摘要：
Although draft genome sequences of two of the major human schistosomes, Schistosoma japonicum and Schistosoma mansoni are available, the structures and characteristics of most genes and the influence of exogenous genes on the metabolism of schistosomes remain uncharacterized. Furthermore, which functional genomics approaches will be tractable for schistosomes are not yet apparent. Here, the vesicular stomatitis virus glycoprotein (VSVG)-pseudotyped pantropic retroviral vector pBABE-puro was modified to incorporate the human telomerase reverse transcriptase gene (hTERT) as a reporter, under the control of the retroviral long terminal repeat (LTR). Pseudotyped virions were employed to transduce S. japonicum to investigate the utility of retrovirus-mediated transgenesis of S japonicum and the activity of human telomerase reverse transcriptase as a reporter transgene in schistosomes. Schistosomules perfused from experimentally infected rabbits were cultured for 6 days after exposure to the virions after which genomic DNAs from virus exposed and control worms were extracted. Analysis of RNA from transduced parasites and immunohistochemistry of thin parasite sections revealed expression of hTERT in the transduced worms. Expression of hTERT was also confirmed by immunoblot analysis. These findings indicated that S. japonicum could be effectively transduced by VSVG-pseudotyped retrovirus carrying the hTERT gene. Given the potential of hTERT to aid in derivation of immortalized cells, these findings suggest that this pantropic retroviral approach can be employed to transduce cells from specific tissues and organs of schistosomes to investigate the influence of transgene hTERT on growth and proliferation of schistosome cells. (C) 2010 Elsevier B.V. All rights reserved.
作者机构：
[Liu, Hao-Ran; Zeng, Chun-Jiao; Liu, Ying-Lin; Li, Yan-Lai] Hunan Univ, Dept Pharmacol, Coll Chem & Chem Engn, Changsha 410082, Hunan, Peoples R China.;[He, Hai-Bo] Zhejiang Univ, Inst Chinese Herb Med, Coll Pharmaceut Sci, Hangzhou 310058, Zhejiang, Peoples R China.;[Peng, Xiang-Dong] Cent S Univ, Xiangya Hosp 3, Dept Pharm, Changsha, Hunan, Peoples R China.;[He, Gui-Xia] Hunan Univ TCM, Coll Pharm, Dept Nat Pharmaceut Chem, Changsha, Hunan, Peoples R China.
通讯机构：
[He, Hai-Bo] Z;Zhejiang Univ, Inst Chinese Herb Med, Coll Pharmaceut Sci, Hangzhou 310058, Zhejiang, Peoples R China.
摘要：
Total saponin of Solanum lyratum Thunb (TSSLT), a species of natural biologically active substances isolated from Solanum lyratum Thunb, possesses various bioactivities. It has been proposed that the induction of apoptosis may be the basis of its antitumor activity. However, the molecular mechanism underlying the total saponin-induced apoptotic process remains unknown. In the present study, we describe the anti-proliferative effect of TSSLT on human cervical cancer cells (Hela). The TSSLT induced apoptosis of Hela in a time-dependent manner with an IC50 for cell viability of 6 μg/ml. The TSSLT-induced cell death was characterized by changes in cell morphology, DNA fragmentation, activation of caspase-like activities, poly (ADP-ribose) polymerase (PARP) cleavage and release of cytochrome c (cyt c) into cytosol. TSSLT activated various caspases such as caspase-3, -8, and -9 (like) activities but not caspase-1 like activity. The cell death was completely prevented by the pan caspase inhibitor benzyloxy carbonyl-Val-Ala-Asp- fluoromethyl-ketone (Z-VAD-FMK). More than 80% cell survival was observed in the presence of a caspase-3 inhibitor. In addition, treatment with TSSLT induced the increase of Bax:Bcl-2 ratio in Hela cells. These results suggest that the induction of apoptosis by TSSLT involves multiple pathways antigen including death receptor and mitochondrial pathway and strongly suggest that the mitochondrial pathway was mediated by low expression of Bcl-2 and upregulation of Bax, release of cyt c and subsequent activation of caspase-3 followed by down stream events leading to apoptotic cell death.
