19 results on '"Catizone, Angela"'
Search Results
2. Microgravity-Induced Metabolic Response in 2D and 3D TCam-2 Cell Cultures
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Morabito, Caterina, primary, Guarnieri, Simone, additional, Berardini, Marika, additional, Gesualdi, Luisa, additional, Ferranti, Francesca, additional, Reale, Anna, additional, Ricci, Giulia, additional, Catizone, Angela, additional, and Mariggiò, Maria A., additional
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- 2023
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3. Microgravity Exposure Alterations of Cellular Junctions Proteins in TCam-2 Cells: Localization and Interaction
- Author
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Berardini, Marika, primary, Gesualdi, Luisa, additional, Ferranti, Francesca, additional, Mariggiò, Maria Addolorata, additional, Morabito, Caterina, additional, Guarnieri, Simone, additional, Ricci, Giulia, additional, and Catizone, Angela, additional
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- 2023
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4. Simulated Microgravity Exposure Induces Antioxidant Barrier Deregulation and Mitochondria Enlargement in TCam-2 Cell Spheroids.
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Berardini, Marika, Gesualdi, Luisa, Morabito, Caterina, Ferranti, Francesca, Reale, Anna, Zampieri, Michele, Karpach, Katsiaryna, Tinari, Antonella, Bertuccini, Lucia, Guarnieri, Simone, Catizone, Angela, Mariggiò, Maria A., and Ricci, Giulia
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REDUCED gravity environments ,MITOCHONDRIA ,GRAVITATION ,CELL culture ,GERM cells ,REACTIVE oxygen species ,PLANT mitochondria ,HOMEOSTASIS - Abstract
One of the hallmarks of microgravity-induced effects in several cellular models is represented by the alteration of oxidative balance with the consequent accumulation of reactive oxygen species (ROS). It is well known that male germ cells are sensitive to oxidative stress and to changes in gravitational force, even though published data on germ cell models are scarce. We previously studied the effects of simulated microgravity (s-microgravity) on a 2D cultured TCam-2 seminoma-derived cell line, considered the only human cell line available to study in vitro mitotically active human male germ cells. In this study, we used a corresponding TCam-2 3D cell culture model that mimics cell–cell contacts in organ tissue to test the possible effects induced by s-microgravity exposure. TCam-2 cell spheroids were cultured for 24 h under unitary gravity (Ctr) or s-microgravity conditions, the latter obtained using a random positioning machine (RPM). A significant increase in intracellular ROS and mitochondria superoxide anion levels was observed after RPM exposure. In line with these results, a trend of protein and lipid oxidation increase and increased pCAMKII expression levels were observed after RPM exposure. The ultrastructural analysis via transmission electron microscopy revealed that RPM-exposed mitochondria appeared enlarged and, even if seldom, disrupted. Notably, even the expression of the main enzymes involved in the redox homeostasis appears modulated by RPM exposure in a compensatory way, with GPX1, NCF1, and CYBB being downregulated, whereas NOX4 and HMOX1 are upregulated. Interestingly, HMOX1 is involved in the heme catabolism of mitochondria cytochromes, and therefore the positive modulation of this marker can be associated with the observed mitochondria alteration. Altogether, these data demonstrate TCam-2 spheroid sensitivity to acute s-microgravity exposure and indicate the capability of these cells to trigger compensatory mechanisms that allow them to overcome the exposure to altered gravitational force. [ABSTRACT FROM AUTHOR]
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- 2023
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5. ERK Signaling Pathway Is Constitutively Active in NT2D1 Non-Seminoma Cells and Its Inhibition Impairs Basal and HGF-Activated Cell Proliferation.
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Gesualdi, Luisa, Berardini, Marika, Scicchitano, Bianca Maria, Castaldo, Clotilde, Bizzarri, Mariano, Filippini, Antonio, Riccioli, Anna, Schiraldi, Chiara, Ferranti, Francesca, Liguoro, Domenico, Mancini, Rita, Ricci, Giulia, and Catizone, Angela
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CELL proliferation ,PHOSPHATIDYLINOSITOL 3-kinases ,PROTEIN kinases ,GERM cell tumors ,MITOGEN-activated protein kinases ,CELLULAR signal transduction - Abstract
c-MET/hepatocyte growth factor (HGF) system deregulation is a well-known feature of malignancy in several solid tumors, and for this reason this system and its pathway have been considered as potential targets for therapeutic purposes. In previous manuscripts we reported c-MET/HGF expression and the role in testicular germ cell tumors (TGCTs) derived cell lines. We demonstrated the key role of c-Src and phosphatidylinositol 3-kinase (PI3K)/AKT adaptors in the HGF-dependent malignant behavior of the embryonal carcinoma cell line NT2D1, finding that the inhibition of these onco-adaptor proteins abrogates HGF triggered responses such as proliferation, migration, and invasion. Expanding on these previous studies, herein we investigated the role of mitogen-activated protein kinase (MAPK)/extracellular signal regulated kinase (ERK) pathways in the HGF-dependent and HGF-independent NT2D1 cells biological responses. To inhibit MAPK/ERK pathways we chose a pharmacological approach, by using U0126 inhibitor, and we analyzed cell proliferation, collective migration, and chemotaxis. The administration of U0126 together with HGF reverts the HGF-dependent activation of cell proliferation but, surprisingly, does not exert the same effect on NT2D1 cell migration. In addition, we found that the use of U0126 alone significantly promotes the acquisition of NT2D1 «migrating phenotype», while collective migration of NT2D1 cells was stimulated. Notably, the inhibition of ERK activation in the absence of HGF stimulation resulted in the activation of the AKT-mediated pathway, and this let us speculate that the paradoxical effects obtained by using U0126, which are the increase of collective migration and the acquisition of partial epithelium–mesenchyme transition (pEMT), are the result of compensatory pathways activation. These data highlight how the specific response to pathway inhibitors, should be investigated in depth before setting up therapy. [ABSTRACT FROM AUTHOR]
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- 2023
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6. Myo-Inositol Reverses TGF-β1-Induced EMT in MCF-10A Non-Tumorigenic Breast Cells.
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Monti, Noemi, Dinicola, Simona, Querqui, Alessandro, Fabrizi, Gianmarco, Fedeli, Valeria, Gesualdi, Luisa, Catizone, Angela, Unfer, Vittorio, and Bizzarri, Mariano
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TRANSFORMING growth factors-beta ,REVERSE transcriptase polymerase chain reaction ,ANIMAL experimentation ,CELL lines ,EPITHELIAL cells ,BREAST tumors ,INOSITOL - Abstract
Simple Summary: Inflammatory conditions can enact the emergence of cancer, especially by promoting an epithelial-mesenchymal transition (EMT). Furthermore, EMT is a critical requirement for the dissemination of cancerous cells. The discovery of pharmacological agents able to inhibit EMT has become a critical issue in recent times. Herein we demonstrate that myo-inositol (myo-Ins), can efficiently rescue normal breast cells committed to an inflammatory phenotype upon the addition of the inflammatory TGF-β1 stimulus. Myo-Ins was able to almost completely inhibit the resulting invasive–migrating phenotype, namely by reverting EMT. A critical step is the reconstitution of proper E-cadherin-based cell-to-cell junctions, which are instrumental in the recovery of a tissue-like structure. Moreover, myo-Ins re-established a normal gene expression pattern, while normalizing the microenvironment by reducing collagen and metalloproteinase release. These results highlight the relevance of inflammation in promoting a precancerous state and provide useful new perspectives in cancer treatment and prevention with natural compounds. Epithelial-Mesenchymal Transition (EMT), triggered by external and internal cues in several physiological and pathological conditions, elicits the transformation of epithelial cells into a mesenchymal-like phenotype. During EMT, epithelial cells lose cell-to-cell contact and acquire unusual motility/invasive capabilities. The associated architectural and functional changes destabilize the epithelial layer consistency, allowing cells to migrate and invade the surrounding tissues. EMT is a critical step in the progression of inflammation and cancer, often sustained by a main driving factor as the transforming growth factor-β1 (TGF-β1). Antagonizing EMT has recently gained momentum as an attractive issue in cancer treatment and metastasis prevention. Herein, we demonstrate the capability of myo-inositol (myo-Ins) to revert the EMT process induced by TGF-β1 on MCF-10A breast cells. Upon TGF-β1 addition, cells underwent a dramatic phenotypic transformation, as witnessed by structural (disappearance of the E-cadherin–β-catenin complexes and the emergence of a mesenchymal shape) and molecular modifications (increase in N-cadherin, Snai1, and vimentin), including the release of increased collagen and fibronectin. However, following myo-Ins, those changes were almost completely reverted. Inositol promotes the reconstitution of E-cadherin–β-catenin complexes, decreasing the expression of genes involved in EMT, while promoting the re-expression of epithelial genes (keratin-18 and E-cadherin). Noticeably, myo-Ins efficiently inhibits the invasiveness and migrating capability of TGF-β1 treated cells, also reducing the release of metalloproteinase (MMP-9) altogether with collagen synthesis, allowing for the re-establishment of appropriate cell-to-cell junctions, ultimately leading the cell layer back towards a more compact state. Inositol effects were nullified by previous treatment with an siRNA construct to inhibit CDH1 transcripts and, hence, E-cadherin synthesis. This finding suggests that the reconstitution of E-cadherin complexes is an irreplaceable step in the inositol-induced reversion of EMT. Overall, such a result advocates for the useful role of myo-Ins in cancer treatment. [ABSTRACT FROM AUTHOR]
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- 2023
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7. The Role of Inositols in the Hyperandrogenic Phenotypes of PCOS: A Re-Reading of Larner's Results.
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Fedeli, Valeria, Catizone, Angela, Querqui, Alessandro, Unfer, Vittorio, and Bizzarri, Mariano
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ANDROGEN receptors , *POLYCYSTIC ovary syndrome , *PHENOTYPES , *OVARIAN cysts , *MENSTRUATION disorders , *INSULIN resistance - Abstract
Polycystic ovarian syndrome (PCOS) is the most common endocrinological disorder in women, in which, besides chronic anovulation/oligomenorrhea and ovarian cysts, hyperandrogenism plays a critical role in a large fraction of subjects. Inositol isomers—myo-Inositol and D-Chiro-Inositol—have recently been pharmacologically effective in managing many PCOS symptoms while rescuing ovarian fertility. However, some disappointing clinical results prompted the reconsideration of their specific biological functions. Surprisingly, D-Chiro-Ins stimulates androgen synthesis and decreases the ovarian estrogen pathway; on the contrary, myo-Ins activates FSH response and aromatase activity, finally mitigating ovarian hyperandrogenism. However, when the two isomers are given in association—according to the physiological ratio of 40:1—patients could benefit from myo-Ins enhanced FSH and estrogen responsiveness, while taking advantage of the insulin-sensitizing effects displayed mostly by D-Chiro-Ins. We need not postulate insulin resistance to explain PCOS pathogenesis, given that insulin hypersensitivity is likely a shared feature of PCOS ovaries. Indeed, even in the presence of physiological insulin stimulation, the PCOS ovary synthesizes D-Chiro-Ins four times more than that measured in control theca cells. The increased D-Chiro-Ins within the ovary is detrimental in preserving steroidogenic control, and this failure can easily explain why treatment strategies based upon high D-Chiro-Ins have been recognized as poorly effective. Within this perspective, two factors emerge as major determinants in PCOS: hyperandrogenism and reduced aromatase expression. Therefore, PCOS could no longer be considered a disease only due to increased androgen synthesis without considering the contemporary downregulation of aromatase and FSH receptors. Furthermore, these findings suggest that inositols can be specifically effective only for those PCOS phenotypes featured by hyperandrogenism. [ABSTRACT FROM AUTHOR]
- Published
- 2023
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8. B4GALT1 as a New Biomarker of Idiopathic Pulmonary Fibrosis.
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De Vitis, Claudia, D'Ascanio, Michela, Sacconi, Andrea, Pizzirusso, Dario, Salvati, Valentina, Mancini, Massimiliano, Scafetta, Giorgia, Cirombella, Roberto, Ascenzi, Francesca, Bruschini, Sara, Esposito, Antonella, Castelli, Silvia, Salvucci, Claudia, Teodonio, Leonardo, Sposato, Bruno, Catizone, Angela, Di Napoli, Arianna, Vecchione, Andrea, Ciliberto, Gennaro, and Sciacchitano, Salvatore
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IDIOPATHIC pulmonary fibrosis ,CELL culture ,BIOMARKERS ,HUMAN cell culture ,GENE expression ,RESPIRATORY insufficiency - Abstract
Idiopathic pulmonary fibrosis (IPF) is a disease characterized by progressive scarring of the lung that involves the pulmonary interstitium. The disease may rapidly progress, leading to respiratory failure, and the long-term survival is poor. There are no accurate biomarkers available so far. Our aim was to evaluate the expression of the B4GALT1 in patients with IPF. Analysis of B4GALT1 gene expression was performed in silico on two gene sets, retrieved from the Gene Expression Omnibus database. Expression of B4GALT1 was then evaluated, both at the mRNA and protein levels, on lung specimens obtained from lung biopsies of 4 IPF patients, on one IPF-derived human primary cell and on 11 cases of IPF associated with cancer. In silico re-analysis demonstrated that the B4GALT1 gene was overexpressed in patients and human cell cultures with IPF (p = 0.03). Network analysis demonstrated that B4GALT1 upregulation was correlated with genes belonging to the EMT pathway (p = 0.01). The overexpression of B4GALT1 was observed, both at mRNA and protein levels, in lung biopsies of our four IPF patients and in the IPF-derived human primary cell, in other fibrotic non-lung tissues, and in IPF associated with cancer. In conclusion, our results indicate that B4GALT1 is overexpressed in IPF and could represent a novel marker of this disease. [ABSTRACT FROM AUTHOR]
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- 2022
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9. Radioresistance Mechanisms in Prostate Cancer Cell Lines Surviving Ultra-Hypo-Fractionated EBRT: Implications and Possible Clinical Applications.
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Sideri, Silvia, Petragnano, Francesco, Maggio, Roberto, Petrungaro, Simonetta, Catizone, Angela, Gesualdi, Luisa, De Martino, Viviana, Battafarano, Giulia, Del Fattore, Andrea, Liguoro, Domenico, De Cesaris, Paola, Filippini, Antonio, Marampon, Francesco, and Riccioli, Anna
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ANIMAL experimentation ,RABBITS ,METASTASIS ,CELL proliferation ,DOCETAXEL ,CELL lines ,RADIOTHERAPY ,TUMOR markers ,PROSTATE tumors ,PHENOTYPES - Abstract
Simple Summary: Radiotherapy is an option for curing localized and locally advanced prostate cancer. However, radioresistance can occur, determining treatment failure and poor prognosis. Herein, we developed a model of radio-resistant prostate cancer cells by irradiating the bone metastasis-derived PC3 highly metastatic prostate cancer cell line and the brain-derived moderately metastatic DU-145 prostate cancer cell line, both castration-resistant. Ultra-hypo-fractionated radiotherapy was used, with doses and intervals similar to the ones used in clinical practice. These in vitro models were tested to gain information on the molecular mechanisms used by prostate cancer cells to survive radiation-induced death. Results from bioassays and molecular assays show that in the highly metastatic cells (PC3), the acquired radioresistance—though enhancing clonogenic efficiency, enrichment of cancer stem cells, proliferation rate and migration ability—interestingly results in significantly higher sensitivity to Docetaxel. This behaviour was not observed using the moderately metastatic DU-145 prostate cancer cells. It can be hypothesised that subgroups of patients with highly metastatic prostate cancer could benefit from chemotherapy immediately after the failure of radiotherapy, before a re-challenge with hormonal treatment or other strategies. The use of a higher dose per fraction to overcome the high radioresistance of prostate cancer cells has been unsuccessfully proposed. Herein, we present PC3 and DU-145, castration-resistant prostate cancer cell lines that survived a clinically used ultra-higher dose per fraction, namely, radioresistant PC3 and DU-145 cells (PC3RR and DU-145RR). Compared to PC3, PC3RR showed a higher level of aggressive behaviour, with enhanced clonogenic potential, DNA damage repair, migration ability and cancer stem cell features. Furthermore, compared to PC3, PC3RR more efficiently survived further radiation by increasing proliferation and down-regulating pro-apoptotic proteins. No significant changes of the above parameters were described in DU-145RR, suggesting that different prostate cancer cell lines that survive ultra-higher dose per fraction do not display the same grade of aggressive phenotype. Furthermore, both PC3RR and DU-145RR increased antioxidant enzymes and mesenchymal markers. Our data suggest that different molecular mechanisms could be potential targets for future treatments plans based on sequential strategies and synergistic effects of different modalities, possibly in a patient-tailored fashion. Moreover, PC3RR cells displayed an increase in specific markers involved in bone remodeling, indicating that radiotherapy selects a PC3 population capable of migrating to secondary metastatic sites. Finally, PC3RR cells showed a better sensitivity to Docetaxel as compared to native PC3 cells. This suggests that a subset of patients with castration-resistant metastatic disease could benefit from upfront Docetaxel treatment after the failure of radiotherapy. [ABSTRACT FROM AUTHOR]
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- 2022
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10. Microgravity Induces Transient EMT in Human Keratinocytes by Early Down-Regulation of E-Cadherin and Cell-Adhesion Remodeling.
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Ricci, Giulia, Cucina, Alessandra, Proietti, Sara, Dinicola, Simona, Ferranti, Francesca, Cammarota, Marcella, Filippini, Antonio, Bizzarri, Mariano, and Catizone, Angela
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REDUCED gravity environments ,KERATINOCYTES ,CELL-matrix adhesions ,GRAVITATIONAL fields ,BIOLOGICAL assay ,TREE-rings - Abstract
Featured Application: Studies on the effects of microgravity on cell–cell and cell–matrix interaction may help increase the safety and improve the health of astronauts in space. Changes in cell–matrix and cell-to-cell adhesion patterns are dramatically fostered by the microgravity exposure of living cells. The modification of adhesion properties could promote the emergence of a migrating and invasive phenotype. We previously demonstrated that short exposure to the simulated microgravity of human keratinocytes (HaCaT) promotes an early epithelial–mesenchymal transition (EMT). Herein, we developed this investigation to verify if the cells maintain the acquired invasive phenotype after an extended period of weightlessness exposure. We also evaluated cells' capability in recovering epithelial characteristics when seeded again into a normal gravitational field after short microgravity exposure. We evaluated the ultra-structural junctional features of HaCaT cells by Transmission Electron Microscopy and the distribution pattern of vinculin and E-cadherin by confocal microscopy, observing a rearrangement in cell–cell and cell–matrix interactions. These results are mirrored by data provided by migration and invasion biological assay. Overall, our studies demonstrate that after extended periods of microgravity, HaCaT cells recover an epithelial phenotype by re-establishing E-cadherin-based junctions and cytoskeleton remodeling, both being instrumental in promoting a mesenchymal–epithelial transition (MET). Those findings suggest that cytoskeletal changes noticed during the first weightlessness period have a transitory character, given that they are later reversed and followed by adaptive modifications through which cells miss the acquired mesenchymal phenotype. [ABSTRACT FROM AUTHOR]
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- 2021
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11. Microgravity-Induced Cell-to-Cell Junctional Contacts Are Counteracted by Antioxidant Compounds in TCam-2 Seminoma Cells.
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Catizone, Angela, Morabito, Caterina, Cammarota, Marcella, Schiraldi, Chiara, Corano Scheri, Katia, Ferranti, Francesca, Mariggiò, Maria A., and Ricci, Giulia
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SEMINOMA ,WESTERN immunoblotting ,MICROTUBULES ,GERM cells ,CELL membranes ,CELL metabolism - Abstract
The direct impact of microgravity exposure on male germ cells, as well as on their malignant counterparts, has not been largely studied. In previous works, we reported our findings on a cell line derived from a human seminoma lesion (TCam-2 cell line) showing that acute exposure to simulated microgravity altered microtubule orientation, induced autophagy, and modified cell metabolism stimulating ROS production. Moreover, we demonstrated that the antioxidant administration prevented both TCam-2 microgravity-induced microtubule disorientation and autophagy induction. Herein, expanding previous investigations, we report that simulated microgravity exposure for 24 h induced the appearance, at an ultrastructural level, of cell-to-cell junctional contacts that were not detectable in cells grown at 1 g. In line with this result, pan-cadherin immunofluorescence analyzed by confocal microscopy, revealed the clustering of this marker at the plasma membrane level on microgravity exposed TCam-2 cells. The upregulation of cadherin was confirmed by Western blot analyses. Furthermore, we demonstrated that the microgravity-induced ROS increase was responsible for the distribution of cadherin nearby the plasma membrane, together with beta-catenin since the administration of antioxidants prevented this microgravity-dependent phenomenon. These results shed new light on the microgravity-induced modifications of the cell adhesive behavior and highlight the role of ROS as microgravity activated signal molecules. [ABSTRACT FROM AUTHOR]
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- 2020
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12. Microgravity Exposure Induces Antioxidant Barrier Deregulation and Mitochondrial Structure Alterations in TCam-2 Cells †.
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Gesualdi, Luisa, Berardini, Marika, Ferranti, Francesca, Reale, Anna, Zampieri, Michele, Karpach, Katsiaryna, Mariggiò, Maria A., Morabito, Caterina, Guarnieri, Simone, Catizone, Angela, and Ricci, Giulia
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REDUCED gravity environments ,CELL culture ,GRAVITATION ,MITOCHONDRIA ,GERM cells ,TRANSMISSION electron microscopy - Abstract
One of the hallmarks of microgravity-induced alterations in several cell models is an alteration in oxidative balance. Notably, male germ cells, sensitive to oxidative stress, have also been shown susceptibility to changes in gravitational force. To gain more insights into the mechanisms of male germ cells' response to altered gravity, a 3D cell culture model was established from TCam-2 cells, a seminoma cell line and the only available in vitro model to study mitotically active human male germ cells. TCam-2 spheroids were cultured for 24 hours under unitary gravity (UG) or simulated microgravity conditions (SM), which was achieved using a random positioning machine (RPM). Apoptosis and necrosis analyses performed on the UG- and SM exposed samples revealed no significant differences in all of the cell death markers. Notably, the Mitosox assay revealed significant oxidation of mitochondria, after microgravity exposure, at least at this culture time. In the SM-treated samples, gene expression levels (evaluated by real-time PCR) of the main enzymes of the antioxidant barrier, GPX1 and NCF1, were reduced, indicating an influence of SM on mitochondrial function. Notably, the expression of HMOX, involved in the heme catabolism of mitochondrial cytochromes, was increased. The SOD, XDH, CYBA, NCF-2, TXN, and TXNRD genes were not affected. The ultrastructural analysis by transmission electron microscopy revealed that SM significantly altered TCam-2 spheroid mitochondria, which appeared swollen and, in some cases, disrupted. Indeed, mitophagy, or mitochondrial autophagy, appears to be more represented in the samples exposed to simulated microgravity. This result seems to be in line with the increase, mediated by the simulated microgravity, in the enzyme HMOX. All together, these preliminary data demonstrate TCam-2 spheroids' sensitivity to acute SM exposure, strongly indicating a microgravity-dependent modulation of mitochondrial morphology and activity and encouraging us to perform further investigations on the chronical exposure to SM of TCam-2 spheroids. [ABSTRACT FROM AUTHOR]
- Published
- 2023
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13. Microgravity-Induced Metabolic Response in 2D and 3D TCam-2 Cell Cultures †.
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Morabito, Caterina, Guarnieri, Simone, Berardini, Marika, Gesualdi, Luisa, Ferranti, Francesca, Reale, Anna, Ricci, Giulia, Catizone, Angela, and Mariggiò, Maria A.
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CELL culture ,HUMAN space flight ,MALE reproductive organs ,ANAEROBIC metabolism ,REACTIVE oxygen species ,OXIDANT status - Abstract
The past few decades have seen an increasing number of both space travels and studies aimed at investigating the effects induced by space flights and the environment on humans. One of the main features of these conditions is the presence of altered gravity, mostly represented by microgravity experienced by astronauts. Microgravity is well known to induce deleterious effects at cellular, organ and systemic levels, including alterations in the male and female reproductive systems. In the present study, we investigated the effect of simulated microgravity on the metabolic activity of male germ cells using TCam-2 line as a cell model. These cells were cultured in the Random Positioning Machine that simulated microgravity conditions, and were grown as 2D monolayers or 3D spheroids to assay the effects on single cells or on organ-like structures. After a 24 hour-exposure to simulated microgravity, TCam-2 monolayers showed: (1) a decreased proliferation rate and a delay in cell cycle progression; (2) increased anaerobic metabolism; (3) increased levels of reactive oxygen species and superoxide anion; (4) modifications in mitochondrial morphology. After the same 24 hour-exposure, TCam-2 spheroids showed: (1) an increased anaerobic and aerobic activity in 40% and 26% of samples, respectively; (2) alterations in the redox balance with a decrease in catalase activity in about 65% of cell samples, and therefore, a deficit in the cellular antioxidant capacity; (3) increases in oxidative damage to proteins and lipids in more than 50% of cell samples. In conclusion, these data demonstrated a clear inference of simulated microgravity on the metabolic activity of TCam-2 cells, which is expressed through the activation of an oxidative stress state, that, if not compensated for, could be deleted over time. [ABSTRACT FROM AUTHOR]
- Published
- 2023
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14. Microgravity Exposure Alterations of Cellular Junctions Proteins in TCam-2 Cells: Localization and Interaction †.
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Berardini, Marika, Gesualdi, Luisa, Ferranti, Francesca, Mariggiò, Maria Addolorata, Morabito, Caterina, Guarnieri, Simone, Ricci, Giulia, and Catizone, Angela
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REDUCED gravity environments ,CELL culture ,CYTOPLASMIC filaments ,TIGHT junctions ,TRANSMISSION electron microscopy ,CONFOCAL microscopy ,SPACE environment - Abstract
One of the most important hazards of the space environment is microgravity, which causes an alteration in the physiology of different systems, including the reproductive one. It is widely accepted that cytoskeleton is the microgravity-sensitive apparatus of the cells, and that cytoskeletal modifications are responsible for microgravity-triggered cell alterations. We established a 3D free-floating culture system from TCam-2 cell, a human seminoma cell line, and then exposed the obtained TCam-2 spheroids for 24 h at unitary gravity (UG), or under a simulated microgravity condition (SM), using the random position machine (RPM). We tested the cytoskeletal and junctional features of these samples using Western blot and confocal microscopy analysis to elucidate the impact of microgravity on the adherent and occluding junctions of TCam-2 spheroids. The junctional ultrastructure was studied using transmission electron microscopy (TEM). TEM analysis revealed the presence of occluding junctions both in UG or SM samples. Even if Western blot revealed no quantitative difference in actin and occludin proteins both in UG and SM exposed samples, fluorescence colocalization analysis showed a significative increase in the colocalization area of occludin and actin proteins in the superficial layer of TCam-2 spheroids grown in RPM conditions. This result let us speculate that tight junction functionality is different in UG and SM exposed spheroids. As far as adherent junctions are concerned, TEM analysis revealed adherent junctions both in UG or SM samples. Moreover, we observed by Western blot a trend in terms of the increase in the vimentin expression in SM exposed spheroids. Confocal microscopy analyses confirmed this significant increase. All together, these data suggest that simulated microgravity conditions in TCam-2 spheroids alter the tight junction assembly, while the increase in the intermediate filament's structures can in part be associated with an enrichment in the adherent junctions. A functional investigation is needed to more deeply clarify this hypothesis. [ABSTRACT FROM AUTHOR]
- Published
- 2023
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15. Microgravity Modifies the Phenotype of Fibroblast and Promotes Remodeling of the Fibroblast–Keratinocyte Interaction in a 3D Co-Culture Model.
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Fedeli, Valeria, Cucina, Alessandra, Dinicola, Simona, Fabrizi, Gianmarco, Catizone, Angela, Gesualdi, Luisa, Ceccarelli, Simona, Harrath, Abdel Halim, Alwasel, Saleh H., Ricci, Giulia, Pedata, Paola, Bizzarri, Mariano, and Monti, Noemi
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REDUCED gravity environments ,FIBROBLASTS ,PHENOTYPES ,PHENOTYPIC plasticity ,WEIGHTLESSNESS ,NUCLEOPLASM - Abstract
Microgravity impairs tissue organization and critical pathways involved in the cell–microenvironment interplay, where fibroblasts have a critical role. We exposed dermal fibroblasts to simulated microgravity by means of a Random Positioning Machine (RPM), a device that reproduces conditions of weightlessness. Molecular and structural changes were analyzed and compared to control samples growing in a normal gravity field. Simulated microgravity impairs fibroblast conversion into myofibroblast and inhibits their migratory properties. Consequently, the normal interplay between fibroblasts and keratinocytes were remarkably altered in 3D co-culture experiments, giving rise to several ultra-structural abnormalities. Such phenotypic changes are associated with down-regulation of α-SMA that translocate in the nucleoplasm, altogether with the concomitant modification of the actin-vinculin apparatus. Noticeably, the stress associated with weightlessness induced oxidative damage, which seemed to concur with such modifications. These findings disclose new opportunities to establish antioxidant strategies that counteract the microgravity-induced disruptive effects on fibroblasts and tissue organization. [ABSTRACT FROM AUTHOR]
- Published
- 2022
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16. Survival Pathways Are Differently Affected by Microgravity in Normal and Cancerous Breast Cells.
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Monti, Noemi, Masiello, Maria Grazia, Proietti, Sara, Catizone, Angela, Ricci, Giulia, Harrath, Abdel Halim, Alwasel, Saleh H., Cucina, Alessandra, and Bizzarri, Mariano
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BREAST ,REDUCED gravity environments ,APOPTOSIS ,CELLS - Abstract
Metazoan living cells exposed to microgravity undergo dramatic changes in morphological and biological properties, which ultimately lead to apoptosis and phenotype reprogramming. However, apoptosis can occur at very different rates depending on the experimental model, and in some cases, cells seem to be paradoxically protected from programmed cell death during weightlessness. These controversial results can be explained by considering the notion that the behavior of adherent cells dramatically diverges in respect to that of detached cells, organized into organoids-like, floating structures. We investigated both normal (MCF10A) and cancerous (MCF-7) breast cells and found that appreciable apoptosis occurs only after 72 h in MCF-7 cells growing in organoid-like structures, in which major modifications of cytoskeleton components were observed. Indeed, preserving cell attachment to the substrate allows cells to upregulate distinct Akt- and ERK-dependent pathways in MCF-7 and MCF-10A cells, respectively. These findings show that survival strategies may differ between cell types but cannot provide sufficient protection against weightlessness-induced apoptosis alone if adhesion to the substrate is perturbed. [ABSTRACT FROM AUTHOR]
- Published
- 2021
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17. The PI3K/AKT Pathway Is Activated by HGF in NT2D1 Non-Seminoma Cells and Has a Role in the Modulation of Their Malignant Behavior.
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Gesualdi, Luisa, Leonetti, Erica, Cucina, Alessandra, Scicchitano, Bianca Maria, Sorrentino, Silvia, Tarsitano, Maria Grazia, Isidori, Andrea, Bizzarri, Mariano, Filippini, Antonio, Riccioli, Anna, Cammarota, Marcella, Gigantino, Vincenzo, Ricci, Giulia, and Catizone, Angela
- Subjects
TERATOCARCINOMA ,PHOSPHATIDYLINOSITOL 3-kinases ,CELL migration ,TESTICULAR cancer ,CELLS - Abstract
Overactivation of the c-MET/HGF system is a feature of many cancers. We previously reported that type II testicular germ cell tumor (TGCT) cells express the c-MET receptor, forming non-seminomatous lesions that are more positive compared with seminomatous ones. Notably, we also demonstrated that NT2D1 non-seminomatous cells (derived from an embryonal carcinoma lesion) increase their proliferation, migration, and invasion in response to HGF. Herein, we report that HGF immunoreactivity is more evident in the microenvironment of embryonal carcinoma biopsies with respect to seminomatous ones, indicating a tumor-dependent modulation of the testicular niche. PI3K/AKT is one of the signaling pathways triggered by HGF through the c-MET activation cascade. Herein, we demonstrated that phospho-AKT increases in NT2D1 cells after HGF stimulation. Moreover, we found that this pathway is involved in HGF-dependent NT2D1 cell proliferation, migration, and invasion, since the co-administration of the PI3K inhibitor LY294002 together with HGF abrogates these responses. Notably, the inhibition of endogenous PI3K affects collective cell migration but does not influence proliferation or chemotactic activity. Surprisingly, LY294002 administered without the co-administration of HGF increases cell invasion at levels comparable to the HGF-administered samples. This paradoxical result highlights the role of the testicular microenvironment in the modulation of cellular responses and stimulates the study of the testicular secretome in cancer lesions. [ABSTRACT FROM AUTHOR]
- Published
- 2020
- Full Text
- View/download PDF
18. Active Fraction from Embryo Fish Extracts Induces Reversion of the Malignant Invasive Phenotype in Breast Cancer through Down-Regulation of TCTP and Modulation of E-cadherin/β-catenin Pathway.
- Author
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Proietti, Sara, Cucina, Alessandra, Pensotti, Andrea, Biava, Pier Mario, Minini, Mirko, Monti, Noemi, Catizone, Angela, Ricci, Giulia, Leonetti, Erica, Harrath, Abdel Halim, Alwasel, Saleh H., and Bizzarri, Mariano
- Subjects
TUMOR microenvironment ,CADHERINS ,CATENINS ,BREAST cancer ,PHENOTYPES ,DOWNREGULATION ,CYTOSKELETON - Abstract
Some yet unidentified factors released by both oocyte and embryonic microenvironments demonstrated to be non-permissive for tumor development and display the remarkable ability to foster cell/tissue reprogramming, thus ultimately reversing the malignant phenotype. In the present study we observed how molecular factors extracted from Zebrafish embryos during specific developmental phases (20 somites) significantly antagonize proliferation of breast cancer cells, while reversing a number of prominent aspects of malignancy. Embryo extracts reduce cell proliferation, enhance apoptosis, and dramatically inhibit both invasiveness and migrating capabilities of cancer cells. Counteracting the invasive phenotype is a relevant issue in controlling tumor spreading and metastasis. Moreover, such effect is not limited to cancerous cells as embryo extracts were also effective in inhibiting migration and invasiveness displayed by normal breast cells undergoing epithelial–mesenchymal transition upon TGF-β1 stimulation. The reversion program involves the modulation of E-cadherin/β-catenin pathway, cytoskeleton remodeling with dramatic reduction in vinculin, as well as downregulation of TCTP and the concomitant increase in p53 levels. Our findings highlight that—contrary to the prevailing current "dogma", which posits that neoplastic cells are irreversibly "committed"—the malignant phenotype can ultimately be "reversed", at least partially, in response to environmental morphogenetic influences. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF
19. c-Src Recruitment is Involved in c-MET-Mediated Malignant Behaviour of NT2D1 Non-Seminoma Cells.
- Author
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Leonetti, Erica, Gesualdi, Luisa, Corano Scheri, Katia, Dinicola, Simona, Fattore, Luigi, Masiello, Maria Grazia, Cucina, Alessandra, Mancini, Rita, Bizzarri, Mariano, Ricci, Giulia, and Catizone, Angela
- Subjects
CANCER treatment ,GERM cell tumors ,CANCER chemotherapy ,DRUG resistance ,HEPATOCYTE growth factor ,SEMINOMA - Abstract
c-MET pathway over-activation is the signature of malignancy acquisition or chemotherapy resistance of many cancers. We recently demonstrated that type II Testicular Germ Cell Tumours (TGCTs) express c-MET receptor. In particular, we elucidated that the non-seminoma lesions express c-MET protein at higher level, compared with the seminoma ones. In line with this observation, NTERA-2 clone D1 (NT2D1) non-seminoma cells increase their proliferation, migration and invasion in response to Hepatocyte Growth Factor (HGF). One of the well-known adaptor-proteins belonging to c-MET signaling cascade is c-Src. Activation of c-Src is related to the increase of aggressiveness of many cancers. For this reason, we focused on the role of c-Src in c-MET-triggered and HGF-dependent NT2D1 cell activities. In the present paper, we have elucidated that this adaptor-protein is involved in HGF-dependent NT2D1 cell proliferation, migration and invasion, since Src inhibitor-1 administration abrogates these responses. Despite these biological evidences western blot analyses have not revealed the increase of c-Src activation because of HGF administration. However, notably, immunofluorescence analyses revealed that cytoplasmic and membrane-associated localization of c-Src shifted to the nuclear compartment after HGF stimulation. These results shed new light in the modality of HGF-dependent c-Src recruitment, and put the basis for novel investigations on the relationship between c-Src, and TGCT aggressiveness. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF
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