Novel application of acetaldehyde dehydrogenase 2 gene
An acetaldehyde dehydrogenase and application technology, applied in the field of medical detection, can solve the problems of high heterogeneity of gastric cancer, few chemotherapy drugs, and delayed targeted therapy.
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Embodiment 1
[0047] Example 1: Establishment of gastric cancer cell lines with ALDH2 knockdown expression
[0048] Human gastric cancer cell SGC-7901 was purchased from the cell bank of Shanghai Institute of Cell Biology, Chinese Academy of Sciences, and embryonic kidney cell 293T was stored in the Institute itself. Cell culture conditions: SGC-7901 was cultured in RPMI-1640 cell culture medium containing 10% calf serum at 37°C and 5% CO2, and cells in logarithmic growth phase were used for experiments.
[0049] ALDH2 knockdown expression retrovirus vector construction, retrovirus packaging method steps as described above, using retrovirus infection of SGC-7901 cells for 48 hours, using the final concentration of 1 μg / ml puromycin to screen to obtain a stable knockdown expression cell line , using qRT-PCR, Western Blot and immunofluorescence methods to verify the knockdown effect. The results showed that small interfering RNA could significantly knock down the expression of ALDH2 in SGC-7...
Embodiment 2
[0051] Example 2: Drug Sensitivity Experiment of Gastric Cancer Cells with ALDH2 Knockdown Expression
[0052] The differences in the sensitivity of gastric cancer cells to the chemotherapy drugs 5-Fu, cisplatin and DOX after knocking down the ALDH2 gene were compared. We used gradient concentrations (0 μM, 0.01 μM, 0.1 μM, 1 μM, 10 μM and 100 μM concentrations) of several chemotherapy drugs to treat the ALDH2 knockdown expression gastric cancer cells SGC-7901 (ALDH2 shRNA1 and shRNA2 cells) from Example 1 and the control group. Gastric cancer cell SGC-7901 (shRNA empty vector), the relative survival rate of cells in each group was compared.
[0053] The study showed that the relative survival rate of ALDH2shRNA1 and shRNA2 cells was significantly lower than that of the shRNA empty vector group (P=0.0014, P=0.0082) after treating gastric cancer cells with DOX for 24 hours, suggesting that ALDH2 knockdown cells are more sensitive to DOX treatment. Gastric cancer cells were tre...
Embodiment 3
[0059] Example 3: Drug Sensitivity Study of Knockdown ALDH2 Gene Gastric Cancer Cells Inoculated in Nude Mice
[0060] In order to evaluate the difference in drug sensitivity of ALDH2 gene-deficient gastric cancer cells in vivo, we used immunodeficient nude mice for xenograft inoculation and experimental treatment studies. We used a total of 14 4-week-old BALB / C nude mice, and subcutaneously inoculated 3×106 ALDH2-shRNA1 and shRNA empty vector SGC-7901 cells in the left and right thighs of the nude mice, and the transplanted tumors grew to 150mm 3 , each of the experimental group and the control group were given intraperitoneal injection of 2 mg / Kg body weight of DOX and DMSO, respectively, twice a week, and the size of the tumor was measured before each injection of the drug, and treated for 3 consecutive weeks, and 3 days after the last injection of the drug The tumor size was measured and the mice were sacrificed. It was found that the tumor weight of subcutaneously transpl...
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