The Effect Of Irbesartan On Cardiac Inflammation And Fibrosis In Db/db Mice And Its Mechanism

ObjectiveTo investigate the protective effects of irbesartan on cardiac inflammation and fibrosis associated with diabetes and obesity in type 2 diabetic db/db mice and explore the underlying mechanisms.Methods and Contents1.Groups and treatments:36 ten-week-old diabetic db/db mice were equally randomized into model group,irbesartan treatment(50mg/kg per day)group and positive control group(a parthenolide(PTL)derivative treatment(25mg/kg per day)group),using 12 nondiabetic littermates(db/+)as the controls.The mice were treated with irbesartan,PTL derivative or saline vehicle through oral gavage for 16 consecutive weeks.2.General condition and biochemical indicators:their heart weight,body weight,serum levels of fasting blood glucose(FBG),Triglycerides(TG),total cholesterol(TC)were measured.3.The HE staining of left ventricle myocardial tissue:their heart morphology was observed.4.Masson trichrome staining:the content of myocardial collagen fibers was observed5.Western blotting and p65 assessed by immunohistochemistry:l)The protein levels of P-IκBα,IκBα and β-actin were analyzed by Western blotting.The activity and expression of nuclear factor-kappaB(NF-κB)p65 in the myocardium were assessed by immunohistochemistry.2)The protein levels of P-JAK2,JAK2,P-STAT3 and STAT3 were detected by Western blotting.6.Quantitative real-time PCR(qPCR):The mRNA expression of proinflammatory cytokines IL-6,TNF-a and profibrogenic cytokine TGF-β1 in myocardium were detected.Results1.General condition and biochemical indicators:Compared with db/+ mice,the saline,irbesartan or PTL derivative-treated db/db mice developed obesity,hyperglycemia and hyperlipidemia.2.The HE staining of left ventricle myocardial tissue:Compared with db/+ mice,histopathological examination of heart tissue of model group revealed disorders of myocardial fiber arrangement,increased myocardial interstitium and focal inflammatory cell infiltration.The situations were improved by irbesartan treatment,while effect of PTL derivative treatment was better.3.Masson trichrome staining:the collagen fibers around myocardial interstitial and blood vessels in the model group were significantly increased than those in the normal group,accompanied with uneven distribution,disordered arrangement and net-like connection.The situations were improved by treatment with irbesartan,while collagen fibers were significantly reduced by PTL derivative.4.Western blotting and p65 assessed by immunohistochemistry:l)Compared with db/+ mice,db/db mice showed increased P-IκBα and decreased IκBα protein levels,enhanced activity and expression of NF-κB in the hearts.2)The protein expression of P-JAK2 and P-STAT3 was significantly up-regulated.All of these changes were improved by the chronic treatment with irbesartan;and the improvement by PTL derivative treatment was more pronounced.However,the expression of JAK2 and STAT3 protein in the four groups were unchanged.5.Quantitative real-time PCR(qPCR):The mRNA expression of proinflammatory cytokines IL-6,TNF-α and profibrogenic cytokine TGF-β1 in the model group were higher than those in the normal group.These levels of mRNA expression were reduced by the chronic treatment with irbesartan,and significantly inhibited by chronic treatment with PTL derivative.Conclusions1.NF-κB and JAK2/STAT3 signaling pathways are activated,the expression of IL-6,TNF-α and TGF-β1 mRNA are increased and cardiac inflammation and fibrosis are promoted in type 2 diabetic db/db mice.2.The chronic treatment with irbesartan reduces the mRNA expression of IL-6 and TNF-α genes,attenuates cardiac inflammation in type 2 diabetic db/db mice,and these effects were probably associated with the suppression of cardiac angiotensin Ⅱand NF-κB signaling pathway..3.The chronic treatment with irbesartan down-regulates the JAK2/STAT3 signaling pathway,suppresses the expression of TGF-β1 mRNA and improves myocardial fibrosis in type 2 diabetic db/db mice,probably associated with inhibition of NF-κB signaling pathway.4.Early intervention with irbesartan may be a valuable adjuvant drug to the clinical treatment of diabetic cardiomyopathy,especially in diabetic patients with hypertension.