The retrovirus-mediated antisense human telomerase RNA (hTR) gene limits the growth of hepatocellular carcinoma growth in cell culture and animals.
OBJECTIVE
To investigate the inhibitory effect of retrovirus-mediated antisense human telomerase RNA (hTR) gene therapy on hepatocelluar carcinoma.


METHODS
We first constructed the sense and antisense hTR vectors and then transfected these into HepG2 cells. Telomerase activity, cell growth curves, proliferating cell nuclear antigen expression (PCNA), cell cycle distribution, and cell apoptosis were detected by the means of telemere repeat amplification protocol (TRAP), MTT assay, immunofluorescence, flow cytometric analysis, and transferase-mediated nick end labeling (TUNEL), respectively. In order to further confirm the therapeutic effect of this gene therapy, we developed an experimental line of HepG2 tumor-bearing nude mice by and directly injected these with retrovirus expressing the antisense hTR gene. Tumor growth was determined by tumor volume, and cell apoptosis was analyzed by TUNEL.


RESULTS
The antisense hTR gene was shown to be successfully integrated into the target cells' genome. HepG2 cells transfected with the antisense hTR gene showed down-regulated telomerase activity, inhibited cell growth, decreased PCNA expression, and increased apoptotic rate. Moreover, flow cytometry revealed a decrease of cells in the S phase with cell cycle arrest at the G2/M phase. In the antisense hTR-treated group, tumor growth was significantly reduced and showed an increase of apoptotic cells.


CONCLUSIONS
The results indicate that the specific inhibitor of the hTR template is likely to be a very efficient tool for hepatocellular carcinoma research and may possess potential therapeutic significance in the future clinical practice.
Liu
Ji-yong
JY
Department of Gastroenterology, Shandong Provincial Hospital, Shandong University, Jing 5 Wei 7 Road, no. 324, Ji'nan, Shandong Province 250021, China. liujiyong@medmail.com.cn
Zhu
Qiang
Q
Li
Jianfeng
J
Zhao
Shulei
S
Li
Luning
L
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000473:pathology / Q000628:therapy*; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D015316:Genetic Therapy; D005822:Genetic Vectors; D008113:Liver Neoplasms / Q000473:pathology / Q000628:therapy*; D051379:Mice; D008807:Mice, Inbred BALB C; D012313:RNA; D016372:RNA, Antisense; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D012190:Retroviridae; D019098:Telomerase
D016372:RNA, Antisense; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; C422952:telomerase RNA; D012313:RNA; D019098:Telomerase