Characteristic And Inflammation,fibrosis Change Of Different Adipose Tissue Depots In Patients With Primary Aldosteronism

BackgroundTodate,accumulating evidence has show that the incidence of primary aldosteronism is much higher than before.Compared with patients with essential hypertension matched for blood pressure,gender and age,primary aldosteronism(PA)patients has an increased rate of cardiovascular events and mortality.But,the mechanism underlying this phenomenon is still not clear.The underlying mechanisms involved may include the direct effect of aldosterone in cardiovascular and the indirect effect of aldosterone in adipose tissue.Whether Chronic aldosterone increase in patients with PA will direct affect adipose tissue inflammation and fibrosis which induce PA patients metabolize dysfunction is still not clarified yet.Research aim at PA patients different spot adipose tissue inflammation,fibrosis 1s very limit.ObjectiveTo investigate the characteristic of different spot adipose tissue in patients with aldosterone-producing adenoma and non-adrenal-disease patients,which include inflammation,fibrosis and lipid metabolism,adipokines.And assess whether the direet effect of aldosterone involved in the underlying mechanism of increase rate of cardiovascular.Method1.To analyse the characteristic and inflammation,fibrosis change of different adipose tissue depots in patients with primary aldosteronism.Adipose tissue was obtained from patients with APA and non-adrenal-disease patients(NAD).NAD include two group:normotension patients(NT)and essential hypertension(EH).Total RNA was isolated from APA patients's adipose tissue(surround adrenal(n=14),perirenal adipose tissue(n=6),extraperitoneal adipose tissue(n=8)and subcutaneous adiposue tissue(n=9)),NT patients adipose tissue(perirenal(n=7)and subcutaneous(n=6))and EH patients's adipose tissue(perirenal(n=5)and subcutaneous(n=5)).Real-time quantitative PCR for IL-6,TNF-?,EGR1,TIMP1,MMP2,FN,COL,TGFpl,ATGL,COX2 and adipokines include PPAR?,C/EBP?,C/EBP?,UCP1,PRDM16,PGCla,CIDEA and 18S mRNA was performed.Immunohistochemical staining was performed to assess the perirenal adipose tissuse protein expression of TNF-?.Masson's trichrome was used to assess the degree of perirenal adipose tissue fibrosis.Western Blot was used to measure IL-6 protein in perirenal adipose tissue.2.To analyse the impact of aldosterone on SVF cellsSVF cells were obtained from non-adrenal-disease patient's perirenal adipose tissue.SVF cells were cultured and differentiated into adipocytes.Differentiated SVF cells were cultured in FBS free DMEM for 12 hours and then stimulated with 10nMol/L aldosterone for 24 hours.Total RNA was extracted from SVF cells and Real-time quantitative PCR for the above-mentioned detecting index mRNA was performed.3.To analyse the effect of aldosterone on culture adipocyte3T3-L1 preadipocyte and mouse brown preadipocytes(WT-1)were differentiated with standard differentiation protocol,in the absence or the presence of 10nMol/L aldosterone.Total RNA was extracted from 3T3-LI,WT-1 cells and Real-time quantitative PCR for the above-mentioned detecting index mRNA was performed.Results1.The expression of inflammation,fibrosis factors and adipokine in adipose tissues of patients with APA1.1 The expression of inflammation and fibrosis factors in adipose tissues of patients with APA.IL-6,TNF-? and EGR1,FN,COL,TGF?1 mRNA levels from prirenal adipose tissue were significantly higher in APA patients than in NT,EH patients(p<0.05).MMP2 and ATGL mRNA levels were significantly higher in APA patients's prirenal adipose tissue than in EH patients(p<0.05).The immunohistochemical staining results indicate that the expression of TNF-? in perirenal adipose tissue is significantly higher in APA patients than NT and EH patients(p<0.05).Masson's trichrome staining showed that perirenal adipose tissue from APA patients developed more fibrosis compared with NT and EH patients(p<0.05).Western Blot analyses show that IL-6 protein expression is significantly higher in perirenal adipose tissue in patients with APA than EH patients(p<0.05).1.2 The expression of adipokine gene mRNA in adipose tissues of patients with APA.In patients with APA,PPAR?,C/EBP? and C/EBP?,UCP1,PGCl?,CIDEA mRNA were significantly higher in perirenal adipose tissue than NT,EH patients(p<0.05).In subcutaneous adipose tissue,C/EBP?,PGC1? and CIDEA mRNA were higher in APA patients than in NT,EH patients(p<0.05).2.Effects of aldosterone on adipokines,lipid metabolism and inflammation,fibrosis gene mRNA in human perirenal adipose tissue stromal vascular fraction(SVF)cells.10nMol/L aldosterone significantly induced differentiated SVF cells IL-6,EGR1,MMP2,FN,COL,TGF?1 and ATGL mRNA expression(p<0.05).And also induced a marked up-regulation of PPAR?,C/EBP? and CIDEA mRNA levels.3.Effects of aldosterone on culture adipocyte3.1 Effects of 10nMol/L aldosterone on 3T3-L1 preadipocyte.3.1.1 Effects of 10nMol/L aldosterone on 3T3-L1 preadipocyte differentiation.During differentiation,10nMol/L aldosterone significantly increase 3T3-L1 cells PPAR?,C/EBP? and C/EBP? mRNA level(p<0.05).Aldosterone also induce ATGL and IL-6,TNF-?,EGR1,FN,COL,TGF?1mRNA expression significantly mcrease(p<0.05).3.1.2 Effects of 10nMol/L aldosterone on Differentiated 3T3-L1 cells.lOnMol/L aldosterone significantly induced differentiated 3T3-L1 cells PPAR?,C/EBP?,UCP1 and PGC1? mRNA expression(p<0.05).And also induced a marked up-regulation of ATGL and IL-6,TNF-?,EGR1,FN,COL,TGFpl mRNA levels(p<0.05).3.2 Effects of 10nMol/L aldosterone on WT-1 preadipocyte.3.2.1 Effects of 10nMol/L aldosterone on WT-1 preadipocyte differentiation.During differentiation,10nMol/L aldosterone significantly increase 3T3-L1 cells PPAR?,C/EBP?,UCP1,PGC1? and IL-6,TNF-?,FN,COL,TGF?1 mRNA level(p<0.05).3.2.2 Effects of 10nMol/L aldosterone on Differentiated WT-1 cells.10nMol/L aldosterone significantly induced differentiated WT-1 cells PPAR?,C/EBP?,UCP1 and PGCla mRNA expression(P<0.05).And also induced a marked up-regulation of ATGL and IL-6,TNF-?,EGR1,FN,COL,TGFpl mRNA levels(p<0.05).ConclusionCompared with NT and EH patients,gene expression of inflammation and fibrosis in perirenal and subcutaneous adipose tissue of patients with APA were significantly up-regulated.And adipokines mRNA in perirenal and subcutaneous adipose tissues in patients with APA also indicate APA induce change in adipose tissue.Effects of Aldosterone on perirenal adipose tissue SVF cells further comfirm the result that aldosterone induce differented SVF cells inflammation and fibrosis gene mRNA expression up-regulated,and influence adipokines mRNA levels.Aldosterone also can induce 3T3-L1 and WT-1 preadipocyte differentiation,and increase differented 3T3-L1 and WT-1 cells inflammation,fibrosis gene mRNA.The results indicate that aldosterone may induced adipose tissue dysfunction and lead to inflammation and fibrosis,which may involved in primary aldosteronism patients high rate of cardiovascular events and mortality.