[Role of human telomerase reverse transcriptase in apoptosis of human umbilical vein endothelial cells induced by trichostatin A].
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.


CONCLUSIONS
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.
Wu
Peng
P
Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science & Technology, Wuhan 430030, China.
Xi
Ling
L
Chen
Gang
G
Wang
Bei-bei
BB
Luo
Dan-feng
DF
Lu
Yun-ping
YP
Zhou
Jian-feng
JF
Ma
Ding
D
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology*; D010957:Plasmids / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D019098:Telomerase / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014471:Umbilical Veins / Q000166:cytology
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004791:Enzyme Inhibitors; D006877:Hydroxamic Acids; D012333:RNA, Messenger; C012589:trichostatin A; C509186:TERT protein, human; D019098:Telomerase