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Hepatocyte Cell Adhesion Molecule Inhibits Renal Cell Carcinoma Proliferation through Regulating C-MYC

Author: ZhangQiaoLin
Tutor: LuoChunLi
School: Chongqing Medical University
Course: Clinical Laboratory Science
Keywords: hepaCAM renal cell carcinoma c-Myc cell cycle
CLC: R737.11
Type: Master's thesis
Year: 2011
Downloads: 13
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Abstract


PART ONEObjective: To investigate the effect of hepatocyte cell adhesion molecule (hepaCAM) on proliferation of renal cell carcinoma and its mechanisms.Methods: The vector containing hepaCAM gene was transfeceted into 786-0 cells, the expression of hepaCAM and c-Myc in 786-0 cells were measured by RT-PCR and western blot; Cell proliferation was detected by cell counting; the cell cycle was analyzed by FCM.Results: Up-regulated the expression of hepaCAM gene in 786-0 cells can inhibit cell proliferation, which accompany with down-regulation of c-Myc.Conclusion: we demonstrated that re-expression of hepaCAM caused an accumulation in G0/G1 phase in 786-0 cells. This reaction was accompanied by a substantial reduction of c-Myc expression through using an ectopic hepaCAM expression system. Nevertheless, re-expression of hepaCAM can result in apparent reduction of c-Myc protein while no corresponding reduction of c-Myc mRNA. This suggests that this reaction might take place at a post-transcriptional level rather than transcriptional one.PART TWOObjective: To investigate the effect of the small molecule c-Myc inhibitor, 10058-F4, on proliferation of renal cell carcinoma and the mechanisms of hepaCAM inhibits renal cell carcinoma proliferation through regulating c-Myc.Methods: Cell proliferation was detected by MTT assay; The proliferation of 786-0 cells was analyzed by FCM; Western blot was used to detected the effects of hepaCAM on the stability and phosphorylation of c-Myc and the targets of c-Myc, P21 and cyclin D1.Results: we showed a comparable decrease in proliferation and G0/G1 accumulation of 786-0 and RC-2 cells after treatment with a small molecule c-Myc inhibitor, 10058-F4. re-expression of hepaCAM decreased c-Myc stability by increasing the proportion of c-Myc phosphorylation on T58 and the expression of P21 was increased and the expression of cyclin D1 was decreased.Conclusion: we showed a comparable decrease in proliferation and G0/G1 accumulation of 786-0 and RC-2 cells after treatment with a small molecule c-Myc inhibitor, 10058-F4. It demonstrated that down regulation of c-Myc was an essential process in controlling growth inhibitory actions of hepaCAM. Nevertheless, re-expression of hepaCAM can result in apparent reduction of c-Myc protein while no corresponding reduction of c-Myc mRNA. This suggests that this reaction might take place at a post-transcriptional level rather than transcriptional one. Consistent with these findings, hepaCAM decreased c-Myc stability by increasing the proportion of c-Myc phosphorylation on T58 which can be abrogated by a proteasomal inhibitor (MG132). Thus, our results imply that the decrease in c-Myc protein expression, resulting from ectopic expression of hepaCAM, may contribute to inhibition of proliferation in these cells.

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