Objective: The aim of this study was designed to investigate the effect of TSA on human umbilical vein endothelial cells and to reveal its possible mechanisms and relationship between apoptosis and activity of telomerase reverse transcriptase.
Methods: sulforhodamine B method was employed to determine the growth rate of umbilical vein endothelial cells. The cell apoptotic rate was measured by flow cytometry (FCM). The hTERT and p21(Waf1) mRNA expression before and after TSA treatment were detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). The quantitative of hTERT protein expression in cells were detected by flow cytometry. After transfection, the cell telomerase activity was detected by PCR and telomeric repeat amplification protocol assay (PCR-TRAP-ELISA) and early apoptosis was measured by Annexin V/PI stain and flow cytometry.
Results: After being treated with TSA, the proliferation of umbilical vein endothelial cells was inhibited. Slight apoptosis and cell cycle arrest were detected. However, the same concentration of TSA induced serious apoptosis in HeLa cells. Up-regulation of hTERT mRNA expression was observed within 48 h after TSA treatment, but the change of p21(Waf1) expression was not significant. The umbilical vein endothelial cells hTERT protein expression level was increased within 24 h. After transfection of the dominant negative, wild type and control hTERT plasmid, a significant difference of telomerase activity in these cells was observed by PCR-TRAP-ELISA assay. WT-hTERT-transfected cells were more resistant to apoptosis induced by trichostatin A.
Conclusion: Human umbilical vein endothelial cells could be resistant to apoptosis induced by high concentrate TSA, and hTERT might play an important role in this process.