The Correlation Study Of MicroRNA Targeting Regulation Of ACE-AngⅡ-AT1 Axis In The Syndrome Of Yin Deficiency And Yang Hyperactivity In Essential Hypertension

Objective:To determine the relationship between plasma levels of ACE-AngⅡ-AT1 axis related signaling factors and their targeted micrornas.Based on ACE-AngⅡ-AT1 axis,to investigate the differences of targeted micrornas between essential hypertension with Yin deficiency and Yang excess syndrome,non-yin deficiency and Yang excess syndrome and healthy subjects,and to explore the possible correlation between the differentially expressed micrornas and the disease with Yin deficiency and Yang excess syndrome.Methods:(1)From March 2022 to October 2022,92 outpatients and inpatients with essential hypertension were collected from the Department of Cardiovascular Medicine of Anhui Provincial Hospital of Traditional Chinese Medicine and Anhui Provincial Acupuncture and Moxibustion Hospital,including 24 cases of Yin deficiency and Yang excess syndrome,hyperactive liver fire syndrome,phlegm dampness syndrome,and 20 cases of both Yin and Yang deficiency syndrome.Twenty-four normal subjects were collected from the Physical examination Center of Anhui Provincial Hospital of Traditional Chinese Medicine.(2)5ml venous blood was collected on an empty stomach and stored in a test tube without anticoagulation.Immediately after self-coagulation at room temperature,the blood plasma was collected in a water bath at 37℃ for 20 min,centrifuged at room temperature for 3000r·min-1 for 10 min,and frozen at-30 ℃.(3)The target genes of miRNAs were predicted online using two target gene databases,Targetscan and miRDB.The target gene sets obtained by the two target gene prediction software were analyzed by GO function and KEGG enrichment(KEGG Pathway)in the DAVID database.The target miRNAs of ACE-AngⅡ-AT1 axis were identified.(4)The plasma levels of ACE,AngⅡ and AGTR1 were determined by ELISA-Quantikine kit(R&D Systems,Minneapolis,MN,USA),according to the instructions.(5)Real-time quantitative PCR was used to determine the relative expression levels of miR-145-5p and miR-126-3p.The reverse primers were provided by the kit,and the forward primers were synthesized by Shanghai Tianhao Biotechnology Co.,LTD.There were three replicates for each reaction.Results:(1)DAVID database was used to perform GO function and KEGG enrichment analysis on the predicted 503 miR-145-5p target gene sets.The target genes of hsa-miR-145-5p were enriched in aldosterone synthesis and secretion,PI3K-Akt signaling pathway,cancer transcriptional dysregulation,MAPK signaling pathway and other related signaling pathways.The DAVID database was used to perform GO function and KEGG enrichment analysis of the 13 predicted miR-126-3p target gene sets,and the miR-126-3p target genes were enriched in aldosterone regulated sodium reabsorption,cancer pathway and other related signaling pathways.(2)Compared with the healthy control group,the relative expression of miR-145-5p in the Yin deficiency and Yang hyperactivity group,the liver fire hyperactivity group,the phum-dampness excess group,and the Yin and Yang deficiency group was significantly different(P < 0.05).The relative expression of miR-145-5p in the hyperactive liver fire group,the phum-dampness group,and the yin-yang deficiency group was compared with the Yin deficiency and Yang hyperactivity group,respectively(P < 0.05),and the difference was statistically significant.Among them,the relative expression of miR-145-5p in the Yin deficiency and Yang excess group was significantly lower than that in the liver fire hyperactivity group,phum-dampness excess group,Yin and Yang deficiency group and the healthy control group.The expression level of miR-145-5p is down-regulated in hypertension with Yin deficiency and Yang excess syndrome.(3)The relative expression of miR-145-5p was negatively correlated with the plasma levels of ACE,AngⅡ and AGTR1(P < 0.05).Conclusions:(1)This study found a significant negative correlation between the relative expression level of miR-145-5p and the plasma levels of ACe-ANGi I-AT1 axis-related signaling factors ACE,AngⅡ and AGTR1.(2)This study found that the expression level of miR-145-5p was down-regulated in the syndrome of Yin deficiency and Yang hyperactivity of essential hypertension.(3)This study found that ACE-AngⅡ-AT1 axis targeting micro RNA: miR-145-5p may be the specific expression of miRNA in the formation of Yin deficiency and Yang hyperactivity syndrome of essential hypertension.