Csaba Matta

Publications of Csaba Matta

• PKCdelta is a positive regulator of chondrogenesis in chicken high density micromass cell cultures.

  Authors: Csaba Matta, Tamás Juhász, Zsolt Szíjgyártó, Bernadett Kolozsvári, Csilla Somogyi, Georgina Nagy, Pál Gergely, Róza Zákány

  Biochimie. 02/2011; 93(2):149-59.

  We aimed to elucidate the role of the Ca-independent PKC isoenzyme PKCdelta in the regulation of spontaneous in vitro chondrogenesis occurring in a 6-day-long culturing period in chicken limbWe aimed to elucidate the role of the Ca-independent PKC isoenzyme PKCdelta in the regulation of spontaneous in vitro chondrogenesis occurring in a 6-day-long culturing period in chicken limb bud-derived high density cell cultures (HDC). PKCdelta expression and activity were detectable throughout the entire culturing period with a peak on days 2 and 3, when most of the chondroblasts differentiate. To inhibit the activity of PKCdelta, either the natural compound rottlerin was transiently applied to the culture medium of HDC in 2.5, 5 or 10 μM concentrations, or gene silencing was performed by using PKCdelta shRNA. Rottlerin significantly reduced the overall PKC activity in enzyme activity assays of cell-free samples of untreated control HDC, probably via the inhibition of PKCdelta. On the contrary, we were unable to detect any consistent change of PKC enzyme activity assayed in samples of HDC treated with rottlerin during culturing. PKCdelta gene silencing resulted in a significantly lower PKC activity. Both rottlerin and PKCdelta shRNA caused a severe reduction in cartilage formation, furthermore protein and phospho-protein levels of Sox9, the key transcription factor of chondrogenesis, were also significantly decreased. Rottlerin lowered, while PKCdelta gene silencing elevated the phosphorylation status of ERK1/2. Our data suggest that PKCdelta stimulates chondrogenesis via influencing Sox9 and ERK1/2 phosphorylation, but the inhibition of cartilage formation in the rottlerin-treated HDC is probably PKCdelta independent and rottlerin might have different effects when applied to cells or to an in vitro enzyme activity assay.

• Switch of voltage-gated K+ channel expression in the plasma membrane of chondrogenic cells affects cytosolic Ca2+-oscillations and cartilage formation.

  Authors: Zoltan Varga, Tamás Juhász, Csaba Matta, János Fodor, Éva Katona, Adam Bartok, Tamás Oláh, Attila Sebe, László Csernoch, Gyorgy Panyi, Róza Zákány

  PloS one. 01/2011; 6(11):e27957.

  Understanding the key elements of signaling of chondroprogenitor cells at the earliest steps of differentiation may substantially improve our opportunities for the application of mesenchymal stemUnderstanding the key elements of signaling of chondroprogenitor cells at the earliest steps of differentiation may substantially improve our opportunities for the application of mesenchymal stem cells in cartilage tissue engineering, which is a promising approach of regenerative therapy of joint diseases. Ion channels, membrane potential and Ca(2+)-signaling are important regulators of cell proliferation and differentiation. Our aim was to identify such plasma membrane ion channels involved in signaling during chondrogenesis, which may serve as specific molecular targets for influencing chondrogenic differentiation and ultimately cartilage formation. Using patch-clamp, RT-PCR and Western-blot experiments, we found that chondrogenic cells in primary micromass cell cultures obtained from embryonic chicken limb buds expressed voltage-gated Na(V)1.4, K(V)1.1, K(V)1.3 and K(V)4.1 channels, although K(V)1.3 was not detectable in the plasma membrane. Tetrodotoxin (TTX), the inhibitor of Na(V)1.4 channels, had no effect on cartilage formation. In contrast, presence of 20 mM of the K(+) channel blocker tetraethyl-ammonium (TEA) during the time-window of the final commitment of chondrogenic cells reduced K(V) currents (to 27±3% of control), cell proliferation (thymidine incorporation: to 39±4.4% of control), expression of cartilage-specific genes and consequently, cartilage formation (metachromasia: to 18.0±6.4% of control) and also depolarized the membrane potential (by 9.3±2.1 mV). High-frequency Ca(2+)-oscillations were also suppressed by 10 mM TEA (confocal microscopy: frequency to 8.5±2.6% of the control). Peak expression of TEA-sensitive K(V)1.1 in the plasma membrane overlapped with this period. Application of TEA to differentiated chondrocytes, mainly expressing the TEA-insensitive K(V)4.1 did not affect cartilage formation. These data demonstrate that the differentiation and proliferation of chondrogenic cells depend on rapid Ca(2+)-oscillations, which are modulated by K(V)-driven membrane potential changes. K(V)1.1 function seems especially critical during the final commitment period. We show the critical role of voltage-gated cation channels in the differentiation of non-excitable cells with potential therapeutic use.

• Inhibition of calcineurin by cyclosporine A exerts multiple effects on human melanoma cell lines HT168 and WM35.

  Authors: Tamás Juhász, Csaba Matta, Gábor Veress, Georgina Nagy, Zsolt Szíjgyártó, Zsanett Molnár, János Fodor, Róza Zákány, Pál Gergely

  International journal of oncology. 05/2009; 34(4):995-1003.

  The immunosuppressant cyclosporine A (CsA) is a specific pharmacological inhibitor of calcineurin, the Ca2+-calmodulin activated phospho-Ser/Thr-specific protein phosphatase. AlthoughThe immunosuppressant cyclosporine A (CsA) is a specific pharmacological inhibitor of calcineurin, the Ca2+-calmodulin activated phospho-Ser/Thr-specific protein phosphatase. Although calcineurin-inhibiting compounds are applied for local treatment of psoriasis or atopic dermatitis in dermatological practice, little is known about the functions of calcineurin in epidermis-derived malignancies. We investigated the effects of CsA on two human melanoma cell lines, the metastasis forming HT168 and WM35 established from an RGP primary lesion. CsA of 2 microM lowered the enzyme activity by 50% and caused elevation in both mRNA and protein expression of calcineurin. Cell proliferation was diminished, as well as the cellular morphology and the actin organization were altered in both cell lines. CsA increased cell death moderately in both cell lines and reduced the metabolic activity of HT168 cells, but not that of WM35 cells. CsA also elevated the expressions of both Bcl-2 and ERK1/2. Fibronectin guided migration of HT168 cells was stimulated under the effect of CsA, while that of WM35 cells was reduced, moreover, HT168 cells switched from the expression of beta3 to beta1 integrin, but WM35 cells continued to express beta3. Based on our results we propose a multiple, partly malignancy-dependent role of calcineurin in these melanoma cell lines.

• Ionotropic purinergic receptor P2X(4) is involved in the regulation of chondrogenesis in chicken micromass cell cultures.

  Authors: János Fodor, Csaba Matta, Tamás Juhász, Tamás Oláh, Mónika Gönczi, Zsolt Szíjgyártó, Pál Gergely, László Csernoch, Róza Zákány

  Cell calcium. 04/2009;

  We have previously demonstrated that elevation of free cytosolic Ca(2+) concentration at the time of differentiation of chondroblasts was mainly due to a Ca(2+) influx and it was indispensable toWe have previously demonstrated that elevation of free cytosolic Ca(2+) concentration at the time of differentiation of chondroblasts was mainly due to a Ca(2+) influx and it was indispensable to cartilage formation in chicken high density mesenchymal cell cultures (HDC) [C. Matta, J. Fodor, Z. Szijgyarto, T. Juhasz, P. Gergely, L. Csernoch, R. Zakany, Cytosolic free Ca(2+) concentration exhibits a characteristic temporal pattern during in vitro cartilage differentiation: a possible regulatory role of calcineurin in Ca-signalling of chondrogenic cells, Cell Calcium 44 (2008) 310-323]. Here, we report that chondrogenic cells secreted ATP and administration of ATP to the culture medium evoked Ca(2+) transients exclusively in the presence of extracellular Ca(2+) and only on day 3 of culturing, when the final commitment of chondroblasts occurs. Moreover, ATP caused elevated protein expression of the chondrogenic transcription factor Sox9 and stimulated cartilage matrix production. Expression pattern of different types of both ionotropic and metabotropic purinergic receptors was detected. Agonists of metabotropic receptors, ADP and UDP did not evoke any Ca(2+) transients and had no influence on cartilage formation, while UTP caused transient elevation of cytosolic Ca(2+) concentration in 3-day-old HDC without stimulating matrix production. Suramin, which blocks all P2X receptors but not P2X(4) did not impede the effects of ATP, furthermore, P2X(4) appeared in the plasma membrane fraction and gave signals with immunocytochemistry only from day 3. In summary, we suggest a role of ionotropic purinergic signalling of P2X(4) in the generation of ATP-dependent Ca(2+) transients of differentiating chondroblasts.

• Cytosolic free Ca(2+) concentration exhibits a characteristic temporal pattern during in vitro cartilage differentiation: A possible regulatory role of calcineurin in Ca-signalling of chondrogenic cells.

  Authors: Csaba Matta, János Fodor, Zsolt Szíjgyártó, Tamás Juhász, Pál Gergely, László Csernoch, Róza Zákány

  Cell calcium. 03/2008;

  We measured changes of cytosolic Ca(2+) concentration during chondrogenesis, which occurs in high-density cultures (HDC) of chondrifying chicken mesenchymal cells. A significant, transient elevationWe measured changes of cytosolic Ca(2+) concentration during chondrogenesis, which occurs in high-density cultures (HDC) of chondrifying chicken mesenchymal cells. A significant, transient elevation was detected in Fura-2-loaded cells on day 3 of culturing, when majority of chondrogenic cells of HDC become differentiated. This 140nM peak of cytosolic Ca(2+) concentration is a result of increased Ca-influx and is indispensable to proper chondrogenesis, because addition of 0.8mM EGTA to culture medium on day 2 or 3 significantly decreased the intracellular Ca(2+) concentration abolishing the Ca(2+)-peak of day 3 and inhibited cartilage formation. Uncontrolled Ca(2+) influx evoked by a Ca(2+) ionophore exerted dual effects on chondrogenesis in a concentration-dependent manner; 0.1mg/L A23187 increased, whereas 5mg/L A23187 almost totally blocked cartilage formation. Intracellular Ca-stores seemed not to have any significant participation in the regulation of changes of cytosolic Ca(2+) concentration of chondrifying cells. Activity of Ca-calmodulin-dependent protein phosphatase, calcineurin responded to changes of intracellular Ca(2+) concentration induced by EGTA or A23187 in a differentiation stage-dependent manner. Since inhibition of calcineurin with cyclosporine A eliminated the peak in the cytosolic Ca(2+) concentration, an active regulatory role of calcineurin on Ca(2+) influx of chondrifying cells can be supposed.

• Oxidative stress-induced poly(ADP-ribosyl)ation in chick limb bud-derived chondrocytes.

  Authors: Róza Zákány, Edina Bakondi, Tamás Juhász, Csaba Matta, Zsolt Szíjgyártó, Katalin Erdélyi, Eva Szabó, László Módis, László Virág, Pál Gergely

  International journal of molecular medicine. 05/2007; 19(4):597-605.

  Oxidative stress has been implicated in the pathogenesis of various diseases affecting chondrogenesis or the function of articular cartilage. DNA damage caused by oxidative stress may trigger theOxidative stress has been implicated in the pathogenesis of various diseases affecting chondrogenesis or the function of articular cartilage. DNA damage caused by oxidative stress may trigger the activation of the nuclear enzyme, poly(ADP-ribose) polymerase-1 (PARP-1) which may contribute to tissue injury. We aimed at investigating the effects of peroxynitrite (100-600 microM) and hydrogen peroxide (0.1-4 mM) on PARP activation and extracellular matrix production of high density micromass cultures (HDC) prepared from chick limb bud mesenchymal cells. We found that both oxidative species strongly inhibited matrix formation of HDCs treated on day 2 but not on day 5. The PARP inhibitor 3-aminobenzamide (3-AB) stimulated matrix production in non-stressed cells and prevented suppressed matrix production in oxidatively stressed cells. Both hydrogen peroxide and peroxynitrite induced PARP activation and poly(ADP-ribose) accumulation. Decreased proliferation, viability and NAD+ content were not or only slightly improved by 3-AB, indicating that 3-AB directly affects matrix formation. In conclusion, oxidative stress stimulates poly(ADP-ribose) metabolism and inhibits extracellular matrix production of HDCs in a PARP-dependent manner. Our findings may have implications for potential therapeutic approaches aimed at restoring the matrix production capacity of chondrogenic cells.

• Hydrogen peroxide inhibits formation of cartilage in chicken micromass cultures and decreases the activity of calcineurin: implication of ERK1/2 and Sox9 pathways.

  Authors: Róza Zákány, Zsolt Szíjgyártó, Csaba Matta, Tamás Juhász, Csilla Csortos, Kornélia Szucs, Gabriella Czifra, Tamás Bíró, László Módis, Pál Gergely

  Experimental cell research. 05/2005; 305(1):190-9.

  Calcineurin was found as a positive regulator of chondrogenesis in chondrifying chicken micromass cultures (HDCs), as cyclosporine A (CsA) reduced both the amount of cartilage and the expression ofCalcineurin was found as a positive regulator of chondrogenesis in chondrifying chicken micromass cultures (HDCs), as cyclosporine A (CsA) reduced both the amount of cartilage and the expression of mRNAs of aggrecan and the chondrogenic transcription factor Sox9. Cartilage formation was inhibited by H(2)O(2) in a concentration-dependent manner without loss of cellular viability or severe decrease of cell number. Expression of both the mRNA and the unphosphorylated protein Sox9 was decreased, while its phosphorylation was stimulated by either H(2)O(2) or CsA. Oxidative stress decreased the activity of calcineurin but the phosphorylation of the member of MAPK family ERK1/2 was extremely elevated either by 1 mM H(2)O(2) or 2 muM CSA. The ERK inhibitor PD098059 attenuated the depletion of cartilage matrix as well as decreased the expression and phosphorylation of Sox9 in cultures treated with H(2)O(2) or CsA. Our results suggest that the chondrogenesis-inhibiting effect of H(2)O(2) is mediated, at least partly, by inhibition of calcineurin and by activation of ERK1/2. We also propose a regulatory role of calcineurin in the phosphorylation level of either ERK1/2 or Sox9 and a positive role of ERK1/2 in regulating both the expression level and the phosphorylation state of Sox9 in chicken HDCs.

• Cytosolic free Ca2+ concentration exhibits a characteristic temporal pattern during in vitro cartilage differentiation: A possible regulatory role of calcineurin in Ca-signalling of chondrogenic cells

  Authors: Csaba Matta, János Fodor, Zsolt Szíjgyártó, Tamás Juhász, Pál Gergely, László Csernoch, Róza Zákány

  Cell Calcium.

  We measured changes of cytosolic Ca2+ concentration during chondrogenesis, which occurs in high-density cultures (HDC) of chondrifying chicken mesenchymal cells. A significant, transient elevationWe measured changes of cytosolic Ca2+ concentration during chondrogenesis, which occurs in high-density cultures (HDC) of chondrifying chicken mesenchymal cells. A significant, transient elevation was detected in Fura-2-loaded cells on day 3 of culturing, when majority of chondrogenic cells of HDC become differentiated. This 140 nM peak of cytosolic Ca2+ concentration is a result of increased Ca-influx and is indispensable to proper chondrogenesis, because addition of 0.8 mM EGTA to culture medium on day 2 or 3 significantly decreased the intracellular Ca2+ concentration abolishing the Ca2+-peak of day 3 and inhibited cartilage formation. Uncontrolled Ca2+ influx evoked by a Ca2+ ionophore exerted dual effects on chondrogenesis in a concentration-dependent manner; 0.1 mg/L A23187 increased, whereas 5 mg/L A23187 almost totally blocked cartilage formation. Intracellular Ca-stores seemed not to have any significant participation in the regulation of changes of cytosolic Ca2+ concentration of chondrifying cells. Activity of Ca–calmodulin-dependent protein phosphatase, calcineurin responded to changes of intracellular Ca2+ concentration induced by EGTA or A23187 in a differentiation stage-dependent manner. Since inhibition of calcineurin with cyclosporine A eliminated the peak in the cytosolic Ca2+ concentration, an active regulatory role of calcineurin on Ca2+ influx of chondrifying cells can be supposed.

• Ionotropic purinergic receptor P2X4 is involved in the regulation of chondrogenesis in chicken micromass cell cultures

  Authors: János Fodor, Csaba Matta, Tamás Juhász, Tamás Oláh, Mónika Gönczi, Zsolt Szíjgyártó, Pál Gergely, László Csernoch, Róza Zákány

  Cell Calcium.

  We have previously demonstrated that elevation of free cytosolic Ca2+ concentration at the time of differentiation of chondroblasts was mainly due to a Ca2+ influx and it was indispensable toWe have previously demonstrated that elevation of free cytosolic Ca2+ concentration at the time of differentiation of chondroblasts was mainly due to a Ca2+ influx and it was indispensable to cartilage formation in chicken high density mesenchymal cell cultures (HDC) [C. Matta, J. Fodor, Z. Szijgyarto, T. Juhasz, P. Gergely, L. Csernoch, R. Zakany, Cytosolic free Ca2+ concentration exhibits a characteristic temporal pattern during in vitro cartilage differentiation: a possible regulatory role of calcineurin in Ca-signalling of chondrogenic cells, Cell Calcium 44 (2008) 310–323]. Here, we report that chondrogenic cells secreted ATP and administration of ATP to the culture medium evoked Ca2+ transients exclusively in the presence of extracellular Ca2+ and only on day 3 of culturing, when the final commitment of chondroblasts occurs. Moreover, ATP caused elevated protein expression of the chondrogenic transcription factor Sox9 and stimulated cartilage matrix production. Expression pattern of different types of both ionotropic and metabotropic purinergic receptors was detected. Agonists of metabotropic receptors, ADP and UDP did not evoke any Ca2+ transients and had no influence on cartilage formation, while UTP caused transient elevation of cytosolic Ca2+ concentration in 3-day-old HDC without stimulating matrix production. Suramin, which blocks all P2X receptors but not P2X4 did not impede the effects of ATP, furthermore, P2X4 appeared in the plasma membrane fraction and gave signals with immunocytochemistry only from day 3. In summary, we suggest a role of ionotropic purinergic signalling of P2X4 in the generation of ATP-dependent Ca2+ transients of differentiating chondroblasts.

• Hydrogen peroxide inhibits formation of cartilage in chicken micromass cultures and decreases the activity of calcineurin: implication of ERK1/2 and Sox9 pathways

  Authors: Róza Zákány, Zsolt Szíjgyártó, Csaba Matta, Tamás Juhász, Csilla Csortos, Kornélia Szűcs, Gabriella Czifra, Tamás Bíró, László Módis, Pál Gergely

  Experimental Cell Research.

  Calcineurin was found as a positive regulator of chondrogenesis in chondrifying chicken micromass cultures (HDCs), as cyclosporine A (CsA) reduced both the amount of cartilage and the expression ofCalcineurin was found as a positive regulator of chondrogenesis in chondrifying chicken micromass cultures (HDCs), as cyclosporine A (CsA) reduced both the amount of cartilage and the expression of mRNAs of aggrecan and the chondrogenic transcription factor Sox9. Cartilage formation was inhibited by H2O2 in a concentration-dependent manner without loss of cellular viability or severe decrease of cell number. Expression of both the mRNA and the unphosphorylated protein Sox9 was decreased, while its phosphorylation was stimulated by either H2O2 or CsA. Oxidative stress decreased the activity of calcineurin but the phosphorylation of the member of MAPK family ERK1/2 was extremely elevated either by 1 mM H2O2 or 2 μM CSA. The ERK inhibitor PD098059 attenuated the depletion of cartilage matrix as well as decreased the expression and phosphorylation of Sox9 in cultures treated with H2O2 or CsA. Our results suggest that the chondrogenesis-inhibiting effect of H2O2 is mediated, at least partly, by inhibition of calcineurin and by activation of ERK1/2. We also propose a regulatory role of calcineurin in the phosphorylation level of either ERK1/2 or Sox9 and a positive role of ERK1/2 in regulating both the expression level and the phosphorylation state of Sox9 in chicken HDCs.