Résumé
Objective@#To explore whether PIK3R3 is dependent on PPARA to regulate fat metabolism and proliferation of liver cancer cells.@* Methods@#The PIK3R3 overexpression plasmids and its specific siRNA were transfected into hepatocellular carcinoma cells HepG2 and SMMC-7721. The level of PPARA was detected by Western blotting（WB）and the ketone body content in supernatant was measured by colorimetric method.DNA synthesis and proliferation of hepatocellular carcinoma cells were detected by BrdU/PI incorporation method and CCK8 method. Then we used PIK3R3 overexpression or silenced hepatocellular carcinoma cell lines to observe the effect of PIK3R3 on the proliferation of hepatocellular carcinoma cells by plate cloning experiments. We intervened the expression of PIK3R3 and then observed the lipid metabolism and cell proliferation，so as to determine whether the role of PIK3R3 in hepatocellular carcinoma cells depends on PPARA. Finally, we added MG132（proteasome inhibitor）or CHX（cycloheximide）to PIK3R3 overexpression or silencing hepatocellular carcinoma cells to detect the level of PPARA by WB，next we constructed the plasmids of PPARA promoter luciferases and observed the effect of PIK3R3 on the activity of PPARA promoter. @*Results@#PIK3R3 influenced the expression of PPARA，lipid metabolism and proliferation of hepatocellular carcinoma cells，and PPARA partly reversed the effects of PIK3R3.@*Conclusion@#PIK3R3 can regulate the fat metabolism and proliferation of hepatoma cells by regulating the expression of PPARA.
Résumé
Objective:To investigate the effect and the related mechanism of c-Myc on the proliferation,invasion and migration ability of malignant melanoma B16-F10 cells. Methods:We detected the expression of PIK3R3 in malignant melanoma and normal tis-sues. Efficiency of gene silencing of c-Myc and PIK3R3 was determined by qPCR and Western blot. We detected the proliferate ability of B16-F10 cells after silencing c-Myc and PIK3R3 using EdU assay. We detected the migration and invasion ability of B16-F10 cells after silencing c-Myc and PIK3R3 using wound healing assays and Transwell matrigel invasion assays. The expression of miR-199a after silencing c-Myc and PIK3R3 using qPCR. The expression of c-Myc and PIK3R3 was detected by qPCR after transfecting miR-199a mimics or miR-199a inhibitor. Dual-luciferase reporter assay system was used to detect miR-199a regulating c-Myc and PIK3R3. Results:Compared normal skin tissue,expression of PIK3R3 was significantly increased in malignant melanoma tissue;after silencing c-Myc and PIK3R3 gene,the proliferation,invasion and metastasis of melanoma cell line B16-F10 were significantly reduced;expression of miR-199a upregulated after silencing c-Myc and PIK3R3 genes, expression of c-Myc and PIK3R3 decreased after transfecting miR-199a mimics, expression of c-Myc and PIK3R3 upregulated after transfecting miR-199a inhibitor, dual luciferase reporter system test results revealed miR-199a can directly regulate c-Myc and PIK3R3 transcription activity. Conclusion: miR199a regulated the expression of c-Myc,then promote proliferation,migration and invasion in malignant melanoma cells.
Résumé
Objective:To explore the role ofinterleukin (IL)-6 in gastric cancer cells and the mechanisms.Methods:Gastric cancer cells MGC-803 were treated with 50 ng/mL of recombinant IL-6 protein,and then cell viability and cell migration were detected by MTT assay and wound-healing assay,respectively.The mRNA and protein expressions of E-cadherin,N-cadherin,vimentin,Snaill and miR-152 were analyzed by RT-qPCR and Western blot,respectively.Moreover,MGC-803 cells were simultaneously or separately treated with IL-6 and transfected with miR-152 mimics,and then the mRNA expression of PIK3R3 and the protein levels of PIK3R3,Akt and p-Akt were determined.Results:IL-6 stimulation significantly promoted cell proliferation and migration,reduced the expression of E-cadherin and miR-152,and increased the expression of N-cadherin,vimentin,Snaill,PIK3R3 and p-Akt (All P＜0.05).The protein levels of PIK3R3 and p-Akt were significantly decreased after transfecting miR-152 mimics into MGC-803 cells (P＜0.01).miR-152 overexpression down-regulated IL-6-induced the protein expression of PIK3R3 and p-Akt (P＜0.01).The levels of Akt in each group were not changed.Conclusion:IL-6 up-regulates PIK3R3 expression and activates PI3K/Akt signaling pathway through down-regulating miR-152 expression,which consequently promotes gastric cancer cell proliferation,migration,and epithelial-mesenchymal transition.