Survival And Metabolism Effects Of Stenabolic On Human T98G Cells

ObjectiveTo investigate the effects of SR9009 on T98 G cell proliferation,cell viability,metabolic activity,and treatment time,and to elucidate the potential mechanism of its role.MethodsHuman T98 G cells were cultured in vitro and Hep G2 cell lines were used as controls,which were compared with DMSO,SR9009 and bortezomib(BOR)after incubation.The activity of T98 G cells after SR9009 treatment was determined by MTT assay,and the sensitive effect period of SR9009 or BOR on the activity of T98 G cells was also measured.The two drugs SR9009 and BOR were combined on T98 G cells to observe their synergistic effects.The cell cycle of SR9009-treated T98 G cells was determined by flow cytometry.The REDOX status of T98 G cells treated with SR9009 was detected by fluorescent tracer probe luciferin diacetate(FDA).LD levels in SR9009 or BOR treated cells were determined by flow cytometry with Nile red staining and confocal microscopy,and glutamine synthase,vimentin,GFAP,REV-ERB beam were detected by immunohistochemistry.ResultsAfter the T98 G cells treated with SR 9009 were incubated at concentrations of 20 and 40μM for 48-72 h,the cell vitality decreased significantly.When SR9009(20μM concentration)treated T98 G cells for 48 h,MTT assay was used to analyze the cell vitality.The results showed that there was an obvious time-dependent effect of drug treatment,and the inhibition effect was the highest between 18 and 30 hours,which showed that the cell vitality was the lowest during this time period(P<0.05).The activity of T98 G cells treated withBOR(500n M)was the lowest between 12 and 24 h after culture(P<0.01).The cell activity of Hep G2 cells treated with SR9009 was also significantly reduced.Flow cytometry cell cycle analysis showed that 58% of T98 G cells treated with SR9009 were in G0/G1 phase,which was higher than 43% of those treated with DMSO(P<0.01).In contrast,the proportion of S-phase cells in DMSO-treated cells was higher than that in SR9009 treated cells(P<0.05).Flow cytometry measured and analyzed ROS levels and found that ROS levels in SR9009-treated cells were significantly lower than those in DMSO control group(P<0.01).ROS levels of T98 G cells treated with BOR(500n M)increased(P<0.05).In addition,compared with the DMSO group,the ROS level of Hep G2 cells treated with SR9009(40μM)for 96 h also decreased significantly(P<0.01).Nile red staining showed that the average LD size and content of SR9009 and BOR treated cells were higher than that of DMSO treated cells,while the combination of the two drugs showed a synergic effect(P<0.01).In addition,compared with the DMSO group,the LD level of Hep G2 cells was also increased after SR9009 treatment(P<0.001).ConclusionsSR9009 has obvious anti-tumor activity in T98 G cells,showing obvious cytotoxic effect and affecting the metabolic level of tumor cells,thus reducing the survival rate of tumor cells.The mechanism may be that SR9009 regulates the intrinsic clock component protein REV-ERB.