c-Myc regulates glucose transporters 4 to activate the janus kinase/signal transducer and activators of transcription to promote triple negative breast cancer
Objective To analyze the mechanism of c-Myc regulating glucose transporters 4(GLUT4)to activate janus kinase(JAK)/signal transducer and activators of transcription(STAT)to pro-mote the progression of triple negative breast cancer(TNBC),and to explore new therapeutic targets.Methods The relationship between GLUT4 and c-Myc and the clinicopathological features of breast cancer were analyzed by bioinformatics.Clinical samples of TNBC patients were collected,and the expression lev-els of GLUT4 and c-Myc were detected by immunohistochemistry.MDA-MB-231 cell line with c-Myc knockout was constructed,and the expression of GLUT4 was detected by real-time quantitative reverse tran-scriptase-polymerase chain reaction(RT-qPCR)and Western blotting.Cell counting kit-8(CCK-8)assay was used to detect proliferation,flow cytometry was used to detect apoptosis,and Transwell assay was used to detect migration and invasion.Western blotting was used to further analyze the effect of c-Myc regulating GLUT4 on JAK/STAT.The measurement data of the two groups of normal distribution data were compared using the independent sample t test,and the data between numerous groups were compared using analysis of variance.Results Bioinformatics analysis showed that GLUT4 was significantly down-regulated in breast cancer(P＜0.01)and correlated with TNM stage,pathological grade and PAM50 subtype.Accord-ing to the immunohistochemical score,the expression levels of c-Myc in TNBC cancer tissues were signifi-cantly higher than paracancerous tissues(7.233 vs.4.567,t=4.551,P＜0.01),while GLUT4 expres-sion was significantly lower(4.800 vs.7.933,t=5.702,P＜0.01).When c-Myc was knocked out in MDA-MB-231 cells,RT-qPCR results showed that the expression of GLUT4 in c-Myc knockout group was higher than that in negative control group(2.486±0.273 vs.1.070±0.140,F=55.005,P＜0.01).Western blotting results showed that the expression of GLUT4 in c-Myc knockout group was higher than that in negative control group(0.691±0.041 vs.0.431±0.055,F=29.027,P＜0.01).MDA-MB-231 cell line with low GLUT4 expression was constructed,and RT-qPCR showed that there were significant differences in the GLUT4 expression between GLUT4 silencing group and negative control group(0.507±0.076/0.137±0.025/0.317±0.085 vs.1.150±0.200,F=45.016,P＜0.01).CCK-8 results showed that the cell survival rate increased after GLUT4 knockdown(128.614±13.780 vs.95.875±9.130,F=26.927,P＜0.01),and the cell survival rate was relatively reduced after further knocking out c-Myc(70.977± 5.906 vs.95.875±9.130,F=26.927,P＜0.01).The results of flow cytometry showed that apoptosis was increased after knocking out GLUT4 and c-Myc simultaneously(12.170±1.283 vs.3.263±0.691,F=102.946,P＜0.01).Transwell results showed that the number of migrating cells[(237.333±14.364)cells vs.(156.667±10.693)cells,F=91.074,P＜0.01],and the number of invasive cells[(257.000± 9.539)cells vs.(152.333±5.132)cells,F=90.437,P＜0.01]in GLUT4 knockdown group were signifi-cantly greater than those in the negative control group.After knocking out GLUT4 and c-Myc simultaneously,the number of migrating cells[(67.000±14.177)cells vs.(156.667±10.693)cells,F=91.074,P＜0.01]and the number of invasive cells[(91.667±13.650)cells vs.(152.333±5.132)cells,F=90.437,P＜0.01]were significantly less than those in the negative control group.Western blotting results showed that the expression of c-Myc(0.814±0.060 vs.0.447±0.028),p-JAK2(0.629±0.038 vs.0.380±0.030)and p-STAT3(0.698±0.029 vs.0.288±0.016)was up-regulated after GLUT4 knock-out(F=17.541,P＜0.01).The GLUT4 expression was relatively increased after c-Myc knockdown(0.618±0.029 vs.0.529±0.049,F=68.772,P＜0.05),and the expression of p-JAK2(0.510± 0.033 vs.0.629±0.038)and p-STAT3(0.410±0.050 vs.0.698±0.029)decreased(F=68.772,P＜0.01).Conclusion c-Myc has the ability to suppress GLUT4 expression and control the JAK/STAT to limit the growth,invasion,and death of TNBC cells.
Triple negative breast cancerC-MycGlucose transporters 4Janus kinase/signal transducer and activators of transcription signaling pathway
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