43 results on '"Sitzia, C"'
Search Results
2. Novel insight into stem cell trafficking in dystrophic muscles
- Author
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Farini A, Villa C, Manescu A, Fiori F, Giuliani A, Razini P, Sitzia C, Del Fraro G, Belicchi M, Meregalli M, Rustichelli F, and Torrente Y
- Subjects
Medicine (General) ,R5-920 - Abstract
Andrea Farini,1,* Chiara Villa,1,* Adrian Manescu,2 Fabrizio Fiori,2 Alessandra Giuliani,2 Paola Razini,1 Clementina Sitzia,1 Giulia Del Fraro,1 Marzia Belicchi,1 Mirella Meregalli,1 Franco Rustichelli,2 Yvan Torrente1 1Stem Cell Laboratory, Department of Neurological Sciences, Fondazione IRCCS Cà Granda Ospedale Maggiore Policlinico di Milano, Centro Dino Ferrari, Università di Milano, Milano, Italy; 2Università Politecnica delle Marche, Di.S.C.O. - Sezione di Biologia, Biochimica e Fisica Ancona, Italy*These authors contributed equally to this paperAbstract: Recently published reports have described possible cellular therapy approaches to regenerate muscle tissues using arterial route delivery. However, the kinetic of distribution of these migratory stem cells within injected animal muscular dystrophy models is unknown. Using living X-ray computed microtomography, we established that intra-arterially injected stem cells traffic to multiple muscle tissues for several hours until their migration within dystrophic muscles. Injected stem cells express multiple traffic molecules, including VLA-4, LFA-1, CD44, and the chemokine receptor CXCR4, which are likely to direct these cells into dystrophic muscles. In fact, the majority of intra-arterially injected stem cells access the muscle tissues not immediately after the injection, but after several rounds of recirculation. We set up a new, living, 3D-imaging approach, which appears to be an important way to investigate the kinetic of distribution of systemically injected stem cells within dystrophic muscle tissues, thereby providing supportive data for future clinical applications.Keywords: iron nanoparticles, micro-CT, CD133+ stem cells, dystrophic muscles
- Published
- 2012
3. Preliminary Evidences of Safety and Efficacy of Flavonoids- and Omega 3-Based Compound for Muscular Dystrophies Treatment: A Randomized Double-Blind Placebo Controlled Pilot Clinical Trial
- Author
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Sitzia, C, Meregalli, M, Belicchi, M, Farini, A, Arosio, M, Bestetti, D, Villa, C, Valenti, L, Brambilla, P, Torrente, Y, Sitzia, Clementina, Meregalli, Mirella, Belicchi, Marzia, Farini, Andrea, Arosio, Maddalena, Bestetti, Denise, Villa, Chiara, Valenti, Luca, Brambilla, Paolo, Torrente, Yvan, Sitzia, C, Meregalli, M, Belicchi, M, Farini, A, Arosio, M, Bestetti, D, Villa, C, Valenti, L, Brambilla, P, Torrente, Y, Sitzia, Clementina, Meregalli, Mirella, Belicchi, Marzia, Farini, Andrea, Arosio, Maddalena, Bestetti, Denise, Villa, Chiara, Valenti, Luca, Brambilla, Paolo, and Torrente, Yvan
- Abstract
Background: Nutritional compounds can exert both anti-inflammatory and anti-oxidant effects. Since these events exacerbate the pathophysiology of muscular dystrophies, we investigated nutraceutical supplementation as an adjuvant therapy in dystrophic patients, to low costs and easy route of administration. Moreover, this treatment could represent an alternative therapeutic strategy for dystrophic patients who do not respond to corticosteroid treatment. Objective: A 24 weeks randomized double-blind placebo-controlled clinical study was aimed at evaluating the safety and efficacy of daily oral administration of flavonoids- and omega3-based natural supplement (FLAVOMEGA) in patients affected by muscular dystrophy with recognized muscle inflammation. Design: We screened 60 patients diagnosed for Duchenne (DMD), Facioscapulohumeral (FSHD), and Limb Girdle Muscular Dystrophy (LGMD). Using a computer-generated random allocation sequence, we stratified patients in a 2:1:1 ratio (DMD:FSHD:LGMD) to one of two treatment groups: continuous FLAVOMEGA, continuous placebo. Of 29 patients included, only 24 completed the study: 15 were given FLAVOMEGA, 14 placebo. Results: FLAVOMEGA was well tolerated with no reported adverse events. Significant treatment differences in the change from baseline in 6 min walk distance (6MWD; secondary efficacy endpoint) (P = 0.033) and in isokinetic knee extension (P = 0.039) (primary efficacy endpoint) were observed in LGMD and FSHD subjects. Serum CK levels (secondary efficacy endpoint) decreased in all FLAVOMEGA treated groups with significant difference in DMD subjects (P = 0.039). Conclusions: Although the small number of patients and the wide range of disease severity among patients reduced statistical significance, we obtained an optimal profile of safety and tolerability for the compound, showing valuable data of efficacy in primary and secondary endpoints.
- Published
- 2019
4. Reversine selectively induces cell death in tumor cells
- Author
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Piccoli M, Palazzolo G, Papini N, Dileo L, Sitzia C, Conforti E, Tettamanti G, Venerando B, Anastasia L, Piccoli, M., Palazzolo, G., Papini, N., Dileo, L., Sitzia, C., Conforti, E., Tettamanti, G., Venerando, B., Anastasia, L., Piccoli, M, Palazzolo, G, Papini, N, Dileo, L, Sitzia, C, Conforti, E, Tettamanti, G, Venerando, B, and Anastasia, L
- Abstract
Reversine is a synthetic 2,6-distribuited purine1, which has been shown to increase cell plasticity with the ability to reprogram lineage-committed cells to a more primitive multipotent state. In particular, reversine-treated dermal fibroblasts could be then induced to differentiate to skeletal muscle both in vitro and in vivo2. Nevertheless, reversine mechanism of action is still not fully demonstrated, although it has been shown that the molecule inhibits MEK1, nonmuscle myosin II heavy chain3, and aurora kinases. Although it is still unclear if these effects are strictly related to de-differentiation, it is now quite clear that the molecule inhibits cell growth. Thus, it has been already reported that reversine inhibits cell growth and colony formation in tumor cells. Surprisingly and remarkably, in our hands, treatment of several types of cancer cells (including neuroblastoma, fibrosarcoma, and glioblastoma cancer stem cells) with reversine at micromolar concentration, not only inhibited tumor cells growth, but extensively induced cell death. On the other hand, we did not observe the same lethal effects on normal cells (including human and murine primary dermal fibroblast, murine myoblasts, rat and human mesenchymal stem cells, and murine mesangioblasts), but only a reversible cell growth arrest. Moreover, we found that tumor cells seem to undergo different death pathways. In fact, while neuroblastoma SK-N-BE cells revealed all the canonical signs of apoptosis (chromatin condensation, caspase 3 activation), we could not find the same evidences in fibrosarcoma cells HT1080. Nevertheless, cell cycle analysis and morphological features, after reversine treatment, seem to point out that cell death may be due to mitotic catastrophe, which may be caspase dependent or independent. In fact, reversine treatment leads to the formation of large cells with several micronuclei, possibly due to the aberrant chromosome segregation and the inhibition of cytokinesis. Reversine inhibition of aurora kinase and nonmuscle myosin II heavy chain, which hold crucial roles in the correct mitotic division, may result in the observed effects. Cell cycle analysis shows that reversine induces endoreplication, but while normal cells can control this effect and block cells in G1 phase, tumor cells, where G1 checkpoint is de-regulated, keep cycling to a point where micronucleated polyploid populations start to die.
- Published
- 2008
5. NEU3 sialidase strictly modulates GM3 levels in skeletal myoblasts C2C12 thus favoring their differentiation and protecting them from apoptosis
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Palazzolo G, Anastasia L, Papini N Colazzo F, Tringali C, Dileo L, Piccoli M, Conforti E, Sitzia C, Monti E, Sampaolesi M, Tettamanti G, Venerando B, Palazzolo, G, Anastasia, L, Papini, N Colazzo F, Tringali, C, Dileo, L, Piccoli, M, Conforti, E, Sitzia, C, Monti, E, Sampaolesi, M, Tettamanti, G, and Venerando, B
- Published
- 2008
6. Stem Cell-Mediated Exon Skipping of the Dystrophin Gene by the Bystander Effect
- Author
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Meregalli, M, Farini, A, Sitzia, C, Beley, C, Razini, P, Cassinelli, L, Colleoni, F, Frattini, P, Santo, N, Galbiati, E, Prosperi, D, Tavelli, A, Belicchi, M, Garcia, L, Torrente, Y, Torrente, Y., CASSINELLI, LETIZIA MARIA, GALBIATI, ELISABETTA, PROSPERI, DAVIDE, Meregalli, M, Farini, A, Sitzia, C, Beley, C, Razini, P, Cassinelli, L, Colleoni, F, Frattini, P, Santo, N, Galbiati, E, Prosperi, D, Tavelli, A, Belicchi, M, Garcia, L, Torrente, Y, Torrente, Y., CASSINELLI, LETIZIA MARIA, GALBIATI, ELISABETTA, and PROSPERI, DAVIDE
- Abstract
Duchenne muscular dystrophy (DMD) is characterized by the loss of a functional dystrophin protein; the muscles of DMD patients progressively degenerate as a result of mechanical stress during contractions, and the condition eventually leads to premature death. By means antisense oligonucleotides (AONs), it is possible to modulate pre-mRNA splicing eliminating mutated exons and restoring dystrophin open reading frame. To overcome the hurdles in using AONs for therapeutic interventions, we exerted engineered human DMD stem cells with a lentivirus, which permanently and efficiently delivered the cloned AONs. Here we describe for the first time the exosome-mediated release of AONs from engineered human DMD CD133+ stem cells allowing the rescue of murine dystrophin expression. Finally, upon release, AONs could be internalized by host cells suggesting a potential role of exosomes acting as vesicular carriers for DMD gene therapy.
- Published
- 2015
7. P4.52 Combining gene and stem cell therapy in the treatment of dysferlinophaties
- Author
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Meregalli, M., primary, Sitzia, C., additional, Navarro, C., additional, Parolini, D., additional, Belicchi, M., additional, Razini, P., additional, Farini, A., additional, Khran, M., additional, Garcia, L., additional, Levy, N., additional, and Torrente, Y., additional
- Published
- 2011
- Full Text
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8. O.9 Stem cell therapy of muscular dystrophies using exon skipping approach in GRMD dogs
- Author
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Meregalli, M., primary, Farini, A., additional, Belicchi, M., additional, Parolini, D., additional, Sitzia, C., additional, Cassinelli, L., additional, Del Fraro, G., additional, Razini, P., additional, Angeloni, V., additional, Jardim, L., additional, da Silva Bizario, J.C., additional, Garcia, L., additional, and Torrente, Y., additional
- Published
- 2011
- Full Text
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9. Vibration of Thick Circular Plate--Determination of Elastic Properties Through the Resonant Behaviour
- Author
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Priolo, P., primary and Sitzia, C., additional
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- 1983
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10. Efficiency of annular finite elements for flexural vibrations of thick disks
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Priolo, P., primary and Sitzia, C., additional
- Published
- 1984
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11. Preliminary Evidences of Safety and Efficacy of Flavonoids- and Omega 3-Based Compound for Muscular Dystrophies Treatment: A Randomized Double-Blind Placebo Controlled Pilot Clinical Trial
- Author
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Luca Valenti, Denise Bestetti, Marzia Belicchi, Mirella Meregalli, Maddalena Arosio, Clementina Sitzia, Chiara Villa, Andrea Farini, Yvan Torrente, Paolo Brambilla, Sitzia, C, Meregalli, M, Belicchi, M, Farini, A, Arosio, M, Bestetti, D, Villa, C, Valenti, L, Brambilla, P, and Torrente, Y
- Subjects
0301 basic medicine ,musculoskeletal diseases ,Duchenne muscular dystrophy ,safety ,medicine.medical_specialty ,BIO/12 - BIOCHIMICA CLINICA E BIOLOGIA MOLECOLARE CLINICA ,Placebo ,lcsh:RC346-429 ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,medicine ,Adjuvant therapy ,Muscular dystrophy ,tolerability ,Adverse effect ,strength recovery ,lcsh:Neurology. Diseases of the nervous system ,business.industry ,medicine.disease ,Clinical Trial ,Clinical trial ,030104 developmental biology ,Tolerability ,Neurology ,Neurology (clinical) ,nutraceutical supplementation ,business ,030217 neurology & neurosurgery ,Limb-girdle muscular dystrophy - Abstract
Background: Nutritional compounds can exert both anti-inflammatory and anti-oxidant effects. Since these events exacerbate the pathophysiology of muscular dystrophies, we investigated nutraceutical supplementation as an adjuvant therapy in dystrophic patients, to low costs and easy route of administration. Moreover, this treatment could represent an alternative therapeutic strategy for dystrophic patients who do not respond to corticosteroid treatment. Objective: A 24 weeks randomized double-blind placebo-controlled clinical study was aimed at evaluating the safety and efficacy of daily oral administration of flavonoids- and omega3-based natural supplement (FLAVOMEGA) in patients affected by muscular dystrophy with recognized muscle inflammation. Design: We screened 60 patients diagnosed for Duchenne (DMD), Facioscapulohumeral (FSHD), and Limb Girdle Muscular Dystrophy (LGMD). Using a computer-generated random allocation sequence, we stratified patients in a 2:1:1 ratio (DMD:FSHD:LGMD) to one of two treatment groups: continuous FLAVOMEGA, continuous placebo. Of 29 patients included, only 24 completed the study: 15 were given FLAVOMEGA, 14 placebo. Results: FLAVOMEGA was well tolerated with no reported adverse events. Significant treatment differences in the change from baseline in 6 min walk distance (6MWD; secondary efficacy endpoint) (P = 0.033) and in isokinetic knee extension (P = 0.039) (primary efficacy endpoint) were observed in LGMD and FSHD subjects. Serum CK levels (secondary efficacy endpoint) decreased in all FLAVOMEGA treated groups with significant difference in DMD subjects (P = 0.039). Conclusions: Although the small number of patients and the wide range of disease severity among patients reduced statistical significance, we obtained an optimal profile of safety and tolerability for the compound, showing valuable data of efficacy in primary and secondary endpoints. Trial registration number: NCT03317171 Retrospectively registered 25/10/2017.
- Published
- 2019
12. Stem Cell-Mediated Exon Skipping of the Dystrophin Gene by the Bystander Effect
- Author
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Elisabetta Galbiati, Cyriaque Beley, Nadia Santo, Letizia Cassinelli, Paola Frattini, Marzia Belicchi, Paola Razini, Alessandro Tavelli, A. Farini, Mirella Meregalli, Luis Garcia, Yvan Torrente, Federica Colleoni, Clementina Sitzia, Davide Prosperi, Meregalli, M, Farini, A, Sitzia, C, Beley, C, Razini, P, Cassinelli, L, Colleoni, F, Frattini, P, Santo, N, Galbiati, E, Prosperi, D, Tavelli, A, Belicchi, M, Garcia, L, and Torrente, Y
- Subjects
musculoskeletal diseases ,congenital, hereditary, and neonatal diseases and abnormalities ,RNA Splicing ,Duchenne muscular dystrophy ,Mice, SCID ,Dystrophin ,Mice ,Exon ,Drug Discovery ,Genetics ,medicine ,Animals ,Humans ,Muscular dystrophy ,Muscle, Skeletal ,Molecular Biology ,Cells, Cultured ,Genetics (clinical) ,biology ,Stem Cells ,Myogenic stem cell ,Bystander Effect ,Exons ,Genetic Therapy ,Oligonucleotides, Antisense ,medicine.disease ,Molecular biology ,Exon skipping ,Microvesicles ,Cell biology ,Muscular Dystrophy, Duchenne ,RNA splicing ,biology.protein ,Molecular Medicine ,Delivery of exon skipping machinery ,Stem cell - Abstract
Duchenne muscular dystrophy (DMD) is characterized by the loss of a functional dystrophin protein; the muscles of DMD patients progressively degenerate as a result of mechanical stress during contractions, and the condition eventually leads to premature death. By means antisense oligonucleotides (AONs), it is possible to modulate pre-mRNA splicing eliminating mutated exons and restoring dystrophin open reading frame. To overcome the hurdles in using AONs for therapeutic interventions, we exerted engineered human DMD stem cells with a lentivirus, which permanently and efficiently delivered the cloned AONs. Here we describe for the first time the exosome-mediated release of AONs from engineered human DMD CD133+ stem cells allowing the rescue of murine dystrophin expression. Finally, upon release, AONs could be internalized by host cells suggesting a potential role of exosomes acting as vesicular carriers for DMD gene therapy.
- Published
- 2015
13. NEU3 Sialidase Strictly Modulates GM3 Levels in Skeletal Myoblasts C2C12 Thus Favoring Their Differentiation and Protecting Them from Apoptosis
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Eugenio Monti, Guido Tettamanti, Maurilio Sampaolesi, Marco Piccoli, Nadia Papini, Cristina Tringali, Luigi Anastasia, Bruno Venerando, Francesca Colazzo, Erika Conforti, Giacomo Palazzolo, Clementina Sitzia, L. Dileo, Anastasia, L., Papini, N., Colazzo, F., Palazzolo, G., Tringali, C., Dileo, L., Piccoli, M., Conforti, E., Sitzia, C., Monti, E., Sampaolesi, M., Tettamanti, G., and Venerando, B.
- Subjects
Down-Regulation ,Neuraminidase ,Glycobiology and Extracellular Matrices ,Biology ,Sialidase ,Biochemistry ,Cell Line ,Cell membrane ,Mice ,SKELETAL MYOBLASTS ,DIFFERENTIATION ,APOPTOSIS ,medicine ,Animals ,G(M3) Ganglioside ,Myocyte ,Gene Silencing ,Epidermal growth factor receptor ,skeletal muscle ,Muscle, Skeletal ,Molecular Biology ,ganglioside GM3 ,Sphingolipids ,Ganglioside ,Epidermal Growth Factor ,Hydrolysis ,Skeletal muscle ,C2V12 ,Cell Biology ,sialidase NEU3 ,cell differentiation ,Cell biology ,ErbB Receptors ,medicine.anatomical_structure ,Models, Chemical ,Cancer cell ,biology.protein ,C2C12 - Abstract
Membrane-bound sialidase NEU3, often referred to as the "ganglioside sialidase", has a critical regulatory function on the sialoglycosphingolipid pattern of the cell membrane, with an anti-apoptotic function, especially in cancer cells. Although other sialidases have been shown to be involved in skeletal muscle differentiation, the role of NEU3 had yet to be disclosed. Herein we report that NEU3 plays a key role in skeletal muscle differentiation by strictly modulating the ganglioside content of adjacent cells, with special regards to GM3. Induced down-regulation of NEU3 in murine myoblasts C2C12, even when partial, totally inhibits their capability to differentiate by increasing GM3 level above a critical point, that causes EGFR inhibition (and ultimately its down-regulation), and an higher responsiveness of myoblasts to the apoptotic stimuli. ispartof: Journal of Biological Chemistry vol:283 issue:52 pages:36265-36271 ispartof: location:United States status: published
- Published
- 2008
14. Unveiling IL-33/ST2 Pathway Unbalance in Cardiac Remodeling Due to Obesity in Zucker Fatty Rats.
- Author
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Sitzia C, Vianello E, Dozio E, Kalousová M, Zima T, Brizzola S, Roccabianca P, Tedeschi G, Lamont J, Tacchini L, and Corsi-Romanelli MM
- Subjects
- Animals, Rats, Cardiomegaly genetics, Cardiomegaly metabolism, Disease Models, Animal, Fibrosis genetics, Fibrosis metabolism, Ghrelin genetics, Ghrelin metabolism, Guanine Nucleotide Exchange Factors genetics, Guanine Nucleotide Exchange Factors metabolism, Interleukin-1 Receptor-Like 1 Protein genetics, Interleukin-1 Receptor-Like 1 Protein metabolism, Myocardium metabolism, Rats, Zucker, Interleukin-33 genetics, Interleukin-33 metabolism, Obesity complications, Obesity genetics, Obesity metabolism, Receptors, Interleukin-1 genetics, Receptors, Interleukin-1 metabolism, Ventricular Remodeling genetics, Ventricular Remodeling physiology
- Abstract
Obesity is an epidemic condition linked to cardiovascular disease severity and mortality. Fat localization and type represent cardiovascular risk estimators. Importantly, visceral fat secretes adipokines known to promote low-grade inflammation that, in turn, modulate its secretome and cardiac metabolism. In this regard, IL-33 regulates the functions of various immune cells through ST2 binding and-following its role as an immune sensor to infection and stress-is involved in the pro-fibrotic remodeling of the myocardium. Here we further investigated the IL-33/ST2 effects on cardiac remodeling in obesity, focusing on molecular pathways linking adipose-derived IL-33 to the development of fibrosis or hypertrophy. We analyzed the Zucker Fatty rat model, and we developed in vitro models to mimic the adipose and myocardial relationship. We demonstrated a dysregulation of IL-33/ST2 signaling in both adipose and cardiac tissue, where they affected Epac proteins and myocardial gene expression, linked to pro-fibrotic signatures. In Zucker rats, pro-fibrotic effects were counteracted by ghrelin-induced IL-33 secretion, whose release influenced transcription factor expression and ST2 isoforms balance regulation. Finally, the effect of IL-33 signaling is dependent on several factors, such as cell types' origin and the balancing of ST2 isoforms. Noteworthy, it is reasonable to state that considering IL-33 to have a unique protective role should be considered over-simplistic.
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- 2023
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15. Plasma Levels of Glycohydrolase Activities in Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Patients.
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Massaccesi L, Vianello E, Ranucci M, Galliera E, Sitzia C, and Romanelli MMC
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- Biomarkers blood, Case-Control Studies, Humans, COVID-19 blood, COVID-19 diagnosis, Glycoside Hydrolases blood
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- 2021
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16. Sensitivity of serology assay in Covid-19 diagnosis: does the antigen matter?
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Sitzia C, Pistelli L, Cardani R, Renna LV, Ranucci M, Carrara M, Borlini S, Clerici P, Rampoldi B, Cornetta M, and Corsi-Romanelli M
- Subjects
- Antibodies, Viral, Humans, Immunoglobulin G, Italy, SARS-CoV-2, Sensitivity and Specificity, Spike Glycoprotein, Coronavirus, United States, COVID-19
- Abstract
Since the spreading of Sar-CoV-2 in March 2020, many serologic tests have been developed to identify antibody responses. Indeed, different commercial kits are directed against different antigens and could utilise different methods thereby triggering confusion and criticism. Here, we compared two Food and Drug Administration (FDA)-approved automatized assays that detect IgG responses against spike or nucleocapsid protein of Sars-Cov-2 virus in 127 subjects among healthcare workers of IRCCS Policlinico San Donato (MI), Italy. We observed different kinetics of IgG responses, demonstrating the importance of timing of sampling to correctly interpret the results both for infection diagnosis and for epidemiologic studies. We observed that Anti-N response starts earlier than Anti-S1/S2 response but also decreases earlier, affecting the sensitivity of the tests at different time points. Combining two different assays, designed against different antigens, could reduce false negative results. Finally, we observed a patient who produced anti-nucleocapsid IgG, but not anti-spike IgG. In conclusion, we investigated antibody responses in Covid-19 disease, aiming to direct clinicians and laboratory scientists to correctly interpret serologic results by always paying attention to clinical history correlation, timing of sampling, methods and antigens used, to avoid false negative results and obtain relevant epidemiologic data., (Copyright 2020 Biolife Sas. www.biolifesas.org.)
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- 2021
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17. Defective dystrophic thymus determines degenerative changes in skeletal muscle.
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Farini A, Sitzia C, Villa C, Cassani B, Tripodi L, Legato M, Belicchi M, Bella P, Lonati C, Gatti S, Cerletti M, and Torrente Y
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- Animals, Autophagy physiology, Ghrelin biosynthesis, Macrophages immunology, Male, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Mice, Inbred mdx, Mice, Nude, Muscular Dystrophy, Duchenne pathology, T-Lymphocytes transplantation, T-Lymphocytes, Regulatory immunology, Thymus Gland transplantation, Transcription Factors biosynthesis, AIRE Protein, Immune Tolerance immunology, Muscle, Skeletal pathology, Muscular Atrophy pathology, Muscular Dystrophy, Animal pathology, Thymus Gland pathology
- Abstract
In Duchenne muscular dystrophy (DMD), sarcolemma fragility and myofiber necrosis produce cellular debris that attract inflammatory cells. Macrophages and T-lymphocytes infiltrate muscles in response to damage-associated molecular pattern signalling and the release of TNF-α, TGF-β and interleukins prevent skeletal muscle improvement from the inflammation. This immunological scenario was extended by the discovery of a specific response to muscle antigens and a role for regulatory T cells (Tregs) in muscle regeneration. Normally, autoimmunity is avoided by autoreactive T-lymphocyte deletion within thymus, while in the periphery Tregs monitor effector T-cells escaping from central regulatory control. Here, we report impairment of thymus architecture of mdx mice together with decreased expression of ghrelin, autophagy dysfunction and AIRE down-regulation. Transplantation of dystrophic thymus in recipient nude mice determine the up-regulation of inflammatory/fibrotic markers, marked metabolic breakdown that leads to muscle atrophy and loss of force. These results indicate that involution of dystrophic thymus exacerbates muscular dystrophy by altering central immune tolerance.
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- 2021
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18. Intra-erythrocytes magnesium deficiency could reflect cognitive impairment status due to vascular disease: a pilot study.
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Sitzia C, Sterlicchio M, Crapanzano C, Dozio E, Vianello E, and Corsi Romanelli MM
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- Erythrocytes, Humans, Pilot Projects, Cognitive Dysfunction complications, Magnesium Deficiency complications, Vascular Diseases
- Abstract
Background and Aims: Magnesium is a fundamental cation that regulates neuronal transmission, protein synthesis, energy metabolism. Magnesium deficiency mostly affects nervous and cardiovascular systems determining weakness, tremors, seizure and arrhythmias. This condition retains also a role in memory function and neuronal plasticity. Importantly magnesium deficiency could remain latent and asymptomatic resulting a risk factor for cardiovascular disease. In this sense we aim to determine magnesium status in patient presenting cognitive impairment of vascular origin., Methods: 21 healthy subjects and 27 patients presenting vascular cognitive impairment were included in this study. Both plasma and intraerythrocitary magnesium level were measured to detect magnesium deficiency and cognitive performance was evaluated trough Mini Mental State Evaluation (MMSE)., Results: Here we showed that patients presenting vascular cognitive impairment present intraerythrocitary magnesium level lower than age-matched healthy subjects. To note their plasma magnesium resulted within reference limit., Conclusion: We suggest that intracellular magnesium laboratory measurement is needed to detect occult magnesium deficiency in population at risk. Magnesium supplementation could represent an adjuvant for healthy aging in high risk population.
- Published
- 2020
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19. Soluble Receptor for Advanced Glycation End Products and Its Forms in COVID-19 Patients with and without Diabetes Mellitus: A Pilot Study on Their Role as Disease Biomarkers.
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Dozio E, Sitzia C, Pistelli L, Cardani R, Rigolini R, Ranucci M, and Corsi Romanelli MM
- Abstract
The receptor for advanced glycation end products (RAGE), a well-known player of diabetes mellitus (DM)-related morbidities, was supposed to be involved in coronavirus disease-19 (COVID-19), but no data exist about COVID-19, DM, and the soluble RAGE (sRAGE) forms. We quantified total sRAGE and its forms, the endogenously secretory esRAGE and the membrane-cleaved cRAGE, in COVID-19 patients with and without DM and in healthy individuals to explore how COVID-19 may affect these molecules and their potential role as biomarkers. Circulating sRAGE and esRAGE were quantified by enzyme-linked-immunosorbent assays. cRAGE was obtained by subtracting esRAGE from total sRAGE. sRAGE, esRAGE, cRAGE, and the cRAGE/esRAGE ratio did not differ between DM and non-DM patients and had the same trend when compared to healthy individuals. Levels of total sRAGE, cRAGE, and cRAGE/esRAGE ratio were upregulated, while esRAGE was downregulated. The lack of difference between DM and non-DM COVID-19 patients in the levels of sRAGE and its forms supports the hypothesis that in COVID-19 the RAGE system is modulated regardless of glycemic control. Identifying how sRAGE and its forms associate to COVID-19 prognosis and the potential of RAGE as a therapeutic target to control inflammatory burden seem of relevance to help treatment of COVID-19.
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- 2020
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20. Anti-SARS CoV2 antibody testing in healthcare workers: comparison between rapid-cassette tests, ELISA and CLIA methods.
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Pistelli L, Sitzia C, Carrara M, Borlini S, Tejada M, Sammarco G, Fulceri F, Valaperta R, Giubbilini P, Rigolini R, Rampoldi B, Cornetta M, Mazza S, Corino SD, Pettinato A, Ferrari S, Paduraru C, Moroni A, Jance B, Costa E, and Corsi Romanelli MM
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- Humans, COVID-19 diagnosis, COVID-19 Serological Testing methods, Enzyme-Linked Immunosorbent Assay, Health Personnel, Immunoassay methods
- Published
- 2020
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21. Covid-19-Associated Coagulopathy: Biomarkers of Thrombin Generation and Fibrinolysis Leading the Outcome.
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Ranucci M, Sitzia C, Baryshnikova E, Di Dedda U, Cardani R, Martelli F, and Corsi Romanelli M
- Abstract
Background: Coronavirus Disease 2019 (COVID-19)-associated coagulopathy is characterized by a prothrombotic state not yet comprehensively studied. We investigated the coagulation pattern of patients with COVID-19 acute respiratory distress syndrome (ARDS), comparing patients who survived to those who did not. Methods: In this prospective cohort study on 20 COVID-19 ARDS patients, the following biomarkers were measured: thrombin generation (prothrombin fragment 1 + 2 (PF 1 + 2)), fibrinolysis activation (tissue plasminogen activator (tPA)) and inhibition (plasminogen activator inhibitor 2 (PAI-2)), fibrin synthesis (fibrinopeptide A) and fibrinolysis magnitude (plasmin-antiplasmin complex (PAP) and D-dimers). Measurements were done upon intensive care unit (ICU) admission and after 10-14 days. Results: There was increased thrombin generation; modest or null release of t-PA; and increased levels of PAI-2, fibrinopeptide A, PAP and D-dimers. At baseline, nonsurvivors had a significantly ( p = 0.014) higher PAI-2/PAP ratio than survivors (109, interquartile range (IQR) 18.1-216, vs. 8.7, IQR 2.9-12.6). At follow-up, thrombin generation was significantly ( p = 0.025) reduced in survivors (PF 1 + 2 from 396 pg/mL, IQR 185-585 to 237 pg/mL, IQR 120-393), whereas it increased in nonsurvivors. Fibrinolysis inhibition at follow-up remained stable in survivors and increased in nonsurvivors, leading to a significant ( p = 0.026) difference in PAI-2 levels (161 pg/mL, IQR 50-334, vs. 1088 pg/mL, IQR 177-1565). Conclusion: Severe patterns of COVID-19 ARDS are characterized by a thrombin burst and the consequent coagulation activation. Mechanisms of fibrinolysis regulation appear unbalanced toward fibrinolysis inhibition. This pattern ameliorates in survivors, whereas it worsens in nonsurvivors.
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- 2020
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22. PCSK9 Expression in Epicardial Adipose Tissue: Molecular Association with Local Tissue Inflammation.
- Author
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Dozio E, Ruscica M, Vianello E, Macchi C, Sitzia C, Schmitz G, Tacchini L, and Corsi Romanelli MM
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- Aged, Anthropometry, Body Mass Index, Case-Control Studies, Chemokines metabolism, Coronary Artery Disease complications, Female, Heart Valve Diseases complications, Humans, Lymphocytes metabolism, Male, Middle Aged, Monocytes metabolism, Protein Array Analysis, Risk, Adipose Tissue metabolism, Inflammation metabolism, Pericardium metabolism, Proprotein Convertase 9 metabolism
- Abstract
Epicardial adipose tissue (EAT) has the unique property to release mediators that nourish the heart in healthy conditions, an effect that becomes detrimental when volume expands and proinflammatory cytokines start to be produced. Proprotein convertase subtilisin/kexin type 9 (PCSK9), a proinflammatory mediator involved in atherosclerosis, is also produced by visceral fat. Due to the correlation of inflammation with PCSK9 and EAT enlargement, we evaluated whether PCSK9 was expressed in EAT and associated with EAT inflammation and volume. EAT samples were isolated during surgery. EAT thickness was measured by echocardiography. A microarray was used to explore EAT transcriptoma. The PCSK9 protein levels were measured by Western Blot in EAT and ELISA in plasma. PCSK9 was expressed at both the gene and protein levels in EAT. We found a positive association with EAT thickness and local proinflammatory mediators, in particular, chemokines for monocytes and lymphocytes. No association was found with the circulating PCSK9 level. The expression of PCSK9 in EAT argues that PCSK9 is part of the EAT secretome and EAT inflammation is associated with local PCSK9 expression, regardless of circulating PCSK9 levels. Whether reducing EAT inflammation or PCSK9 local levels may have beneficial effects on EAT metabolism and cardiovascular risk needs further investigations., Competing Interests: The authors declare no conflict of interest., (Copyright © 2020 Elena Dozio et al.)
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- 2020
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23. PTX3 Predicts Myocardial Damage and Fibrosis in Duchenne Muscular Dystrophy.
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Farini A, Villa C, Di Silvestre D, Bella P, Tripodi L, Rossi R, Sitzia C, Gatti S, Mauri P, and Torrente Y
- Abstract
Pentraxin 3 (PTX3) is a main component of the innate immune system by inducing complement pathway activation, acting as an inflammatory mediator, coordinating the functions of macrophages/dendritic cells and promoting apoptosis/necrosis. Additionally, it has been found in fibrotic regions co-localizing with collagen. In this work, we wanted to investigate the predictive role of PTX3 in myocardial damage and fibrosis of Duchenne muscular dystrophy (DMD). DMD is an X-linked recessive disease caused by mutations of the dystrophin gene that affects muscular functions and strength and accompanying dilated cardiomyopathy. Here, we expound the correlation of PTX3 cardiac expression with age and Toll-like receptors (TLRs)/interleukin-1 receptor (IL-1R)-MyD88 inflammatory markers and its modulation by the so-called alarmins IL-33, high-mobility group box 1 (HMGB1), and S100β. These findings suggest that cardiac levels of PTX3 might have prognostic value and potential in guiding therapy for DMD cardiomyopathy., (Copyright © 2020 Farini, Villa, Di Silvestre, Bella, Tripodi, Rossi, Sitzia, Gatti, Mauri and Torrente.)
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- 2020
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24. Circulating Irisin and esRAGE as Early Biomarkers of Decline of Metabolic Health.
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Dozio E, Vianello E, Sitzia C, Ambrogi F, Benedini S, Gorini S, Rampoldi B, Rigolini R, Tacchini L, and Romanelli MMC
- Abstract
A decline in metabolic health may take place before observing any alteration in the levels of the traditional metabolic markers. New indicators of metabolic derangement are therefore compelling. Irisin is a myokine with important metabolic functions. The role of irisin as a metabolic biomarker in humans has not been fully established yet. We quantified plasma irisin and esRAGE in 106 apparently healthy individuals and we performed a cluster analysis to evaluate their associations with metabolic profile. Plasma levels of various traditional markers of metabolic risk (i.e., glucose and lipid levels) were all within the ranges of normality. We identified two clusters of individuals. Compared to cluster 2, individuals in cluster 1 had higher irisin levels, a metabolic profile shifted toward the limits of the reference ranges and lower esRAGE levels. The traditional metabolic blood tests seem not to be enough to identify a metabolic decline early. Irisin increase and esRAGE decrease may reflect a metabolic derangement at the beginning of its development. The role of these molecules as early biomarkers of decline of metabolic health seems an interesting topic to be further explored., Competing Interests: The authors declare no conflict of interest.
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- 2020
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25. Blockade of IGF2R improves muscle regeneration and ameliorates Duchenne muscular dystrophy.
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Bella P, Farini A, Banfi S, Parolini D, Tonna N, Meregalli M, Belicchi M, Erratico S, D'Ursi P, Bianco F, Legato M, Ruocco C, Sitzia C, Sangiorgi S, Villa C, D'Antona G, Milanesi L, Nisoli E, Mauri P, and Torrente Y
- Subjects
- Animals, Binding Sites, Child, Humans, Mice, Mice, Inbred mdx, Myoblasts, Young Adult, Muscle, Skeletal growth & development, Muscular Dystrophy, Duchenne drug therapy, Receptor, IGF Type 2 antagonists & inhibitors, Regeneration
- Abstract
Duchenne muscular dystrophy (DMD) is a debilitating fatal X-linked muscle disorder. Recent findings indicate that IGFs play a central role in skeletal muscle regeneration and development. Among IGFs, insulinlike growth factor 2 (IGF2) is a key regulator of cell growth, survival, migration and differentiation. The type 2 IGF receptor (IGF2R) modulates circulating and tissue levels of IGF2 by targeting it to lysosomes for degradation. We found that IGF2R and the store-operated Ca
2+ channel CD20 share a common hydrophobic binding motif that stabilizes their association. Silencing CD20 decreased myoblast differentiation, whereas blockade of IGF2R increased proliferation and differentiation in myoblasts via the calmodulin/calcineurin/NFAT pathway. Remarkably, anti-IGF2R induced CD20 phosphorylation, leading to the activation of sarcoplasmic/endoplasmic reticulum Ca2+ -ATPase (SERCA) and removal of intracellular Ca2+ . Interestingly, we found that IGF2R expression was increased in dystrophic skeletal muscle of human DMD patients and mdx mice. Blockade of IGF2R by neutralizing antibodies stimulated muscle regeneration, induced force recovery and normalized capillary architecture in dystrophic mdx mice representing an encouraging starting point for the development of new biological therapies for DMD., (© 2019 The Authors. Published under the terms of the CC BY 4.0 license.)- Published
- 2020
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26. Preliminary Evidences of Safety and Efficacy of Flavonoids- and Omega 3-Based Compound for Muscular Dystrophies Treatment: A Randomized Double-Blind Placebo Controlled Pilot Clinical Trial.
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Sitzia C, Meregalli M, Belicchi M, Farini A, Arosio M, Bestetti D, Villa C, Valenti L, Brambilla P, and Torrente Y
- Abstract
Background: Nutritional compounds can exert both anti-inflammatory and anti-oxidant effects. Since these events exacerbate the pathophysiology of muscular dystrophies, we investigated nutraceutical supplementation as an adjuvant therapy in dystrophic patients, to low costs and easy route of administration. Moreover, this treatment could represent an alternative therapeutic strategy for dystrophic patients who do not respond to corticosteroid treatment. Objective: A 24 weeks randomized double-blind placebo-controlled clinical study was aimed at evaluating the safety and efficacy of daily oral administration of flavonoids- and omega3-based natural supplement (FLAVOMEGA) in patients affected by muscular dystrophy with recognized muscle inflammation. Design: We screened 60 patients diagnosed for Duchenne (DMD), Facioscapulohumeral (FSHD), and Limb Girdle Muscular Dystrophy (LGMD). Using a computer-generated random allocation sequence, we stratified patients in a 2:1:1 ratio (DMD:FSHD:LGMD) to one of two treatment groups: continuous FLAVOMEGA, continuous placebo. Of 29 patients included, only 24 completed the study: 15 were given FLAVOMEGA, 14 placebo. Results: FLAVOMEGA was well tolerated with no reported adverse events. Significant treatment differences in the change from baseline in 6 min walk distance (6MWD; secondary efficacy endpoint) ( P = 0.033) and in isokinetic knee extension ( P = 0.039) (primary efficacy endpoint) were observed in LGMD and FSHD subjects. Serum CK levels (secondary efficacy endpoint) decreased in all FLAVOMEGA treated groups with significant difference in DMD subjects ( P = 0.039). Conclusions: Although the small number of patients and the wide range of disease severity among patients reduced statistical significance, we obtained an optimal profile of safety and tolerability for the compound, showing valuable data of efficacy in primary and secondary endpoints. Trial registration number: NCT03317171 Retrospectively registered 25/10/2017.
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- 2019
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27. Creatine kinase elevation: a neglected clue to the diagnosis of polymyositis. A case report.
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Sitzia C, Sansone VA, and Corsi Romanelli MM
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- Electromyography, Humans, Immunoglobulins, Intravenous therapeutic use, Male, Middle Aged, Muscle Strength, Muscle, Skeletal pathology, Myoglobin blood, Polymyositis drug therapy, Steroids therapeutic use, Creatine Kinase blood, Polymyositis diagnosis
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- 2019
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28. Fibrosis Rescue Improves Cardiac Function in Dystrophin-Deficient Mice and Duchenne Patient-Specific Cardiomyocytes by Immunoproteasome Modulation.
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Farini A, Gowran A, Bella P, Sitzia C, Scopece A, Castiglioni E, Rovina D, Nigro P, Villa C, Fortunato F, Comi GP, Milano G, Pompilio G, and Torrente Y
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- Animals, Fibrosis, Humans, Induced Pluripotent Stem Cells immunology, Induced Pluripotent Stem Cells pathology, Male, Mice, Mice, Inbred mdx, Cardiomyopathies immunology, Cardiomyopathies pathology, Muscular Dystrophy, Duchenne immunology, Muscular Dystrophy, Duchenne pathology, Myocytes, Cardiac immunology, Myocytes, Cardiac pathology, Proteasome Endopeptidase Complex immunology
- Abstract
Patients affected by Duchenne muscular dystrophy (DMD) develop a progressive dilated cardiomyopathy characterized by inflammatory cell infiltration, necrosis, and cardiac fibrosis. Standard treatments consider the use of β-blockers and angiotensin-converting enzyme inhibitors that are symptomatic and unspecific toward DMD disease. Medications that target DMD cardiac fibrosis are in the early stages of development. We found immunoproteasome dysregulation in affected hearts of mdx mice (murine animal model of DMD) and cardiomyocytes derived from induced pluripotent stem cells of patients with DMD. Interestingly, immunoproteasome inhibition ameliorated cardiomyopathy in mdx mice and reduced the development of cardiac fibrosis. Establishing the immunoproteasome inhibition-dependent cardioprotective role suggests the possibility of modulating the immunoproteasome as new and clinically relevant treatment to rescue dilated cardiomyopathy in patients with DMD., (Copyright © 2019 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)
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- 2019
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29. Therapeutic Potential of Immunoproteasome Inhibition in Duchenne Muscular Dystrophy.
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Farini A, Sitzia C, Cassani B, Cassinelli L, Rigoni R, Colleoni F, Fusco N, Gatti S, Bella P, Villa C, Napolitano F, Maiavacca R, Bosari S, Villa A, and Torrente Y
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- Animals, Cell Differentiation, Disease Models, Animal, Genetic Therapy, Mice, Mice, Inbred mdx, Muscular Dystrophy, Duchenne immunology, Oligopeptides administration & dosage, Oligopeptides pharmacology, Proteasome Endopeptidase Complex metabolism, Proteasome Inhibitors pharmacology, T-Lymphocytes drug effects, T-Lymphocytes physiology, Immunoproteins antagonists & inhibitors, Muscular Dystrophy, Duchenne drug therapy, Proteasome Inhibitors administration & dosage, T-Lymphocytes immunology
- Abstract
Duchenne muscular dystrophy is an inherited fatal genetic disease characterized by mutations in dystrophin gene, causing membrane fragility leading to myofiber necrosis and inflammatory cell recruitment in dystrophic muscles. The resulting environment enriched in proinflammatory cytokines, like IFN-γ and TNF-α, determines the transformation of myofiber constitutive proteasome into the immunoproteasome, a multisubunit complex involved in the activation of cell-mediate immunity. This event has a fundamental role in producing peptides for antigen presentation by MHC class I, for the immune response and also for cytokine production and T-cell differentiation. Here, we characterized for the first time the presence of T-lymphocytes activated against revertant dystrophin epitopes, in the animal model of Duchenne muscular dystrophy, the mdx mice. Moreover, we specifically blocked i-proteasome subunit LMP7, which was up-regulated in dystrophic skeletal muscles, and we demonstrated the rescue of the dystrophin expression and the amelioration of the dystrophic phenotype. The i-proteasome blocking lowered myofiber MHC class I expression and self-antigen presentation to T cells, thus reducing the specific antidystrophin T cell response, the muscular cell infiltrate, and proinflammatory cytokine production, together with muscle force recovery. We suggest that i-proteasome inhibition should be considered as new promising therapeutic approach for Duchenne muscular dystrophy pathology.
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- 2016
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30. Adaptive Immune Response Impairs the Efficacy of Autologous Transplantation of Engineered Stem Cells in Dystrophic Dogs.
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Sitzia C, Farini A, Jardim L, Razini P, Belicchi M, Cassinelli L, Villa C, Erratico S, Parolini D, Bella P, da Silva Bizario JC, Garcia L, Dias-Baruffi M, Meregalli M, and Torrente Y
- Subjects
- Animals, Cells, Cultured, Disease Models, Animal, Dogs, Follow-Up Studies, Humans, Muscular Dystrophy, Animal immunology, Stem Cells metabolism, Transplantation, Autologous, Treatment Outcome, AC133 Antigen metabolism, Adaptive Immunity, Muscular Dystrophy, Animal therapy, Stem Cell Transplantation methods
- Abstract
Duchenne muscular dystrophy is the most common genetic muscular dystrophy. It is caused by mutations in the dystrophin gene, leading to absence of muscular dystrophin and to progressive degeneration of skeletal muscle. We have demonstrated that the exon skipping method safely and efficiently brings to the expression of a functional dystrophin in dystrophic CD133+ cells injected scid/mdx mice. Golden Retriever muscular dystrophic (GRMD) dogs represent the best preclinical model of Duchenne muscular dystrophy, mimicking the human pathology in genotypic and phenotypic aspects. Here, we assess the capacity of intra-arterial delivered autologous engineered canine CD133+ cells of restoring dystrophin expression in Golden Retriever muscular dystrophy. This is the first demonstration of five-year follow up study, showing initial clinical amelioration followed by stabilization in mild and severe affected Golden Retriever muscular dystrophy dogs. The occurrence of T-cell response in three Golden Retriever muscular dystrophy dogs, consistent with a memory response boosted by the exon skipped-dystrophin protein, suggests an adaptive immune response against dystrophin.
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- 2016
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31. Longitudinal MRI quantification of muscle degeneration in Duchenne muscular dystrophy.
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Godi C, Ambrosi A, Nicastro F, Previtali SC, Santarosa C, Napolitano S, Iadanza A, Scarlato M, Natali Sora MG, Tettamanti A, Gerevini S, Cicalese MP, Sitzia C, Venturini M, Falini A, Gatti R, Ciceri F, Cossu G, Torrente Y, and Politi LS
- Abstract
Objective: The aim of this study was to evaluate the usefulness of magnetic resonance imaging (MRI) in detecting the progression of Duchenne muscular dystrophy (DMD) by quantification of fat infiltration (FI) and muscle volume index (MVI, a residual-to-total muscle volume ratio)., Methods: Twenty-six patients (baseline age: 5-12 years) with genetically proven DMD were longitudinally analyzed with lower limb 3T MRI, force measurements, and functional tests (Gowers, 10-m time, North Star Ambulatory Assessment, 6-min walking test). Five age-matched controls were also examined, with a total of 85 MRI studies. Semiquantitative (scores) and quantitative MRI (qMRI) analyses (signal intensity ratio - SIR, lower limb MVI, and individual muscle MVI) were carried out. Permutation and regression analyses according to both age and functional test-outcomes were calculated. Age-related quantitative reference curves of SIRs and MVIs were generated., Results: FI was present on glutei and adductor magnus in all patients since the age of 5, with a proximal-to-distal progression and selective sparing of sartorius and gracilis. Patients' qMRI measures were significantly different from controls' and among age classes. qMRI were more sensitive than force measurements and functional tests in assessing disease progression, allowing quantification also after loss of ambulation. Age-related curves with percentile values were calculated for SIRs and MVIs, to provide a reference background for future experimental therapy trials. SIRs and MVIs significantly correlated with all clinical measures, and could reliably predict functional outcomes and loss of ambulation., Interpretations: qMRI-based indexes are sensitive measures that can track the progression of DMD and represent a valuable tool for follow-up and clinical studies.
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- 2016
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32. Exome sequencing identifies variants in two genes encoding the LIM-proteins NRAP and FHL1 in an Italian patient with BAG3 myofibrillar myopathy.
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D'Avila F, Meregalli M, Lupoli S, Barcella M, Orro A, De Santis F, Sitzia C, Farini A, D'Ursi P, Erratico S, Cristofani R, Milanesi L, Braga D, Cusi D, Poletti A, Barlassina C, and Torrente Y
- Subjects
- Adult, Exome, Female, Humans, Italy, Transfection, Adaptor Proteins, Signal Transducing genetics, Apoptosis Regulatory Proteins genetics, Intracellular Signaling Peptides and Proteins genetics, LIM Domain Proteins genetics, Muscle Proteins genetics, Myopathies, Structural, Congenital genetics
- Abstract
Myofibrillar myopathies (MFMs) are genetically heterogeneous dystrophies characterized by the disintegration of Z-disks and myofibrils and are associated with mutations in genes encoding Z-disk or Z-disk-related proteins. The c.626 C > T (p.P209L) mutation in the BAG3 gene has been described as causative of a subtype of MFM. We report a sporadic case of a 26-year-old Italian woman, affected by MFM with axonal neuropathy, cardiomyopathy, rigid spine, who carries the c.626 C > T mutation in the BAG3 gene. The patient and her non-consanguineous healthy parents and brother were studied with whole exome sequencing (WES) to further investigate the genetic basis of this complex phenotype. In the patient, we found that the BAG3 mutation is associated with variants in the NRAP and FHL1 genes that encode muscle-specific, LIM domain containing proteins. Quantitative real time PCR, immunohistochemistry and Western blot analysis of the patient's muscular biopsy showed the absence of NRAP expression and FHL1 accumulation in aggregates in the affected skeletal muscle tissue. Molecular dynamic analysis of the mutated FHL1 domain showed a modification in its surface charge, which could affect its capability to bind its target proteins. To our knowledge this is the first study reporting, in a BAG3 MFM, the simultaneous presence of genetic variants in the BAG3 and FHL1 genes (previously described as independently associated with MFMs) and linking the NRAP gene to MFM for the first time., Competing Interests: The authors declare that they have no competing interests.
- Published
- 2016
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33. Inositol 1,4,5-trisphosphate (IP3)-dependent Ca2+ signaling mediates delayed myogenesis in Duchenne muscular dystrophy fetal muscle.
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Farini A, Sitzia C, Cassinelli L, Colleoni F, Parolini D, Giovanella U, Maciotta S, Colombo A, Meregalli M, and Torrente Y
- Subjects
- Animals, Biomarkers metabolism, Biopsy, Calcium metabolism, Calcium Channels metabolism, Fetus pathology, Gene Expression Regulation, Developmental, Mice, Inbred C57BL, Mice, Inbred mdx, Models, Biological, Muscle Fibers, Skeletal metabolism, Muscle Fibers, Skeletal pathology, Muscle, Skeletal metabolism, Muscle, Skeletal pathology, Muscular Dystrophy, Animal metabolism, Muscular Dystrophy, Animal pathology, Muscular Dystrophy, Duchenne pathology, PAX7 Transcription Factor metabolism, Protein Kinase C-alpha metabolism, Calcium Signaling, Fetus metabolism, Inositol 1,4,5-Trisphosphate metabolism, Muscle Development, Muscle, Skeletal embryology, Muscular Dystrophy, Duchenne embryology, Muscular Dystrophy, Duchenne metabolism
- Abstract
Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disorder characterized by muscle wasting and premature death. The defective gene is dystrophin, a structural protein, absence of which causes membrane fragility and myofiber necrosis. Several lines of evidence showed that in adult DMD patients dystrophin is involved in signaling pathways that regulate calcium homeostasis and differentiation programs. However, secondary aspects of the disease, such as inflammation and fibrosis development, might represent a bias in the analysis. Because fetal muscle is not influenced by gravity and does not suffer from mechanical load and/or inflammation, we investigated 12-week-old fetal DMD skeletal muscles, highlighting for the first time early alterations in signaling pathways mediated by the absence of dystrophin itself. We found that PLC/IP3/IP3R/Ryr1/Ca(2+) signaling is widely active in fetal DMD skeletal muscles and, through the calcium-dependent PKCα protein, exerts a fundamental regulatory role in delaying myogenesis and in myofiber commitment. These data provide new insights into the origin of DMD pathology during muscle development., (© 2016. Published by The Company of Biologists Ltd.)
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- 2016
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34. Stem Cell-Mediated Exon Skipping of the Dystrophin Gene by the Bystander Effect.
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Meregalli M, Farini A, Sitzia C, Beley C, Razini P, Cassinelli L, Colleoni F, Frattini P, Santo N, Galbiati E, Prosperi D, Tavelli A, Belicchi M, Garcia L, and Torrente Y
- Subjects
- Animals, Bystander Effect physiology, Cells, Cultured, Dystrophin biosynthesis, Exons genetics, Humans, Mice, Mice, SCID, Muscle, Skeletal pathology, Oligonucleotides, Antisense genetics, RNA Splicing genetics, Dystrophin genetics, Genetic Therapy methods, Muscular Dystrophy, Duchenne genetics, Muscular Dystrophy, Duchenne therapy, Stem Cells cytology
- Abstract
Duchenne muscular dystrophy (DMD) is characterized by the loss of a functional dystrophin protein; the muscles of DMD patients progressively degenerate as a result of mechanical stress during contractions, and the condition eventually leads to premature death. By means antisense oligonucleotides (AONs), it is possible to modulate pre-mRNA splicing eliminating mutated exons and restoring dystrophin open reading frame. To overcome the hurdles in using AONs for therapeutic interventions, we exerted engineered human DMD stem cells with a lentivirus, which permanently and efficiently delivered the cloned AONs. Here we describe for the first time the exosome-mediated release of AONs from engineered human DMD CD133+ stem cells allowing the rescue of murine dystrophin expression. Finally, upon release, AONs could be internalized by host cells suggesting a potential role of exosomes acting as vesicular carriers for DMD gene therapy.
- Published
- 2015
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35. Improvement of endurance of DMD animal model using natural polyphenols.
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Sitzia C, Farini A, Colleoni F, Fortunato F, Razini P, Erratico S, Tavelli A, Fabrizi F, Belicchi M, Meregalli M, Comi G, and Torrente Y
- Subjects
- Animals, Cells, Cultured, Disease Models, Animal, Fibrosis drug therapy, Fibrosis metabolism, Mice, Mice, Inbred C57BL, Muscle, Skeletal metabolism, Muscular Dystrophy, Animal metabolism, Muscular Dystrophy, Duchenne metabolism, Myofibrils drug effects, Myofibrils metabolism, NF-kappa B metabolism, Nitric Oxide metabolism, Muscle, Skeletal drug effects, Muscular Dystrophy, Animal drug therapy, Muscular Dystrophy, Duchenne drug therapy, Polyphenols pharmacology
- Abstract
Duchenne muscular dystrophy (DMD), the most common form of muscular dystrophy, is characterized by muscular wasting caused by dystrophin deficiency that ultimately ends in force reduction and premature death. In addition to primary genetic defect, several mechanisms contribute to DMD pathogenesis. Recently, antioxidant supplementation was shown to be effective in the treatment of multiple diseases including muscular dystrophy. Different mechanisms were hypothesized such as reduced hydroxyl radicals, nuclear factor-κB deactivation, and NO protection from inactivation. Following these promising evidences, we investigated the effect of the administration of a mix of dietary natural polyphenols (ProAbe) on dystrophic mdx mice in terms of muscular architecture and functionality. We observed a reduction of muscle fibrosis deposition and myofiber necrosis together with an amelioration of vascularization. More importantly, the recovery of the morphological features of dystrophic muscle leads to an improvement of the endurance of treated dystrophic mice. Our data confirmed that ProAbe-based diet may represent a strategy to coadjuvate the treatment of DMD.
- Published
- 2015
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36. Advancements in stem cells treatment of skeletal muscle wasting.
- Author
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Meregalli M, Farini A, Sitzia C, and Torrente Y
- Abstract
Muscular dystrophies (MDs) are a heterogeneous group of inherited disorders, in which progressive muscle wasting and weakness is often associated with exhaustion of muscle regeneration potential. Although physiological properties of skeletal muscle tissue are now well known, no treatments are effective for these diseases. Muscle regeneration was attempted by means transplantation of myogenic cells (from myoblast to embryonic stem cells) and also by interfering with the malignant processes that originate in pathological tissues, such as uncontrolled fibrosis and inflammation. Taking into account the advances in the isolation of new subpopulation of stem cells and in the creation of artificial stem cell niches, we discuss how these emerging technologies offer great promises for therapeutic approaches to muscle diseases and muscle wasting associated with aging.
- Published
- 2014
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37. Clinical applications of mesenchymal stem cells in chronic diseases.
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Farini A, Sitzia C, Erratico S, Meregalli M, and Torrente Y
- Abstract
Extraordinary progress in understanding several key features of stem cells has been made in the last ten years, including definition of the niche, and identification of signals regulating mobilization and homing as well as partial understanding of the mechanisms controlling self-renewal, commitment, and differentiation. This progress produced invaluable tools for the development of rational cell therapy protocols that have yielded positive results in preclinical models of genetic and acquired diseases and, in several cases, have entered clinical experimentation with positive outcome. Adult mesenchymal stem cells (MSCs) are nonhematopoietic cells with multilineage potential to differentiate into various tissues of mesodermal origin. They can be isolated from bone marrow and other tissues and have the capacity to extensively proliferate in vitro. Moreover, MSCs have also been shown to produce anti-inflammatory molecules which can modulate humoral and cellular immune responses. Considering their regenerative potential and immunoregulatory effect, MSC therapy is a promising tool in the treatment of degenerative, inflammatory, and autoimmune diseases. It is obvious that much work remains to be done to increase our knowledge of the mechanisms regulating development, homeostasis, and tissue repair and thus to provide new tools to implement the efficacy of cell therapy trials.
- Published
- 2014
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38. Influence of immune responses in gene/stem cell therapies for muscular dystrophies.
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Farini A, Sitzia C, Erratico S, Meregalli M, and Torrente Y
- Subjects
- Extracellular Matrix immunology, Extracellular Matrix metabolism, Genetic Therapy, Humans, Muscular Dystrophies immunology, Muscular Dystrophies pathology, Mutation, Stem Cells cytology, Cell- and Tissue-Based Therapy, Immunity, Innate, Muscular Dystrophies therapy, Stem Cells immunology
- Abstract
Muscular dystrophies (MDs) are a heterogeneous group of diseases, caused by mutations in different components of sarcolemma, extracellular matrix, or enzymes. Inflammation and innate or adaptive immune response activation are prominent features of MDs. Various therapies under development are directed toward rescuing the dystrophic muscle damage using gene transfer or cell therapy. Here we discussed current knowledge about involvement of immune system responses to experimental therapies in MDs.
- Published
- 2014
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39. Full-length dysferlin expression driven by engineered human dystrophic blood derived CD133+ stem cells.
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Meregalli M, Navarro C, Sitzia C, Farini A, Montani E, Wein N, Razini P, Beley C, Cassinelli L, Parolini D, Belicchi M, Parazzoli D, Garcia L, and Torrente Y
- Subjects
- AC133 Antigen, Adult, Animals, Blotting, Western, Cells, Cultured, Distal Myopathies genetics, Distal Myopathies pathology, Dysferlin, Fluorescent Antibody Technique, Humans, Immunoenzyme Techniques, In Situ Hybridization, Fluorescence, Injections, Intramuscular, Lentivirus genetics, Male, Membrane Proteins genetics, Mice, Mice, Inbred A, Mice, SCID, Muscle Proteins genetics, Muscle, Skeletal injuries, Muscle, Skeletal metabolism, Muscular Atrophy genetics, Muscular Atrophy pathology, Mutation genetics, RNA, Messenger genetics, Real-Time Polymerase Chain Reaction, Reverse Transcriptase Polymerase Chain Reaction, Stem Cells metabolism, Antigens, CD metabolism, Distal Myopathies therapy, Glycoproteins metabolism, Membrane Proteins metabolism, Muscle Proteins metabolism, Muscle, Skeletal pathology, Muscular Atrophy therapy, Oligonucleotides, Antisense pharmacology, Peptides metabolism, Stem Cell Transplantation, Stem Cells cytology
- Abstract
The protein dysferlin is abundantly expressed in skeletal and cardiac muscles, where its main function is membrane repair. Mutations in the dysferlin gene are involved in two autosomal recessive muscular dystrophies: Miyoshi myopathy and limb-girdle muscular dystrophy type 2B. Development of effective therapies remains a great challenge. Strategies to repair the dysferlin gene by skipping mutated exons, using antisense oligonucleotides (AONs), may be suitable only for a subset of mutations, while cell and gene therapy can be extended to all mutations. AON-treated blood-derived CD133+ stem cells isolated from patients with Miyoshi myopathy led to partial dysferlin reconstitution in vitro but failed to express dysferlin after intramuscular transplantation into scid/blAJ dysferlin null mice. We thus extended these experiments producing the full-length dysferlin mediated by a lentiviral vector in blood-derived CD133+ stem cells isolated from the same patients. Transplantation of engineered blood-derived CD133+ stem cells into scid/blAJ mice resulted in sufficient dysferlin expression to correct functional deficits in skeletal muscle membrane repair. Our data suggest for the first time that lentivirus-mediated delivery of full-length dysferlin in stem cells isolated from Miyoshi myopathy patients could represent an alternative therapeutic approach for treatment of dysferlinopathies., (© 2013 FEBS.)
- Published
- 2013
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40. Perspectives of stem cell therapy in Duchenne muscular dystrophy.
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Meregalli M, Farini A, Belicchi M, Parolini D, Cassinelli L, Razini P, Sitzia C, and Torrente Y
- Subjects
- Animals, Humans, Muscular Dystrophy, Duchenne metabolism, Stem Cells metabolism, Cell- and Tissue-Based Therapy, Muscular Dystrophy, Duchenne therapy, Stem Cell Transplantation, Stem Cells cytology
- Abstract
Muscular dystrophies are heritable and heterogeneous neuromuscular disorders characterized by the primary wasting of skeletal muscle, usually caused by mutations in the proteins forming the link between the cytoskeleton and the basal lamina. As a result of mutations in the dystrophin gene, Duchenne muscular dystrophy patients suffer from progressive muscle atrophy and an exhaustion of muscular regenerative capacity. No efficient therapies are available. The evidence that adult stem cells were capable of participating in the regeneration of more than their resident organ led to the development of potential stem cell treatments for degenerative disorder. In the present review, we describe the different types of myogenic stem cells and their possible use for the progression of cell therapy in Duchenne muscular dystrophy., (© 2012 The Authors Journal compilation © 2012 FEBS.)
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- 2013
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41. Expression of CD20 reveals a new store-operated calcium entry modulator in skeletal muscle.
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Parolini D, Cassinelli L, Razini P, Sitzia C, Tonna N, Erratico S, Colleoni F, Angeloni V, Maffioli E, Farini A, Maciotta S, Porretti L, Belicchi M, Bianco F, Tedeschi G, Meregalli M, and Torrente Y
- Subjects
- Amino Acid Sequence, Animals, Antibodies pharmacology, Antigens, CD20 chemistry, Antigens, CD20 genetics, Antigens, CD20 immunology, Cell Line, Cell Membrane metabolism, Cell Proliferation drug effects, Cell Survival drug effects, Gene Knockdown Techniques, Humans, Mice, Mice, Inbred C57BL, Mice, SCID, Molecular Sequence Data, Muscle, Skeletal cytology, Muscle, Skeletal metabolism, Myoblasts metabolism, RNA Interference, Antigens, CD20 metabolism, Calcium Signaling drug effects, Gene Expression, Muscle Fibers, Skeletal metabolism
- Abstract
Among the scarce available data about the biological role of the membrane protein CD20, there is some evidence that this protein functions as a store-operated Ca(2+) channel and/or regulates transmembrane Ca(2+) trafficking. Recent findings indicate that store-operated Ca(2+) entry (SOCE) plays a central role in skeletal muscle function and development, but there remain a number of unresolved issues relating to SOCE modulation in this tissue. Here we describe CD20 expression in skeletal muscle, verifying its membrane localization in myoblasts and adult muscle fibers. Additionally, we show that inhibition of CD20 through antibody binding or gene silencing resulted in specific impairment of SOCE in C2C12 myoblasts. Our results provide novel insights into the CD20 expression pattern, and suggest that functional CD20 is required for SOCE to consistently occur in C2C12 myoblasts. These findings may contribute to future identification of mechanisms and molecules involved in the fine regulation of store-operated Ca(2+) entry in skeletal muscle., (Copyright © 2012 Elsevier Ltd. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
42. Absence of T and B lymphocytes modulates dystrophic features in dysferlin deficient animal model.
- Author
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Farini A, Sitzia C, Navarro C, D'Antona G, Belicchi M, Parolini D, Del Fraro G, Razini P, Bottinelli R, Meregalli M, and Torrente Y
- Subjects
- Animals, Complement Membrane Attack Complex metabolism, Disease Models, Animal, Dysferlin, Dystrophin metabolism, Endothelial Cells pathology, Female, Hybridization, Genetic, In Vitro Techniques, Inflammation, Laminin metabolism, Macrophages pathology, Male, Membrane Proteins genetics, Membrane Proteins metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, SCID, Muscle Contraction, Muscle, Skeletal blood supply, Muscle, Skeletal metabolism, Muscle, Skeletal pathology, Muscular Dystrophy, Animal metabolism, Regeneration, Sarcoglycans metabolism, Sarcolemma genetics, Sarcolemma metabolism, Sarcolemma pathology, B-Lymphocytes pathology, Membrane Proteins deficiency, Muscle, Skeletal physiopathology, Muscular Dystrophy, Animal pathology, T-Lymphocytes pathology
- Abstract
Dysferlin mutations cause muscular dystrophy (dysferlinopathy) characterized by adult onset muscle weakness, high serum creatine kinase levels, attenuation of muscle regeneration and a prominent inflammatory infiltrate. In order to verify the role of lymphocytes and immune cells on this disease, we generated the Scid/A/J transgenic mice and compared these animals with the age-matched A/J mice. The absence of T and B lymphocytes in this animal model of dysferlinopathy resulted in an improvement of the muscle regeneration. Scid/A/J mice showed increased specific force in the myosin heavy chain 2A-expressing fibers of the diaphragm and abdominal muscles. Moreover, a partial reduction in complement deposition was observed together with a diminution in pro-inflammatory M1 macrophages. Consistent with this model, T and B lymphocytes seem to have a role in the muscle damaging immune response. The knowledge of the involvement of immune system in the development of dysferlinopathies could represent an important tool for their rescuing. By studying Scid/blAJ mice, we showed that it could be possible to modulate the pathological symptoms of these diseases by interfering with different components of the immune system., (Copyright © 2012 Elsevier Inc. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
43. NEU3 sialidase strictly modulates GM3 levels in skeletal myoblasts C2C12 thus favoring their differentiation and protecting them from apoptosis.
- Author
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Anastasia L, Papini N, Colazzo F, Palazzolo G, Tringali C, Dileo L, Piccoli M, Conforti E, Sitzia C, Monti E, Sampaolesi M, Tettamanti G, and Venerando B
- Subjects
- Animals, Cell Differentiation, Cell Line, Down-Regulation, Epidermal Growth Factor antagonists & inhibitors, Epidermal Growth Factor metabolism, ErbB Receptors metabolism, Gene Silencing, Hydrolysis, Mice, Models, Chemical, Sphingolipids metabolism, Apoptosis, G(M3) Ganglioside metabolism, Muscle, Skeletal metabolism, Neuraminidase metabolism
- Abstract
Membrane-bound sialidase NEU3, often referred to as the "ganglioside sialidase," has a critical regulatory function on the sialoglycosphingolipid pattern of the cell membrane, with an anti-apoptotic function, especially in cancer cells. Although other sialidases have been shown to be involved in skeletal muscle differentiation, the role of NEU3 had yet to be disclosed. Herein we report that NEU3 plays a key role in skeletal muscle differentiation by strictly modulating the ganglioside content of adjacent cells, with special regard to GM3. Induced down-regulation of NEU3 in murine C2C12 myoblasts, even when partial, totally inhibits their capability to differentiate by increasing the GM3 level above a critical point, which causes epidermal growth factor receptor inhibition (and ultimately its down-regulation) and an higher responsiveness of myoblasts to the apoptotic stimuli.
- Published
- 2008
- Full Text
- View/download PDF
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