-conditioned medium (CM) were added to EC to assess sprouting induction., Results: Decrease of endothelial cells yield and percentage is observed in cells extracted from adipose tissue of older patients, whereas ASC percentage increased with age. Clonogenic potential of ASC is stable with age. ASC can differentiate into adipocytes, chondrocytes and osteoblasts, and aging does not alter this potential. Among the 25 analytes quantified, high levels of pro-angiogenic factors were found, but none is significantly modulated with age. ASC induce a significantly longer vascular sprouts compared to fibroblasts, and no difference was found between young and old ASC donors on that parameter. Higher concentrations of FGF-2, G-CSF, HGF and IL-8, and lower concentrations of VEGF-C were quantified in EC/ASC co-cultures compared to EC/fibroblasts co-cultures. EC/ASC from young donors secrete higher levels of VEGF-A compared to old ones. Neither soluble factor nor CM without cells are able to induce organized sprouts, highlighting the requirement of cell communication for sprouting. CM produced by ASC supporting development of long vascular sprouts promote sprouting in co-cultures that establish shorter sprouts., Conclusion: Our results show cells from young and old donors exhibit no difference in all assessed parameters, suggesting all patients could be included in clinical applications. We emphasized the leading role of ASC in angiogenesis, without impairment with age, where secretome is a key but not sufficient actor., Competing Interests: Declarations Competing interests The authors declare no competing interests. Ethics approval and consent to participate Written informed consent was obtained from the patient and surgical residue was collected according to French regulation and declared to research ministry., (© 2024. The Author(s).)

Published
2024
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30. ESOT Roadmap for Advanced Therapy Medicinal Products in Transplantation: Navigating Regulatory Challenges to Enhance Access and Care.

Author

Berishvili E, Piemonti L, de Koning EJP, Lindstedt S, Scholz H, Scott WE, Auxenfans C, Johnson P, Martin DE, Gunther P, Mey D, Potena L, and Thaunat O

Subjects
Humans, Europe, Cell- and Tissue-Based Therapy, Genetic Therapy legislation & jurisprudence, Health Services Accessibility legislation & jurisprudence, Organ Transplantation legislation & jurisprudence, Tissue Engineering legislation & jurisprudence, Tissue Engineering methods
Abstract

The field of organ transplantation is experiencing a transformative shift with the rise of Advanced Therapy Medicinal Products (ATMPs), which include gene therapies, somatic cell therapies, and tissue-engineered products. These therapies offer new, potentially curative treatments for longstanding medical challenges, impacting numerous patients. However, their adoption is hindered by complex regulatory frameworks, high production costs, and inconsistent access across Europe. The ESOT ATMP Task Force's position paper analyzes these challenges from research to clinical application, advocating for a coordinated strategy to position Europe as a leader in ATMP development. It proposes specific actions such as streamlining regulatory pathways to accelerate approvals, boosting funding for ATMP research, and creating specialized facilities for development and implementation. The paper also highlights the critical roles of patient engagement and real-world evidence in optimizing clinical and regulatory practices., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Berishvili, Piemonti, de Koning, Lindstedt, Scholz, Scott, Auxenfans, Johnson, Martin, Gunther, Mey, Potena and Thaunat.)

Published
2024
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31. The obese inflammatory microenvironment may promote breast DCIS progression.

Author

Habanjar O, Nehme R, Goncalves-Mendes N, Cueff G, Blavignac C, Aoun J, Decombat C, Auxenfans C, Diab-Assaf M, Caldefie-Chézet F, and Delort L

Subjects
Humans, Female, Inflammation pathology, Inflammation metabolism, Adipocytes metabolism, Adipocytes pathology, Adipose Tissue pathology, Adipose Tissue metabolism, Cell Line, Tumor, Obesity metabolism, Obesity pathology, Breast Neoplasms pathology, Breast Neoplasms immunology, Breast Neoplasms metabolism, Tumor Microenvironment immunology, Carcinoma, Intraductal, Noninfiltrating pathology, Carcinoma, Intraductal, Noninfiltrating metabolism, Carcinoma, Intraductal, Noninfiltrating immunology, Disease Progression, Coculture Techniques, Macrophages immunology, Macrophages metabolism
Abstract

Introduction: Ductal carcinoma in situ (DCIS), characterized by a proliferation of neoplastic cells confined within the mammary ducts, is distinctly isolated from the surrounding stroma by an almost uninterrupted layer of myoepithelial cells (MECs) and by the basement membrane. Heightened interactions within the adipose microenvironment, particularly in obese patients, may play a key role in the transition from DCIS to invasive ductal carcinoma (IDC), which is attracting growing interest in scientific research. Adipose tissue undergoes metabolic changes in obesity, impacting adipokine secretion and promoting chronic inflammation. This study aimed to assess the interactions between DCIS, including in situ cancer cells and MECs, and the various components of its inflammatory adipose microenvironment (adipocytes and macrophages)., Methods: To this end, a 3D co-culture model was developed using bicellular bi-fluorescent DCIS-like tumoroids, adipose cells, and macrophages to investigate the influence of the inflammatory adipose microenvironment on DCIS progression., Results: The 3D co-culture model demonstrated an inhibition of the expression of genes involved in apoptosis ( BAX, BAG1, BCL2, CASP3, CASP8 , and CASP9 ), and an increase in genes related to cell survival ( TP53 , JUN , and TGFB1 ), inflammation ( TNF-α, PTGS2, IL-6R ), invasion and metastasis (TIMP1 and MMP-9) in cancer cells of the tumoroids under inflammatory conditions versus a non-inflammatory microenvironment. On the contrary, it confirmed the compromised functionality of MECs, resulting in the loss of their protective effects against cancer cells. Adipocytes from obese women showed a significant increase in the expression of all studied myofibroblast-associated genes (myoCAFs), such as FAP and α-SMA . In contrast, adipocytes from normal-weight women expressed markers of inflammatory fibroblast phenotypes (iCAF) characterized by a significant increase in the expression of LIF and inflammatory cytokines such as TNF-α, IL-1β, IL-8 , and CXCL-10 . These changes also influenced macrophage polarization, leading to a pro-inflammatory M1 phenotype. In contrast, myoCAF-associated adipocytes, and the cancer-promoting microenvironment polarized macrophages towards an M2 phenotype, characterized by high CD163 receptor expression and IL-10 and TGF-β secretion., Discussion: Reciprocal interactions between the tumoroid and its microenvironment, particularly in obesity, led to transcriptomic changes in adipocytes and macrophages, may participate in breast cancer progression while disrupting the integrity of the MEC layer. These results underlined the importance of adipose tissue in cancer progression., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Habanjar, Nehme, Goncalves-Mendes, Cueff, Blavignac, Aoun, Decombat, Auxenfans, Diab-Assaf, Caldefie-Chézet and Delort.)

Published
2024
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32. Aspalathus linearis (Rooibos) Targets Adipocytes and Obesity-Associated Inflammation.

Author

Nehme R, Chervet A, Decombat C, Longechamp L, Rossary A, Boutin R, Rousset A, Senejoux F, Vachias C, Auxenfans C, Fraisse D, Guyon JB, Filaire E, Berthon JY, Diab-Assaf M, Delort L, and Caldefie-Chezet F

Subjects
Humans, Leptin, Plant Extracts pharmacology, Adiponectin, Obesity complications, Inflammation, Adipocytes, Cytokines, Tea, Aspalathus
Abstract

Excess weight and obesity are the fifth leading cause of death globally, and sustained efforts from health professionals and researchers are required to mitigate this pandemic-scale problem. Polyphenols and flavonoids found in Aspalathus linearis -a plant widely consumed as Rooibos tea-are increasingly being investigated for their positive effects on various health issues including inflammation. The aim of our study was to examine the effect of Rooibos extract on obesity and the associated low-grade chronic inflammatory state by testing antioxidant activity, cytokine secretions, macrophage polarization and the differentiation of human adipocytes through the development of adipospheroids. Rooibos extract significantly decreased ROS production and the secretion of pro-inflammatory cytokines (IFN-γ, IL-12, IL-2 and IL-17a) in human leukocytes. Additionally, Rooibos extract down-regulated LPS-induced macrophage M1 polarization, shown by a significant decrease in the expression of pro-inflammatory cytokines: TNFα , IL-8 , IL-6 , IL-1β and CXCL10 . In addition, Rooibos inhibited intracellular lipid accumulation and reduced adipogenesis by decreasing the expression of PPARγ , Ap2 and HSL in adipospheroids. A significant decrease in leptin expression was noted and this, more interestingly, was accompanied by a significant increase in adiponectin expression. Using a co-culture system between macrophages and adipocytes, Rooibos extract significantly decreased the expression of all studied pro-inflammatory cytokines and particularly leptin , and increased adiponectin expression. Thus, adding Rooibos tea to the daily diet is likely to prevent the development of obesity associated with chronic low-level inflammation.

Published
2023
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33. Fat Graft Retention: Adipose Tissue, Adipose-Derived Stem Cells, and Aging.

Author

Trotzier C, Sequeira I, Auxenfans C, and Mojallal AA

Subjects
Humans, Aging, Stem Cells, Fibrosis, Graft Survival, Adipose Tissue transplantation, Adipocytes transplantation
Abstract

Summary: Over the past 30 years, there has been a dramatic increase in the use of autologous fat grafting for soft-tissue augmentation and to improve facial skin quality. Several studies have highlighted the impact of aging on adipose tissue, leading to a decrease of adipose tissue volume and preadipocyte proliferation and increase of fibrosis. Recently, there has been a rising interest in adipose tissue components, including adipose-derived stem/stromal cells (ASCs) because of their regenerative potential, including inflammation, fibrosis, and vascularization modulation. Because of their differentiation potential and paracrine function, ASCs have been largely used for fat grafting procedures, as they are described to be a key component in fat graft survival. However, many parameters as surgical procedures or adipose tissue biology could change clinical outcomes. Variation on fat grafting methods have led to numerous inconsistent clinical outcomes. Donor-to-donor variation could also be imputed to ASCs, tissue inflammatory state, or tissue origin. In this review, the authors aim to analyze (1) the parameters involved in graft survival, and (2) the effect of aging on adipose tissue components, especially ASCs, that could lead to a decrease of skin regeneration and fat graft retention., Clinical Relevance Statement: This review aims to enlighten surgeons about known parameters that could play a role in fat graft survival. ASCs and their potential mechanism of action in regenerative medicine are more specifically described., (Copyright © 2022 by the American Society of Plastic Surgeons.)

Published
2023
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34. Spen modulates lipid droplet content in adult Drosophila glial cells and protects against paraquat toxicity.

Author

Girard V, Goubard V, Querenet M, Seugnet L, Pays L, Nataf S, Dufourd E, Cluet D, Mollereau B, and Davoust N

Subjects
Animals, Drosophila Proteins genetics, Drosophila melanogaster drug effects, Drosophila melanogaster metabolism, Herbicides toxicity, Homeodomain Proteins genetics, Male, Neuroglia drug effects, Neuroglia metabolism, Parkinson Disease etiology, Parkinson Disease metabolism, Parkinson Disease pathology, RNA-Binding Proteins genetics, Drosophila Proteins metabolism, Drosophila melanogaster growth & development, Gene Expression Regulation, Developmental, Homeodomain Proteins metabolism, Lipid Droplets chemistry, Neuroglia cytology, Paraquat toxicity, Parkinson Disease prevention & control, RNA-Binding Proteins metabolism
Abstract

Glial cells are early sensors of neuronal injury and can store lipids in lipid droplets under oxidative stress conditions. Here, we investigated the functions of the RNA-binding protein, SPEN/SHARP, in the context of Parkinson's disease (PD). Using a data-mining approach, we found that SPEN/SHARP is one of many astrocyte-expressed genes that are significantly differentially expressed in the substantia nigra of PD patients compared with control subjects. Interestingly, the differentially expressed genes are enriched in lipid metabolism-associated genes. In a Drosophila model of PD, we observed that flies carrying a loss-of-function allele of the ortholog split-ends (spen) or with glial cell-specific, but not neuronal-specific, spen knockdown were more sensitive to paraquat intoxication, indicating a protective role for Spen in glial cells. We also found that Spen is a positive regulator of Notch signaling in adult Drosophila glial cells. Moreover, Spen was required to limit abnormal accumulation of lipid droplets in glial cells in a manner independent of its regulation of Notch signaling. Taken together, our results demonstrate that Spen regulates lipid metabolism and storage in glial cells and contributes to glial cell-mediated neuroprotection.

Published
2020
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35. Encapsulation of Adipose-Derived Mesenchymal Stem Cells in Calcium Alginate Maintains Clonogenicity and Enhances their Secretory Profile.

Author

Capin L, Abbassi N, Lachat M, Calteau M, Barratier C, Mojallal A, Bourgeois S, and Auxenfans C

Subjects
Adult, Alginates metabolism, Cell Differentiation, Cells, Cultured, Colony-Forming Units Assay, Humans, Middle Aged, Pluripotent Stem Cells cytology, Alginates chemistry, Cell Proliferation, Cytokines metabolism, Mesenchymal Stem Cells cytology, Mesenchymal Stem Cells metabolism, Pluripotent Stem Cells metabolism
Abstract

Adipose-derived mesenchymal stem cells (ASCs) are well known for their secretory potential, which confers them useful properties in cell therapy. Nevertheless, this therapeutic potential is reduced after transplantation due to their short survival in the human body and their migration property. This study proposes a method to protect cells during and after injection by encapsulation in microparticles of calcium alginate. Besides, the consequences of encapsulation on ASC proliferation, pluripotential, and secretome were studied. Spherical particles with a mean diameter of 500 µm could be obtained in a reproducible manner with a viability of 70% after 16 days in vitro. Moreover, encapsulation did not alter the proliferative properties of ASCs upon return to culture nor their differentiation potential in adipocytes, chondrocytes, and osteocytes. Concerning their secretome, encapsulated ASCs consistently produced greater amounts of interleukin-6 (IL-6), interleukin-8 (IL-8), and vascular endothelial growth factor (VEGF) compared to monolayer cultures. Encapsulation therefore appears to enrich the secretome with transforming growth factor β1 (TGF-β1) and macrophage inflammatory protein-1β (MIP-1β) not detectable in monolayer cultures. Alginate microparticles seem sufficiently porous to allow diffusion of the cytokines of interest. With all these cytokines playing an important role in wound healing, it appears relevant to investigate the impact of using encapsulated ASCs on the wound healing process.

Published
2020
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36. Common Neurodegeneration-Associated Proteins Are Physiologically Expressed by Human B Lymphocytes and Are Interconnected via the Inflammation/Autophagy-Related Proteins TRAF6 and SQSTM1.

Author

Nataf S, Guillen M, and Pays L

Subjects
Humans, Systems Biology, Autophagy immunology, B-Lymphocytes immunology, Inflammation immunology, Intracellular Signaling Peptides and Proteins immunology, Neurodegenerative Diseases immunology, Sequestosome-1 Protein immunology
Abstract

There is circumstantial evidence that, under neurodegenerative conditions, peptides deriving from aggregated or misfolded specific proteins elicit adaptive immune responses. On another hand, several genes involved in familial forms of neurodegenerative diseases exert key innate immune functions. However, whether or not such observations are causally linked remains unknown. To start addressing this issue, we followed a systems biology strategy based on the mining of large proteomics and immunopeptidomics databases. First, we retrieved the expression patterns of common neurodegeneration-associated proteins in two professional antigen-presenting cells, namely B lymphocytes and dendritic cells. Surprisingly, we found that under physiological conditions, numerous neurodegeneration-associated proteins are abundantly expressed by human B lymphocytes. A survey of the human proteome allowed us to map a unique protein-protein interaction network linking common neurodegeneration-associated proteins and their first shell interactors in human B lymphocytes. Interestingly, network connectivity analysis identified two major hubs that both relate with inflammation and autophagy, namely TRAF6 (TNF Receptor Associated Factor 6) and SQSTM1 (Sequestosome-1). Moreover, the mapped network in B lymphocytes comprised two additional hub proteins involved in both inflammation and autoimmunity: HSPA8 (Heat Shock Protein Family A Member 8 also known as HSC70) and HSP90AA1 (Heat Shock Protein 90 Alpha Family Class A Member 1). Based on these results, we then explored the Immune Epitope Database "IEDB-AR" and actually found that a large share of neurodegeneration-associated proteins were previously reported to provide endogenous MHC class II-binding peptides in human B lymphocytes. Of note, peptides deriving from amyloid beta A4 protein, sequestosome-1 or profilin-1 were reported to bind multiple allele-specific MHC class II molecules. In contrast, peptides deriving from microtubule-associated protein tau, presenilin 2 and serine/threonine-protein kinase TBK1 were exclusively reported to bind MHC molecules encoded by the HLA-DRB1 1501 allele, a recently-identified susceptibility gene for late onset Alzheimer's disease. Finally, we observed that the whole list of proteins reported to provide endogenous MHC class II-binding peptides in human B lymphocytes is specifically enriched in neurodegeneration-associated proteins. Overall, our work indicates that immunization against neurodegeneration-associated proteins might be a physiological process which is shaped, at least in part, by B lymphocytes., (Copyright © 2019 Nataf, Guillen and Pays.)

Published
2019
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37. The Promoter Regions of Intellectual Disability-Associated Genes Are Uniquely Enriched in LTR Sequences of the MER41 Primate-Specific Endogenous Retrovirus: An Evolutionary Connection Between Immunity and Cognition.

Author

Nataf S, Uriagereka J, and Benitez-Burraco A

Abstract

Social behavior and neuronal connectivity in rodents have been shown to be shaped by the prototypical T lymphocyte-derived pro-inflammatory cytokine Interferon-gamma (IFNγ). It has also been demonstrated that STAT1 (Signal Transducer And Activator Of Transcription 1), a transcription factor (TF) crucially involved in the IFNγ pathway, binds consensus sequences that, in humans, are located with a high frequency in the LTRs (Long Terminal Repeats) of the MER41 family of primate-specific HERVs (Human Endogenous Retroviruses). However, the putative role of an IFNγ/STAT1/MER41 pathway in human cognition and/or behavior is still poorly documented. Here, we present evidence that the promoter regions of intellectual disability-associated genes are uniquely enriched in LTR sequences of the MER41 HERVs. This observation is specific to MER41 among more than 130 HERVs examined. Moreover, we have not found such a significant enrichment in the promoter regions of genes that associate with autism spectrum disorder (ASD) or schizophrenia. Interestingly, ID-associated genes exhibit promoter-localized MER41 LTRs that harbor TF binding sites (TFBSs) for not only STAT1 but also other immune TFs such as, in particular, NFKB1 (Nuclear Factor Kappa B Subunit 1) and STAT3 (Signal Transducer And Activator Of Transcription 3). Moreover, IL-6 (Interleukin 6) rather than IFNγ, is identified as the main candidate cytokine regulating such an immune/MER41/cognition pathway. Of note, differences between humans and chimpanzees are observed regarding the insertion sites of MER41 LTRs in the promoter regions of ID-associated genes. Finally, a survey of the human proteome has allowed us to map a protein-protein network which links the identified immune/MER41/cognition pathway to FOXP2 (Forkhead Box P2), a key TF involved in the emergence of human speech. Our work suggests that together with the evolution of immune genes, the stepped self-domestication of MER41 in the genomes of primates could have contributed to cognitive evolution. We further propose that non-inherited forms of ID might result from the untimely or quantitatively inappropriate expression of immune signals, notably IL-6, that putatively regulate cognition-associated genes via promoter-localized MER41 LTRs.

Published
2019
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38. Development and characterization of Lyophilized Transparized Decellularized stroma as a replacement for living cornea in deep anterior lamellar keratoplasty.

Author

Rovere MR, Ouilhon C, Salmon D, Haftek M, Damour O, and Auxenfans C

Subjects
Animals, Corneal Stroma ultrastructure, Histocompatibility Antigens metabolism, Humans, Models, Biological, Swine, Thermogravimetry, Corneal Stroma cytology, Corneal Transplantation methods, Freeze Drying methods, Tissue Engineering methods
Abstract

Corneal disease is the second cause of blindness in developing countries, where the number of corneal grafts needed by far exceeds the number available. In industrialized countries, although corneas are generally available for keratoplasty, onto inflamed and vascularized host beds they are often rejected despite immune-suppression. A non-immunogenic, transparent, cytocompatible stroma is therefore required, which can be lyophilized for long-term conservation. Decellularization methods were tested on porcine corneal stromas before validation on human corneas. Decellularization and lyophilization led to opacification of the stroma, which could be reversed by soaking in 100% glycerol. Cell-depleted transparized stromas were then lyophilized (LTDC) to allow their long-term conservation and water content was measured. The ultrastructure of LTDC corneas was examined by transmission electron microscopy (TEM). Histocompatibility antigens were undetectable on LTDC stromas by antibody staining. Finally, cytocompatibility of LTDC stromas was demonstrated on an ex vivo model of anterior lamellar keratoplasty. Differential staining was used to monitor colonization of LTDC stromas by cells from the receiving cornea. Only SDS-based decellularization produced acellular porcine stromas. The lowest SDS concentration tested (0.1%) was validated on human corneas. Unlike lyophilized corneas, LTDC stromas without residual water, express no histocompatibility markers, although TEM revealed the presence of cellular debris in an ultrastructural arrangement of collagen fibers very close to that of native corneas. This structure is compatible with colonization by cells from the receiver cornea in an ex vivo lamellar graft model. Our procedure produced non-immunogenic, transparent stromas with conserved ultrastructure compatible with long-term conservation.

Published
2019
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39. Paclitaxel Treatment and Proprotein Convertase 1/3 (PC1/3) Knockdown in Macrophages is a Promising Antiglioma Strategy as Revealed by Proteomics and Cytotoxicity Studies.

Author

Duhamel M, Rose M, Rodet F, Murgoci AN, Zografidou L, Régnier-Vigouroux A, Vanden Abeele F, Kobeissy F, Nataf S, Pays L, Wisztorski M, Cizkova D, Fournier I, and Salzet M

Subjects
Animals, Brain Neoplasms metabolism, Cell Line, Cell Survival drug effects, Coculture Techniques, Cytokines metabolism, Glioma metabolism, Macrophages drug effects, Macrophages metabolism, Proteomics, Rats, Antineoplastic Agents, Phytogenic pharmacology, Brain Neoplasms therapy, Glioma therapy, Paclitaxel pharmacology, Proprotein Convertase 1 genetics
Abstract

High grade gliomas are the most common brain tumors in adult. These tumors are characterized by a high infiltration in microglial cells and macrophages. The immunosuppressive tumor environment is known to orient immune cells toward a pro-tumoral and anti-inflammatory phenotype. Therefore, the current challenge for cancer therapy is to find a way to reorient macrophages toward an antitumoral phenotype. Previously, we demonstrated that macrophages secreted antitumoral factors when they were invalidated for the proprotein converstase 1/3 (PC1/3) and treated with LPS. However, achieving an activation of macrophages via LPS/TLR4/Myd88-dependent pathway appears yet unfeasible in cancer patients. On the contrary, the antitumor drug Paclitaxel is also known to activate the TLR4 MyD88-dependent signaling pathway and mimics LPS action. Therefore, we evaluated if PC1/3 knock-down (KD) macrophages could be activated by Paclitaxel and efficient against glioma. We report here that such a treatment of PC1/3 KD macrophages drove to the overexpression of proteins mainly involved in cytoskeleton rearrangement. In support of this finding, we found that these cells exhibited a Ca 2+ increase after Paclitaxel treatment. This is indicative of a possible depolymerization of microtubules and may therefore reflect an activation of inflammatory pathways in macrophages. In such a way, we found that PC1/3 KD macrophages displayed a repression of the anti-inflammatory pathway STAT3 and secreted more pro-inflammatory cytokines. Extracellular vesicles isolated from these PC1/3 KD cells inhibited glioma growth. Finally, the supernatant collected from the coculture between glioma cells and PC1/3 KD macrophages contained more antitumoral factors. These findings unravel the potential value of a new therapeutic strategy combining Paclitaxel and PC1/3 inhibition to switch macrophages toward an antitumoral immunophenotype., (© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.)

Published
2018
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40. Adipocyte/breast cancer cell crosstalk in obesity interferes with the anti-proliferative efficacy of tamoxifen.

Author

Bougaret L, Delort L, Billard H, Le Huede C, Boby C, De la Foye A, Rossary A, Mojallal A, Damour O, Auxenfans C, Vasson MP, and Caldefie-Chezet F

Subjects
Cell Line, Female, Gene Expression, Humans, Leptin metabolism, MCF-7 Cells, Tumor Necrosis Factor-alpha metabolism, Adipocytes pathology, Antineoplastic Agents, Hormonal pharmacology, Breast Neoplasms pathology, Cell Proliferation drug effects, Obesity pathology, Tamoxifen pharmacology
Abstract

Background: Obesity is a well-known risk factor of breast cancer in post-menopausal women that also correlates with a diminished therapeutic response. The influence of adipocytes and their secretome, i.e. adipokines, on the efficacy of hormone therapy has yet to be elucidated., Methods: We investigated, ex vivo, whether mature adipocytes, differentiated from adipose stem cells of normal-weight (MA20) or obese (MA30) women, and their secretions, were able to counteract the effects of tamoxifen (Tx) which is known to decrease neoplastic cell proliferation., Results: In a tridimensional model and in a model of co-culture, the anti-proliferative effect of Tx on MCF-7 cancer cells was counteracted by MA30. These two models highlighted two different specific gene expression profiles for genes encoding cytokines or involved in angiogenesis based on the adipocyte microenvironment and the treatment. Thus it notably showed altered expression of genes such as TNFα that correlated with IL-6. In addition, leptin, IL-6 and TNFα, at concentrations reflecting plasma concentrations in obese patients, decreased the anti-proliferative efficacy of 4-hydroxytamoxifen (a major active metabolite of Tx)., Conclusions: These findings bring insights on adipocytes and mammary cancer cell interactions in Tx therapy, particularly in overweight/obese people. Indeed, patient' adipokine status would give valuable information for developing individual strategies and avoid resistance to treatment.

Published
2018
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41. A Unique TGFB1-Driven Genomic Program Links Astrocytosis, Low-Grade Inflammation and Partial Demyelination in Spinal Cord Periplaques from Progressive Multiple Sclerosis Patients.

Author

Nataf S, Barritault M, and Pays L

Subjects
Astrocytes metabolism, Cell Cycle Proteins metabolism, Computational Biology, Connexin 43 metabolism, Glial Fibrillary Acidic Protein metabolism, Humans, Interleukin-12 metabolism, Interleukin-12 Subunit p35 metabolism, Interleukin-17 metabolism, Interleukin-33 metabolism, Intracellular Signaling Peptides and Proteins metabolism, Lymphokines metabolism, Myelin Sheath metabolism, Oligodendroglia metabolism, Platelet-Derived Growth Factor metabolism, Inflammation metabolism, Multiple Sclerosis metabolism, Spinal Cord metabolism, Transforming Growth Factor beta1 metabolism
Abstract

We previously reported that, in multiple sclerosis (MS) patients with a progressive form of the disease, spinal cord periplaques extend distance away from plaque borders and are characterized by the co-occurrence of partial demyelination, astrocytosis and low-grade inflammation. However, transcriptomic analyses did not allow providing a comprehensive view of molecular events in astrocytes vs. oligodendrocytes. Here, we re-assessed our transcriptomic data and performed co-expression analyses to characterize astrocyte vs. oligodendrocyte molecular signatures in periplaques. We identified an astrocytosis-related co-expression module whose central hub was the astrocyte gene C x 43 / GJA1 (connexin-43, also named gap junction protein α-1) . Such a module comprised GFAP (glial fibrillary acidic protein) and a unique set of transcripts forming a TGFB / SMAD1 / SMAD2 (transforming growth factor β/SMAD family member 1/SMAD family member 2) genomic signature. Partial demyelination was characterized by a co-expression network whose central hub was the oligodendrocyte gene NDRG1 ( N -myc downstream regulated 1), a gene previously shown to be specifically silenced in the normal-appearing white matter (NAWM) of MS patients. Surprisingly, besides myelin genes, the NDRG1 co-expression module comprised a highly significant number of translation/elongation-related genes. To identify a putative cause of NDRG1 downregulation in periplaques, we then sought to identify the cytokine/chemokine genes whose mRNA levels inversely correlated with those of NDRG1 . Following this approach, we found five candidate immune-related genes whose upregulation associated with NDRG1 downregulation: TGFB1 ( transforming growth factor β 1), PDGFC (platelet derived growth factor C), IL17D (interleukin 17D), IL 33 (interleukin 33), and IL 12A (interleukin 12A). From these results, we propose that, in the spinal cord periplaques of progressive MS patients, TGFB1 may limit acute inflammation but concurrently induce astrocytosis and an alteration of the translation/elongation of myelin genes in oligodendrocytes., Competing Interests: The authors declare no conflict of interest.

Published
2017
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42. Skin allograft for severe chronic GvHD.

Author

Crocchiolo R, Dubois V, Nicolini FE, Sobh M, Ducastelle-Lepretre S, Labussiere H, Lebras L, Auxenfans C, Mojallal A, Damour O, Braye F, and Michallet M

Subjects
Adult, Allografts, Female, Humans, Graft vs Host Disease pathology, Graft vs Host Disease surgery, Hematopoietic Stem Cell Transplantation, Leukemia, Myeloid, Acute pathology, Leukemia, Myeloid, Acute therapy, Skin Diseases pathology, Skin Diseases surgery, Skin Transplantation
Published
2017
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43. Supernatants of Adipocytes From Obese Versus Normal Weight Women and Breast Cancer Cells: In Vitro Impact on Angiogenesis.

Author

Bougaret L, Delort L, Billard H, Lequeux C, Goncalves-Mendes N, Mojallal A, Damour O, Vasson MP, and Caldefie-Chezet F

Subjects
Adipocytes drug effects, Adipocytes metabolism, Adipose Tissue pathology, Antibodies, Neutralizing pharmacology, Body Mass Index, Body Weight drug effects, Cell Differentiation drug effects, Cell Movement drug effects, Cell Proliferation drug effects, Culture Media, Conditioned pharmacology, Female, Gene Expression Regulation drug effects, Human Umbilical Vein Endothelial Cells drug effects, Human Umbilical Vein Endothelial Cells metabolism, Humans, MCF-7 Cells, Receptors, Leptin genetics, Receptors, Leptin metabolism, Staining and Labeling, Subcellular Fractions drug effects, Subcellular Fractions metabolism, Vascular Endothelial Growth Factor Receptor-2 genetics, Vascular Endothelial Growth Factor Receptor-2 metabolism, Adipocytes pathology, Breast Neoplasms blood supply, Breast Neoplasms pathology, Neovascularization, Pathologic pathology, Obesity pathology
Abstract

Breast cancer is correlated with a higher risk of metastasis in obese postmenopausal women. Adipokines, whose plasma concentrations are modulated in obese subjects and adipocytes surround mammary cells, suggesting that adipocyte secretome affect mammary tumorogenesis. We hypothesize that mature adipocyte secretions from obese women conditioned or not by breast neoplasic cells, increase changes on the angiogenesis stages. Supernatants of human mature adipocytes, differentiated from stem cells of either adipose tissue of normal weight (MA20) or obese (MA30) women or obtained from co-cultures between MA20 and MA30 and breast cancer cell line MCF-7, were collected. The impact of these supernatants was investigated on proliferation, migration, and tube formation by endothelial cells (HUVEC). MA20 and MA30 showed a preservation of their "metabolic memory" (increase of Leptin, ObR, VEGF, CYP19A1, and a decrease of Adiponectin expression in MA30 compared to MA20). Supernatants from obese-adipocytes increased HUVEC proliferation, migration, and sprouting like with supernatants obtained from co-cultures of MA/MCF-7 regardless the women's BMI. Additional analyses such as the use of neutralizing antibodies, analysis of supernatants (Milliplex®) and variations in gene expression (qRT-PCR), strongly suggest an implication of IL-6, or a synergistic action among adipokines, probably associated with that of VEGF or IL-6. As a conclusion, supernatants from co-cultures of MA30 and MCF-7 cells increase proliferation, migration, and sprouting of HUVEC cells. These results provide insights into the interaction between adipocytes and epithelial cancer cells, particularly in case of obesity. The identification of synergistic action of adipokines would therefore be a great interest in developing preventive strategies. J. Cell. Physiol. 232: 1808-1816, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Published
2017
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44. Eye bank prepared versus surgeon cut endothelial graft tissue for Descemet membrane endothelial keratoplasty: An observational study.

Author

Regnier M, Auxenfans C, Maucort-Boulch D, Marty AS, Damour O, Burillon C, and Kocaba V

Subjects
Aged, Aged, 80 and over, Endothelium, Corneal pathology, Endothelium, Corneal surgery, Female, Fuchs' Endothelial Dystrophy pathology, Graft Rejection, Humans, Male, Middle Aged, Organ Size, Postoperative Complications, Retrospective Studies, Surgeons, Treatment Outcome, Visual Acuity, Descemet Membrane pathology, Descemet Membrane surgery, Descemet Stripping Endothelial Keratoplasty methods, Eye Banks, Fuchs' Endothelial Dystrophy surgery
Abstract

The purpose of this article is to examine outcomes of Descemet membrane endothelial keratoplasty (DMEK) performed with cornea bank (CB) prestripped tissue and surgeon stripped tissue (SST).This retrospective study examined subjects who underwent DMEK with CB or surgeon prepared tissue for Fuchs endothelial corneal dystrophy. Best-corrected visual acuity (BCVA), corneal thickness, endothelial cell count (ECC), and complications were examined before and throughout a 6-month postoperative period.Eleven CB and 22 SST subjects were included. Six months after surgery, BCVA was 20/20 or better in 36.4% of CB and 22.7% of SST subjects (P = .43). Median logMAR BCVA was 0.10 (0.00-0.20, 20/25) in group CB and 0.10 (0.10-0.30, 20/25) in group SST. Median preoperative corneal thickness was 614.0 μm (577.5-662.0 μm) and 658.0 μm (606.0-689.0 μm) in CB and SST subjects, respectively (P = .37). Six months after surgery, median corneal thickness was lower in the CB group (571.0 μm [478.0-592.0 μm]), than in the SST group (576.0 μm [531.0-607.0 μm], P = .02). At 6 months, median ECC was 1500.0 cell/mm (1321.5-2049.0 cell/mm, 41% decrease) in group CB and 1403.0 cell/mm (972.5-2010.7 cell/mm, 46% decrease) in group SST (P = .70). Rebubbling was required in 5 CB (45.5%) and 15 SST (68.2%) subjects (P = .39).Fuchs' dystrophy patients have good anatomic and functional DMEK results. Similar outcomes and complication rates occurred with eye bank and surgeon prepared donor tissue.

Published
2017
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45. Proteomic Analysis of the Spatio-temporal Based Molecular Kinetics of Acute Spinal Cord Injury Identifies a Time- and Segment-specific Window for Effective Tissue Repair.

Author

Devaux S, Cizkova D, Quanico J, Franck J, Nataf S, Pays L, Hauberg-Lotte L, Maass P, Kobarg JH, Kobeissy F, Mériaux C, Wisztorski M, Slovinska L, Blasko J, Cigankova V, Fournier I, and Salzet M

Subjects
Animals, Disease Models, Animal, Humans, Protein Array Analysis, Protein Interaction Maps, Rats, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods, Time Factors, Biomarkers metabolism, Cytokines metabolism, Proteomics methods, Spinal Cord Injuries metabolism
Abstract

Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic burden on the public and private sectors worldwide. Although several studies have been conducted to identify the molecular progression of injury sequel due from the lesion site, still the exact underlying mechanisms and pathways of injury development have not been fully elucidated. In this work, based on OMICs, 3D matrix-assisted laser desorption ionization (MALDI) imaging, cytokines arrays, confocal imaging we established for the first time that molecular and cellular processes occurring after SCI are altered between the lesion proximity, i.e. rostral and caudal segments nearby the lesion (R1-C1) whereas segments distant from R1-C1, i.e. R2-C2 and R3-C3 levels coexpressed factors implicated in neurogenesis. Delay in T regulators recruitment between R1 and C1 favor discrepancies between the two segments. This is also reinforced by presence of neurites outgrowth inhibitors in C1, absent in R1. Moreover, the presence of immunoglobulins (IgGs) in neurons at the lesion site at 3 days, validated by mass spectrometry, may present additional factor that contributes to limited regeneration. Treatment in vivo with anti-CD20 one hour after SCI did not improve locomotor function and decrease IgG expression. These results open the door of a novel view of the SCI treatment by considering the C1 as the therapeutic target., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

Published
2016
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46. Erythropoietin Levels Increase during Cerebral Malaria and Correlate with Heme, Interleukin-10 and Tumor Necrosis Factor-Alpha in India.

Author

Dalko E, Tchitchek N, Pays L, Herbert F, Cazenave PA, Ravindran B, Sharma S, Nataf S, Das B, and Pied S

Subjects
Adult, Antigens, Protozoan metabolism, Chemokine CCL2 blood, Female, Hemopexin metabolism, Humans, India, Malaria, Cerebral parasitology, Male, Plasmodium falciparum physiology, Protozoan Proteins metabolism, Severity of Illness Index, Young Adult, Erythropoietin blood, Heme metabolism, Interleukin-10 blood, Malaria, Cerebral blood, Tumor Necrosis Factor-alpha blood
Abstract

Cerebral malaria (CM) caused by Plasmodium falciparum parasites often leads to the death of infected patients or to persisting neurological sequelae despite anti-parasitic treatments. Erythropoietin (EPO) was recently suggested as a potential adjunctive treatment for CM. However diverging results were obtained in patients from Sub-Saharan countries infected with P. falciparum. In this study, we measured EPO levels in the plasma of well-defined groups of P. falciparum-infected patients, from the state of Odisha in India, with mild malaria (MM), CM, or severe non-CM (NCM). EPO levels were then correlated with biological parameters, including parasite biomass, heme, tumor necrosis factor (TNF)-α, interleukin (IL)-10, interferon gamma-induced protein (IP)-10, and monocyte chemoattractant protein (MCP)-1 plasma concentrations by Spearman's rank and multiple correlation analyses. We found a significant increase in EPO levels with malaria severity degree, and more specifically during fatal CM. In addition, EPO levels were also found correlated positively with heme, TNF-α, IL-10, IP-10 and MCP-1 during CM. We also found a significant multivariate correlation between EPO, TNF-α, IL-10, IP-10 MCP-1 and heme, suggesting an association of EPO with a network of immune factors in CM patients. The contradictory levels of circulating EPO reported in CM patients in India when compared to Africa highlights the need for the optimization of adjunctive treatments according to the targeted population.

Published
2016
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47. Molecular Consequences of Proprotein Convertase 1/3 (PC1/3) Inhibition in Macrophages for Application to Cancer Immunotherapy: A Proteomic Study.

Author

Duhamel M, Rodet F, Delhem N, Vanden Abeele F, Kobeissy F, Nataf S, Pays L, Desjardins R, Gagnon H, Wisztorski M, Fournier I, Day R, and Salzet M

Subjects
Animals, CD4-Positive T-Lymphocytes drug effects, CD4-Positive T-Lymphocytes immunology, CD4-Positive T-Lymphocytes pathology, CD8-Positive T-Lymphocytes drug effects, CD8-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes pathology, Calcium metabolism, Cell Line, Cell Line, Tumor, Cytokines biosynthesis, Cytokines immunology, Gene Expression Regulation, Humans, Killer Cells, Natural drug effects, Killer Cells, Natural immunology, Killer Cells, Natural pathology, Lipopolysaccharides pharmacology, Macrophages, Alveolar cytology, Macrophages, Alveolar drug effects, Macrophages, Peritoneal cytology, Macrophages, Peritoneal drug effects, Mice, Mice, Knockout, Myeloid Differentiation Factor 88 genetics, Myeloid Differentiation Factor 88 immunology, Primary Cell Culture, Proprotein Convertase 1 genetics, Proprotein Convertase 1 immunology, Protein Array Analysis, RNA, Small Interfering genetics, RNA, Small Interfering metabolism, Rats, Signal Transduction, T-Lymphocytes, Helper-Inducer drug effects, T-Lymphocytes, Helper-Inducer immunology, T-Lymphocytes, Helper-Inducer pathology, Toll-Like Receptor 4 genetics, Toll-Like Receptor 4 immunology, Immunotherapy methods, Macrophages, Alveolar immunology, Macrophages, Peritoneal immunology, Proprotein Convertase 1 antagonists & inhibitors
Abstract

Macrophages provide the first line of host immune defense. Their activation triggers the secretion of pro-inflammatory cytokines and chemokines recruiting other immune cells. In cancer, macrophages present an M2 anti-inflammatory phenotype promoting tumor growth. In this way, strategies need to be develop to reactivate macrophages. Previously thought to be expressed only in cells with a neural/neuroendocrine phenotype, the proprotein convertase 1/3 has been shown to also be expressed in macrophages and regulated as a function of the Toll-like receptor immune response. Here, we investigated the intracellular impact of the down-regulation of the proprotein convertase 1/3 in NR8383 macrophages and confirmed the results on macrophages from PC1/3 deficient mice. A complete proteomic study of secretomes and intracellular proteins was undertaken and revealed that inhibition of proprotein convertase 1/3 orient macrophages toward an M1 activated phenotype. This phenotype is characterized by filopodial extensions, Toll-like receptor 4 MyD88-dependent signaling, calcium entry augmentation and the secretion of pro-inflammatory factors. In response to endotoxin/lipopolysaccharide, these intracellular modifications increased, and the secreted factors attracted naïve T helper lymphocytes to promote the cytotoxic response. Importantly, the application of these factors onto breast and ovarian cancer cells resulted in a decrease viability or resistance. Under inhibitory conditions using interleukin 10, PC1/3-knockdown macrophages continued to secrete inflammatory factors. These data indicate that targeted inhibition of proprotein convertase 1/3 could represent a novel type of immune therapy to reactivate intra-tumoral macrophages., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Published
2015
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50. Subcutaneous fat tissue engineering using autologous adipose-derived stem cells seeded onto a collagen scaffold.

Author

Lequeux C, Oni G, Wong C, Damour O, Rohrich R, Mojallal A, and Brown SA

Subjects
Adipocytes, White, Animals, Bromodeoxyuridine, Female, Fluorescent Antibody Technique, Pilot Projects, Subcutaneous Fat physiology, Sus scrofa, Wound Healing, Collagen, Guided Tissue Regeneration methods, Stem Cell Transplantation methods, Subcutaneous Fat cytology, Tissue Engineering methods, Tissue Scaffolds
Abstract

Background: This pilot study examined the efficacy of 5-bromo-2-deoxyuridine-labeled autologous adipose-derived stem cells seeded onto collagen scaffolds to augment and/or regenerate the fat-enriched hypodermal tissue in an acute porcine wound model., Methods: Porcine autologous adipose-derived stem cells were isolated and cultured. At passage 2, the cells were labeled with 5-bromo-2-deoxyuridine, seeded onto a three-dimensional collagen scaffold, and cultured for 10 days. Scaffolds were implanted subcutaneously in adult pigs with two adipose-derived stem cell scaffolds and two control scaffolds. Animals were euthanized at 2, 4, 8, and 12 weeks; all scaffold conditions were explanted for histology and immunohistochemistry analyses., Results: For all time points, adipose-derived stem cell scaffolds had increased connective tissue matrix within the subcutaneous tissue compared with scaffold alone and untreated porcine skin (p < 0.01). The neosynthesized connective tissue was vascularized and composed of small cells within an abundant extracellular matrix organized in layers. 5-Bromo-2-deoxyuridine cells were detectable only up to 4 weeks and mature adipocytes were absent. Levels of collagen types I, III, and VI differed among the experimental groups, with increased extracellular matrix associated with the presence of adipose-derived stem cells., Conclusions: The authors' data clearly show the efficacy of adipose-derived stem cells for soft-tissue repair and skin aging because it induces a significant increase of the dermis thickness. Moreover, the authors' results demonstrate the interest of their acute wound model and allowed them to show the skin thickness variation over time of the experiment, which is one of the challenges with which clinicians struggle in fat grafting.

Published
2012
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