| Objective: Diabetes is one of the diseases that seriously harm human health today,among which Type 2 diabetes(T2DM)is the majority.Insulin resistance(IR)in peripheral tissues such as muscle and fat is the main pathogenesis of T2 DM.Insulin receptor substrate-1(IRS-1)is an important mediator of insulin information transmission.The study found that the expression of IRS-1 decreased in both IR and T2 DM patients,while Gammaaminobutyric acid(GABA)can up-regulate the expression of IRS-1 and improve IR,but the molecular mechanism of GABA regulating IRS1 is still unclear.Histone deacetylases(HDACs)can inhibit transcription of corresponding genes by catalyzing histone deacetylation.Studies have shown that the expression of IRS-1 is regulated by HDAC2/3.Other studies have reported that GABA can inhibit HDACs,suggesting that GABA may up-regulate the expression of IRS-1 by inhibiting HDAC2/3.However,the specific molecular mechanism of GABA inhibiting HDAC2/3 remains unclear.We predicted the microRNA(mi RNA)related to HDAC2/3 through bioinformatics,in which the expression of mi R-34 a can be up-regulated by GABA.We also found that the promoter region of mi R-34 a may be associated with the transcription factor CCAAT/enhancer binding protein alpha(CEBPα),suggesting that CEBPα can up-regulate the expression of mi R-34 a.This study combined with in vivo and in vitro experiments to explore whether GABA can pass CEBPα However,mi R-34 a was up-regulated,which inhibited the expression of HDAC2/3,and finally antagonized the decrease of IRS-1 expression in T2 DM.Methods: T2 DM model mice were established with high-fat diet combined with streptozotocin.The T2 DM model mice were randomly divided into T2 DM group and T2DM+GABA group,and the normal mice were divided into control group and GABA group,with 8 mice in each group.The mice in each group were fed with maintenance diet.GABA group and T2DM+GABA group drank distilled water containing 0.2% GABA,while control group and T2 DM group drank distilled water.Keep feeding for 4w.At the end of the experiment,the mice in each group were fasted overnight,then they were killed by cervical spondylolisthesis.The abdominal adipose tissue was quickly dissected and placed in liquid nitrogen,and stored in the-80℃ refrigerator for use.The expression of IRS-1,HDAC2,HDAC3,mi R-34 a and CEBPα in adipose tissue were analyzed by quantitative real-time PCR(q RT-PCR)and Western blot(WB).3T3-L1 cell line was cultured in vitro.3T3-L1 cells treated with 20 m M glucose were used as T2 DM model cells,and 10 n M GABA was used for intervention.The expression levels of IRS-1,HDAC2,HDAC3,mi R-34 a and CEBPα were analyzed by q RT-PCR and WB.The expression levels of CEBPα and mi R-34 a in 3T3-L1 cells were silenced,and the expression levels of CEBPα and mi R-34 a were analyzed by q RT-PCR and WB.3T3-L1 cells were treated with mi R-34a-5p mimetics and inhibitors respectively,and the control group was set up at the same time.The expression of HDAC2/HDAC3 were analyzed by q RT-PCR and WB methods.Silence HDAC2 in 3T3-L1 cells,and set up a control group at the same time.The expression of HDAC2 and IRS-1 were analyzed by q RT-PCR and WB methods.Silence HDAC3 in 3T3-L1 cells,and set up a control group at the same time.The expression of HDAC3 and IRS-1 were analyzed by q RT-PCR and WB methods.Results: 1.Compared with the control group,the expression of IRS-1,mi R-34 a and CEBPα in adipose tissue of mice in T2 DM group were decreased(P<0.01),and the expression of HDAC2 and HDAC3 were increased(P<0.01).Compared with T2 DM group,the expression of IRS-1,mi R-34 a and CEBPα in adipose tissue of mice in T2DM+GABA group were increased(P<0.01),and the expression of HDAC2 and HDAC3 were decreased(P<0.01).2.Compared with the control group,the expression of IRS-1,mi R-34 a and CEBPα in T2 DM group were decreased(P<0.01),and the expression of HDAC2 and HDAC3 were increased(P<0.01).Compared with T2 DM group,the expression of IRS-1,mi R-34 a and CEBPα were increased in T2DM+GABA group(P<0.01),and the expression of HDAC2 and HDAC3 were decreased in T2DM+GABA group(P<0.01).3.Compared with the control group,the expression of mi R-34 a and CEBPα in si-CEBPαgroup were decreased(P<0.01).4.Compared with the control group,the expression of HDAC2 and HDAC3 in mi R-34 a mimic group decreased(P<0.01),and the expression of HDAC2 and HDAC3 in mi R-34 a inhibitor group increased(P<0.01).5.Compared with the control group,the expression of HDAC2 in si-HDAC2 group decreased(P<0.01),while the expression of IRS-1 increased(P<0.01).Compared with the control group,the expression of HDAC3 in si-HDAC3 group decreased(P<0.01),while the expression of IRS-1 increased(P<0.01).Conclusion: GABA can increase the expression of CEBPα,up-regulate mi R-34 a,then inhibit the expression of HDAC2/3,and finally up-regulate the expression of IRS-1 in T2 DM. |
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