The Studies Of The Effect Of ATP5B Expression On The Invasion And Metastasis Of Tumor Cells

Tumor disease has become a major threat to human survival and health.Metastasis is the main cause of death in patients with malignant tumors.About 90% of patients with malignant tumors die of tumor metastasis.The process of tumor metastasis is affected by many factors,including the attack of immune cells,blood mechanical action,organ micro-environment,and tumor cell viability in the circulation.Metastasis is the most dangerous stage in the development and evolution of malignant tumors.Therefore,understanding the mechanism of invasion and metastasis of malignant tumors,finding appropriate blocking pathways and potential therapeutic targets play an important role in inhibiting the development of malignant tumors phage-displayed 7-mer peptide library to screen the target peptides.In the previous work of the research group,we used normal prostate epithelial cells,prostate cancer low metastatic cell lines: PC-3 cells,and prostate cancer high metastatic cell lines: PC-3M cells.We performed phage-displayed 7-mer peptide library for subtractive screening.The transfer-related short peptide that specifically binds only to PC-3M: B04,and found that the short peptide B04 can effectively inhibit the proliferation and metastasis of PC-3M cells.Subsequently,the receptor protein of the short peptide B04 was detected as ATP5 B on the PC-3M cell membrane by proteomics and other methods,and B04 was found to inhibit its invasive ability by specifically inhibiting ATP5 B expressed on the PC-3M cell membrane.Cholesterol(Chol)can promote the mitochondrial ectopicity of ATP5 B in PC-3M cells to the cell membrane.The membrane ATP5 B promotes prostate cancer invasion and metastasis by activating C-Myc proto-oncogene and upregulating VEGFA.Therefore,in this experiment,we mainly investigate whether ATP5 B is ectopically expressed on the cell membranes of other cells and the effect of ATP5 B expression on the cell migration and invasion ability of the cell membrane.Results:First of all,Expression of ATP5 B in human breast cancer cells MDA-MB-231 and MCF-7,human cervical cancer HELA and SIHA,human ovarian cancer cells H08910 and SKOV3,human gastric cancer cells BGC823 and SGC7901 cellular localizationATP5B was stained in MDA-MB-231,MCF-7,HELA,SIHA,H08910,SKOV3,BGC823,and SGC7901 cells by immunofluorescence staining.The results illustrate that ATP5 B was expressed on the surface of these eight cell membranes.Subsequently,the expression of ATP5 B on various cell membranes was further examined by flow cytometry.Flow cytometry results showed that the percentage of ATP5 B positive expression on the cell membrane of MDA-MB-231 and MCF-7 cells and the fluorescence intensity were 25.57%±0.14% vs44.58%±3.28%,23.79±0.12vs48.3±5.09,and statistical analysis showed that the percentage of positive expression of ATP5 B and the fluorescence intensity of the two cell membranes were significantly different(P<0.05).<0.05).The above results indicated that ATP5 B was expressed on the cell membranes of MDA-MB-231 and MCF-7,and the expression level on MDA-MB-231 cell membrane was significantly higher than that on MCF-7 cells.Similarly,the percentage of positive expression of ATP5 B on the cell membranes of HELA and SIHA cells and the fluorescence intensity were 95.55±1.09%,79.79%±1.97%,and 58.79±8.54 vs 33.62±6.01,respectively.The above results showed that ATP5 B was expressed on the cell membranes of HELA and SIHA,and the expression level on HELA cell membrane was significantly higher than SIHA cells.The percentage of positive expression of ATP5 B on the cell membranes of H08910 and SKOV3 cells and the fluorescence intensity were 99.26%±0.39% vs 68.16%±1.12% and 166.33±1.1vs62.66±1.85,respectively.The above results reveal that ATP5 B was expressed on the cell membranes of H08910 and SKOV3,and the expression level on H08910 cell membrane was significantly higher than that on SKOV3 cells.The percentage of positive expression of ATP5 B on the cell membrane of BGC823 and SGC7901 cells and fluorescence intensity were 92.96±0.1VS81.83±0.25 and 44.16±0.46vs99.28±0.27,respectively.The above results showed that ATP5 B was expressed on BGC823 and SGC7901 cell membranes,and the expression level on BGC823 cell membrane was slightly higher than that on SGC7901 cells.However,the fluorescence intensity of ATP5 B on SGC7901 cells was significantly higher than that of BGC823 cells.Secondly,The effect of ectopic expression of ATP5 B on cell invasion and metastasis Migration and invasion of human breast cancer cells MDA-MB-231 and MCF-7,human cervical cancer cells HELA and SIHA,human ovarian cancer cells H08910 and SKOV3,human gastric cancer cells BGC823 and SGC7901 related researchThe cell migration assay and invasion assay were performed using cell scratch assay and Transwell chamber migration/invasion assay.The results showed that MDA-MB-231 migrated in vitro in human breast cancer cells MDA-MB-231 and MCF-7.The invasion ability was higher than that of MCF-7(p<0.05).In human cervical cancer HELA and SIHA,the ability of HELA to migrate and invade in vitro was higher than that of SIHA(p<0.05);The ability of human ovarian cancer cells H08910 to migrate and invade in vitro was higher than that of SKOV3(p<0.05).The in vitro migration and invasion of human gastric cancer cells BGC823 and SGC7901 were not statistically significant(p>0.05).The above results show that in each group of cells,cells with high ATP5 B expression on the cell membrane surface have a generally high in vitro migration and invasion ability.The above results show that in human breast cancer,human cervical cancer,human ovarian cancer and other cell lines,different cell lines of the same tumor have different in vitro migration and invasion abilities,and the ectopic expression of ATP5 B on the cell membrane surface is high in those with strong in vitro migration and invasion ability.The ectopic expression of ATP5 B may serve as a potential indicator of tumor cell invasion and migration.