PMID: 16643751Apr 29, 2006Paper

Anti-proliferative effect of herpes simplex virus thymidine kinase gene system on human Tenon capsule fibroblasts in vitro

[Zhonghua yan ke za zhi] Chinese journal of ophthalmology
Ji-bing WangYan-tao Wei

Abstract

To observe anti-proliferative effect of herpes simplex virus thymidine kinase gene system (HSV-tk) on human Tenon capsule fibroblasts (HTFs) and it's mechanism. Retroviral vector was constructed containing HSV-tk gene and transfer to packaging cell line PT67. The positive clones was selected with G418 and the supernatant was collected which contains virus particles. The titer of the virus was also calculated. The virus particles were verified by transmission electron microscope (TEM). PCR was carried out to detect the integrity of tk gene into viral DNA genome. HTFs cells were infected with the virus and the positive clones were selected for propagation culture. RT-PCR and Western blot were used to detect tk gene expression. The anti-proliferative effect of tk gene on HTFs cells was determined by MTT. Apoptosis of cells was detected by Hoechst 33258 DNA staining and flow cytometry. The morphologic changes of the cell were observed by TEM. The retroviral vector pL (tk) SN was constructed successfully which was verified by enzyme cutting. The shape of virus particles derived from the package cells appeared to be oval or spherical under TEM with the diameter around 100 nm. The tk gene was detected in the viral DNA genome by PCR. T...Continue Reading

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