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Suppression of Tumor Growth Induced by siRNA-PARP Combined Radiotherapy

Author: WangHongYong
Tutor: WangTieJun；ChenYuBing
School: Jilin University
Course: Clinical
Keywords: RNA interference(RNAi) poly(ADP-ribose) polymerase (PARP) radiotherapy anti-tumor effect
CLC: R73-36
Type: Master's thesis
Year: 2009
Downloads: 70
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Abstract


Epidemiological investigation shows that the incidence and mortality of malignant neoplasms has gradually increased and the prevention and treatment of malignant tumors has become an important topic with the continuous advancement of the economy, the environment of industrialization and urbanization, demographic composition of the population aging, unhealthy lifestyle and behavior. The lung cancer fastest rise in the malignant tumors.The lung cancer has been the most common malignant tumor in our country and is leading one of the causes of death in the whole world, which severely enrangers human health and life. Radiotherapy is the most important means of lung cancer’s treatment, but it can arise a series of questions and seriously affect their efficacy and clinical application. The radiotherapy breaks DNA double-strand, so that tumor cells can not be copied, but the repair of DNA’s damage will affect the treatment efficacy. In recent years,some studies have been shown that the adoption of specific blockers can inhibit the tumor cell DNA repair and enhance radiosensitivity of tumor cells to achieve the purpose of tumor cure. PARP (poly adenosine diphosphate - ribose polymerase) has been chosen as a target gene because it is a crucial role in the DNA repair pathways.RNA interference (RNAi) has been shown to interfere the expression of correlating mRNA in various species. RNAi has been an important tool for understanding gene function and attempting gene therapy.Object The study was constructed the recombinant plasmid pGPU6/GFP/Neo-shRNA as vector carring the PARP siRNA gene into Lung cancer Lewis cell to inhibit the expression of PARP in cell. We investigate the effect of gene hTERT siRNA transfection into Lewis cells, such as cells growth, proliferation, and apoptosis.Using RNAi interfered PARP gene with radiotherapy in our study, we explore to determine potential of PARP as a therapeutic target for lung cancer by potent highlysequence-specific RNAi technique, and to investigate the effect of PARP gene silencing on the growth of Lewis cell in vitro. To evaluate clinical value of siRNA interfered PARP gene may be become therapeutic applications in treating lung cancer. Methods Constructing the recombinant plasmid pGPU6/GFP/Neo-shRNA as vector, it carried the PARP siRNA gene into Lewis cell in vitro. The change of cell cycle distribution and apoptosis of Lewis cells was detected by FCM(flow-cytometry), electron microscopeand, and gel electrophoresis. Cell livability was checked by MTT method. The PARP protein level of lewis cell was measured by Western blot methods and the PARP expression in Lewis cell was detected by RT PCR methods. We use the factor analysis of variance in groups.Results The recombinant plasmid pGPU6/GFP/Neo-shRNA were constructed successfully, and the completion of the amplification and identification of plasmid. We carried the plasmid into Lewis cells and gave X-ray exposure. Compared with control group,the cells of PARP-siRNA combined radiotherapy were slower,but apoptosis on the contrary. MTT method and FCM showed that more Lewis cells proliferated effectively slowlier and apoptosis induced with PARP siRNA combined radiotherapy(p<0.05).And then the expression of PARP of that group significantly decreased at protein level in vitro(p<0.05).Conclusions The specific small hair RNA target PARP gene could inhibit the expression of PARP mRNA and the proliferation of Lewis cells, adjust cell cycle and promote apoptosis of lung cancer.It improve the radiosensitive as a target gene.

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