| Zilongjin (ZLJ) is an effective folk recipe against cancers in our country. We discovered via repeated experiments that it was not only able to produce G2 arrest but also able to influence the growth, proliferation, invasiveness and correlative gene expression of Human Gastric Carcinoma BGC-823 cells as well. With an aim to explore ZLJ's anticancer effects at molecular level, we detected the gene differential expression of Human Gastric Carcinoma BGC-823 by cDNA Array, flow cytometry, RT-PCR and Western-blot.1. The subjects of the thesis were focused on the following aspects: 1.1. The effect of ZLJ on the growth, proliferation of Human Gastric Carcinoma BGC-823 cells:1.1.1. Growth curve: The influences of ZLJ, HMBA on proliferation of Human Gastric Carcinoma BGC-823 cells and Human K562 cells were observed.1.1.2. Colony formation experiment: The influence of ZLJ on colony formation of Human Gastric Carcinoma BGC-823 was investigated.1.2.Gene differential expression of Human Gastric Carcinoma BGC-823 cells was detected by cDNA microarray:Gene differential expression of Human Gastric Carcinoma BGC-823 cells was detected by cDNA microarray: The growth conditions of Human Gastric Carcinoma BGC-823 cells were in rich medium R ( DMEM supplemented with 15% FBS) and were put in 5% carbon dioxide incubator at 37℃. Cell synchronization: The cells were synchronized with thymidine ( 2.5mM ) and N2O (5.5Kg/cm2), and then classified into two groups: I . ZLJ group: The growth conditions of the cells were in rich medium R( DMEM supplemented with 15% FBS , containing 0.5mgZLJ/ml), begun at three hours after the second release from thymidine, and were put in 5% carbon dioxide carbon Incubator at 37℃ for seven hours; II .control group: Rich medium R( DMEM supplemented with 15% FBS) was renewed without being treated with ZLJ, and other growth conditions were the same as ZLJ group. The effect of synchronization and the influence of ZLJ on G2 phase were observed by flow cytometry, and then Mitotic index was calculated to figure out the percentage of G2 phase and M phase. cDNA microarray and data analysis: This part included total RNA isolation, DNase treatment of Total RNA, cDNA Probe Synthesis, Column Chromatography, Hybridization, Exposing the Atlas Array to a phosphorimaging screen and Analysing results by software of Array Gauge V1.0.1.3. some of differential expression genes ( cyclin E2, p16 ) were analysed:Analysis of cyclin E2 and p16 genes: I . RT-PCR: Total RNA was used for first-strand cDNA synthesis, then PCR was performed and 3 -actin gene was set as endogenous control. The ratios of gene expression in ZLJ group versus control samples were calculated from the data normalized against the endogenous control, and the ratios were compared with the results of cDNA microarray. II. Western blot: It involved extracting and measuring proteins, electrophoresing proteins, blotting them to a membrane, and probing the blot with a specific antibody (cyclin E2 or p16 ) to detect a particular protein; the antibody was detected with a labeled secondary antibody, adding luminescence substrate, covering it with Plastic membrane, immediately exposing to film ,developing and fixation. 3 -actin protein was set as endogenocontrol. The density of strips was measured by chemilmager?5500 scaningapparatu.The antisense oligonucleotide approaches of p16 mRNA: The study was designed to block the translation of p16 mRNA by using antisense oligonucleotides that are complementary to the upstream region that included the first codon AUG and to the terminal site of the coding region. Antisense oligonucleotides were modified by sulfur during synthesis. Western blot was performed to detect the influence of p16 mRNA antisense oligonucleotides on p16 protein, and B-actin protein was set as endogenous control. Flow cytometry was used to detect the change of cell cycle.1.4. The effect of ZLJ on the invasiveness of Human Gastric Carcinoma BGC-823 cells:1.4.1. The influence of ZLJ on invasiveness of Human Gastric...
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