A K Khanna1, J D Hosenpud. 1. Cardiovascular Research Center, Medical College of Wisconsin, Milwaukee 53226, USA. akkhanna@mcw.edu
Abstract
BACKGROUND: Current immunosuppression strategies involve inhibition of T cell activation and/or lymphocyte proliferation. During T cell cycle progression/activation, the expression of cyclins and cyclin dependent kinases is increased. In this study, we examined whether cyclosporine A (CsA) suppresses the cell cycle progression through the induction of the cell cycle inhibitor p21. Because CsA induces the expression of transforming growth factor (TGF)-beta, and TGF-beta induces p21 expression, we also determined whether CsA's induction of p21 is dependent on or independent of TGF-beta. METHOD: Using reverse transcription assisted polymerase chain reaction and western blot analysis, we studied the induction of p21 mRNA and protein in human T cells and A-549 cells (human lung adenocarcinoma cells) by CsA. The stimulation of p21 promoter activity was studied by luciferase assay using p21-luc, chimeric plasmid DNA containing a p21 promoter segment, and luciferase reporter gene. The dependence of CsA's induction of p21 was studied using anti-TGF-beta antibody and TGF-beta altered A-549 cells. RESULTS: CsA induced p21 mRNA protein expression and stimulated its promoter activity in lymphoid (T cells) and nonlymphoid (human lung adenocarcinoma, A-549 cells).CsA's induction of p21 was inhibited both by a neutralizing anti-TGF-beta antibody and in TGF-beta-altered A-549 cells, consistent with its effects on p21 requiring TGF-beta. CONCLUSION: These data support the hypothesis that at least one component of CsA's antiproliferative effects may occur through the induction of p21 and that this induction is dependent on TGF-beta. Should p21 induction be a viable immunosuppressive strategy, inducing this molecule independent from the fibrogenic cytokine TGF-beta might reduce the toxicity associated with current immunosuppression.
BACKGROUND: Current immunosuppression strategies involve inhibition of T cell activation and/or lymphocyte proliferation. During T cell cycle progression/activation, the expression of cyclins and cyclin dependent kinases is increased. In this study, we examined whether cyclosporine A (CsA) suppresses the cell cycle progression through the induction of the cell cycle inhibitor p21. Because CsA induces the expression of transforming growth factor (TGF)-beta, and TGF-beta induces p21 expression, we also determined whether CsA's induction of p21 is dependent on or independent of TGF-beta. METHOD: Using reverse transcription assisted polymerase chain reaction and western blot analysis, we studied the induction of p21 mRNA and protein in human T cells and A-549 cells (humanlung adenocarcinoma cells) by CsA. The stimulation of p21 promoter activity was studied by luciferase assay using p21-luc, chimeric plasmid DNA containing a p21 promoter segment, and luciferase reporter gene. The dependence of CsA's induction of p21 was studied using anti-TGF-beta antibody and TGF-beta altered A-549 cells. RESULTS:CsA induced p21 mRNA protein expression and stimulated its promoter activity in lymphoid (T cells) and nonlymphoid (humanlung adenocarcinoma, A-549 cells).CsA's induction of p21 was inhibited both by a neutralizing anti-TGF-beta antibody and in TGF-beta-altered A-549 cells, consistent with its effects on p21 requiring TGF-beta. CONCLUSION: These data support the hypothesis that at least one component of CsA's antiproliferative effects may occur through the induction of p21 and that this induction is dependent on TGF-beta. Should p21 induction be a viable immunosuppressive strategy, inducing this molecule independent from the fibrogenic cytokine TGF-beta might reduce the toxicity associated with current immunosuppression.
Authors: Theodore H Welling; Guanyi Lu; Keri Csencsits; Sherri C Wood; Lamis Jarvinen; D Keith Bishop Journal: Surgery Date: 2007-12-21 Impact factor: 3.982