丹蛭降糖胶囊通过调控miR-144-3p/DUSP1轴抑制高糖诱导的肾小管上皮细胞EMTOA

AIM:Based on bioinformatic analysis,this study aims to investigate the effects of Danzhi-Jiang-tang capsule(DJC)on high glucose(HG)-induced epithelial-mesenchymal transition(EMT)in renal tubular epithelial cells and the underlying mechanisms of microRNA-144-3p(miR-144-3p)/dual-specificity phosphatase 1(DUSP1)axis.METHODS:Differentially expressed genes in renal tissues from patients with diabetic kidney disease were identified using the Gene Expression Omnibus(GEO)database.Key genes were further screened using three machine learning algo-rithms:least absolute shrinkage and selection operator(LASSO)regression,support vector machine-recursive feature elimination(SVM-RFE),and random forest(RF).The final candidate gene was determined based on receiver operating characteristic(ROC)curve analysis,and its upstream miRNAs were predicted.An HG-induced cell model was estab-lished using renal tubular epithelial cells for in vitro experiments.The cells were divided into the following groups:normal glucose,mannitol,HG,and HG+DJC-containing serum(HG+DJC).Cell viability was determined by CCK-8 assay,and the apoptosis was analyzed by flow cytometry.The targeting relationship between miR-144-3p and DUSP1 was validated by dual-luciferase reporter assay.The protein levels of DUSP1 and EMT-related markers were measured by Western blot,and the expression of miR-144-3p and DUSP1 mRNA was detected by RT-qPCR.RESULTS:Diabetic kidney disease-related datasets were screened from the GEO database,and 97 differentially expressed genes were obtained.The three machine learning methods and ROC curve results were used to select DUSP1 as the research object and predict the upstream miR-144-3p.In HG-induced renal tubular epithelial cells,miR-144-3p was up-regulated,whereas DUSP1 mRNA and protein were down-regulated.The dual-luciferase reporter assay validated a targeting relationship between miR-144-3p and DUSP1.Inhibition of miR-144-3p significantly enhanced the viability of HG-induced renal tubular epithelial cells,re-duced apoptosis,increased the level of E-cadherin(E-Cad)protein,and decreased the levels of Snail,N-cadherin(N-Cad),vimentin(VIM)and α-smooth muscle actin(α-SMA)proteins(all P<0.05).Treatment with DJC-containing se-rum significantly suppressed miR-144-3p expression,up-regulated DUSP1,elevated E-Cad level,and reduced the levels of Snail,N-Cad,VIM and α-SMA in HG-induced renal tubular epithelial cells(all P<0.05).CONCLUSION:The DJC can inhibit HG-induced EMT in renal tubular epithelial cells,potentially through the miR-144-3p/DUSP1 axis.