追毒方调控CD147糖基化介导的有氧糖酵解抑制三阴性乳腺癌增殖及侵袭迁移OA

Objective:To study the effects of Zhuidu(追毒)Formula(ZDF)on the proliferation,invasion,and mi-gration of triple-negative breast cancer(TNBC)MDA-MB-231 cells and explore the underlying molecular mechanisms.Methods:MTT assay,Transwell assay,and wound healing assay were employed to observe the effects of different concen-trations of ZDF on the proliferation,invasion,and migration of TNBC cells,respectively.Then,flow cytometry and the lac-tate assay kit were employed to evaluate glucose uptake,intracellular lactate production,and extracellular lactate efflux of TNBC cells treated with different concentrations of ZDF,thus assessing the aerobic glycolysis levels.Western blotting was performed to study the effects of ZDF on the glycosylation of cluster of differentiation 147(CD147,also known as the ex-tracellular matrix metalloproteinase inducer)and the protein levels of glucose transporter 1(GLUT1)and monocarboxy-late transporter 4(MCT4)in TNBC cells.To further confirm that ZDF regulation of aerobic glycolysis in TNBC cells was related to CD147 glycosylation,this study treated the cells with the glycosyltransferase GnT-V activator retinoic acid and observed its reversal effect on the inhibitory effect of ZDF.Results:Compared with the blank control group,the ZDF groups(0.1,0.2,0.4,0.8,1.6,and 3.2 μg/mL)showed reduced cell proliferation(P<0.01),with IC50 values of 5.00,0.25,and 0.02 μg/mL at the time points of 24 h,48 h,and 72 h,respectively.The ZDF groups(0.4,0.8,and 1.6 μg/mL)and the 2-DG(2 101 μg/mL)group showed weakened cell invasion and migration(P<0.01),reductions in CD147 glycosylation level(P<0.01),glucose uptake(P<0.01),and lactate content in cells and the supernatant(P<0.01),and down-regulated expression of aerobic glycolysis-related transmembrane proteins GLUT1/MCT4(P<0.01).Compared with the blank control group,the retinoic acid(30 μg/mL)group showed increases in glucose uptake,intracellular and supernatant lactate content,CD147 glycosylation level,and GLUT1/MCT4 expression(P<0.01 or P<0.05).Compared with the ZDF(0.8 μg/mL)group,the retinoic acid(30 μg/mL)+ZDF(0.8 μg/mL)group demonstrated up-regulation of the aforementioned indicators(P<0.01 or P<0.05).Conclusion:ZDF regulates CD147 glycosylation to suppress GLUT1 and MCT4 expression and down-regulate aerobic glycolysis-mediated glucose flux and lactate production and efflux,thus inhibiting the proliferation,invasion,and migration of TNBC.