| The ends of linear chromosomes, named telomeres, play an important role in the maintenance of chromosomal integrity and function. Human telomeres consist of multiple repeats of the hexametric sequence TTAGGG. Due to the "end replication problem" in linear chromosomes, telomeric DNA is not completely copied during DNA replication and telomeres progressively shorten. After some telomeres have become sufficiently short, normal human cells enter senescence and stop diving.Telomerase is a special ribonucleoprotein complex that extends and maintains the telomeres, and activation of this enzyme is therefore required for cells to overcome replicative senescence and obtain the ability to divide without limits.Studies of the telomerase enzyme complex have revealed it consist of three subunits: a structural RNA component (hTR) that contains a template region that binds the TTAGGG repeats in telomeres, a catalytic subunits with reverse transcriptase activity (hTERT), and a telomerase-associated protein (hTPl). Purified hTERT mixed with hTR is sufficient to reconstitute telomerase activity in vitro. While hTR is constitutively present in normal and cancer cells. The original reports of hTERT cloning showed a correlation between hTERTmRNA expression and telomerase activity in several cell lines and tissues, after that, large studies have showed the close association, so hTERT become a new target in the cancer field.As the closely association of hTERT and cancer, this study is focused on hTERT, to explore its uses in diagnosis and treatment of cancer.In this study, we established an full length antisense hTERT gene eukaryotic expression vector, use it to block the wild type hTERTmRNA expression, and investigated the effects on the type and growth in human pulmonary giant cell carcinoma cell line (PLA-801D) and discuss its potential role in the gene therapy for cancers. Methods: An antisense hTERT gene eukaryotic expression vector pcDNA3.1(-)-hTERT including the full length region of hTERT CDS was constructed with recombinant DNA technique and transfected into the cells of human pulmonary giant cell carcinoma cell line (PLA-801D) with liposome Superfect. Then the expression of hTERTmRNA, P53 mRNA was observed with RT-PCR, telomerase activity with TRAP-ELISA, and cellular proliferation capacity with growth curve. Results: hTERT 3.4kb full length fragment joint into pcDNA3.1(-) vector in reverse orientation have been constructed and was successfully transfected into the PLA-801D cells, the PLA-801D cells of antisense hTERT transfected growth rate was suppressed, and the antisense hTERT gene can act as agents for specific inhibition to hTERTmRNA expression and after the telomerase activity was down-regulation by it. The cycle distribution was stopped at GO/G1 stage, the P53 mRNA expression was down regulation and the P53 protein was decreased. Conclusion: The results suggest that full-length antisense hTERT therapy specific to hTERTmRNA and telomerase activity is a practical approach to selective suppression of tumor growth; antisense hTERTmRNA could be used as an appropriate target for tumor therapy.Second, to find a drug to specific inhibited the hTERT mRNA and telomerase activity, we choice the Arsenic Trioxide which has conformed have good effection on the treatment of APL, and the study of its using on the other cancer therapy is now going on. To vestigate the effection of As2O3 on telomerase activity and human telomerase reverse transcriptase (hTERT), we choice Hela cell and PLA-801D cell as the target cells. Methods: The Helacells and the PLA-801D cells were cultured and exposed to 2umol/L As2O3 during different time, at the different time point, the cell's proliferation activity were measured by MTT, the distribution of cell cycle were determined by flow cytometry, the expression of hTERT mRNA were measured by RT-PCR, the activity of telomerase were measured by TRAP-ELISA. Results: The Hela cell's and PLA-801D cell's proliferation activity were gradually decreased after exposed to 2umol/L As2O...
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