Study on the protective mechanism of Shenshuai Xiezhou decoction on rat aortic VEC cells based on miRNA126 regulation of PI3K/AKT/mTOR pathway
Objective Based on miRNA126 regulating PI3K/AKT/mTOR pathway,we investigated the mechanism of Vascular Endothe-lial Cell(VEC)protection by Shenshuaixiezhuo Decoction at cellular and molecular levels to delay atherosclerosis in Chronic Kidney Dis-ease(CKD).Methods A rat model of chronic kidney disease combined with atherosclerosis was prepared,and 50 SD rats were random-ly divided into the model group,the chlorthalidomide group,and the low,medium and high dose groups of Shenshuaixiezhuo Decoction,with 10 rats in each group,and another 10 rats were taken as the sham-operation group.Each group was treated with the corresponding drugs for 8 weeks and then executed.Separate serum was prepared from the blank control group,model group,cloxacillin group,and low,medium and high dose groups of Shenshuaixiezhuo Decoction to intervene in the VEC cells of rat aorta.The cell morphology was ob-served by light microscopy and Hoechst 33258 staining;the proliferation and apoptosis levels of cells in each group were detected by flow cytometry;and the expression levels of miRNA126-regulated PI3K/AKT/mTOR pathway mRNAs and proteins were detected by qPCR and WB.Results Morphological observation after cell culture showed that more floating cells appeared in the model group compared with the sham-operated group,and the cell morphology of each dosing group improved compared with that of the model group,among which the medium-dose group of Shenshuaixiezhuo Decoction showed better performance;the proliferation activity of cells in the model group was significantly reduced(P＜0.01)and the level of apoptosis was significantly increased(P＜0.01)compared with that of the blank control group,and the level of proliferation and apoptosis in each dosing group improved compared with that of the model group;and the level of proliferation and apoptosis in each dosing group was improved compared with that of the model group.Compared with the model group,the expression of miRNA126 mRNA in the model group decreased significantly(P＜0.05),and the expression levels of PI3K,AKT,and mTOR genes and proteins increased significantly(P＜0.05);compared with the model group,the expression levels of miR-NA126 mRNA in the Cloxartan group,and the Shenshuaixiezhuo Decoction low,medium,and high dose groups increased significantly(P＜0.05).Compared with the model group,miRNA126 mRNA expression increased significantly in the low,medium and high dose groups of chlorosartan group and Shenshuaixiezhuo Decoction group(P＜0.05);PI3K,AKT,mTOR gene and phosphorylated protein expression levels decreased in all groups of Shenshuaixiezhuo Decoction(P＜0.05),and autophagy-associated Bcelin1 protein expres-sion level increased(P＜0.05).Conclusion Shenshuaixiezhuo Decoction may promote cellular autophagy pathway through miRNA126 regulation of PI3K/AKT/mTOR pathway,maintain aortic VEC cellular homeostasis,and play a role in protecting VEC and delaying ath-erosclerosis in chronic kidney disease combined with atherosclerosis.
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