Effect Of Icariside Ⅱ On Myocardial Fibrosis In Spontaneously Hypertensive Rats

Objective: In the current study,we aimed to evaluate the effects of Icariside Ⅱ(ICS Ⅱ)on induced myocardial fibrosis in spontaneously hypertensive rats(SHR)and explore the mechanism underlying this activity.Methods: 28 male SHR rats of 13 weeks of age were randomly divided into the model group(group SHR,n = 7),ICS Ⅱ low(4 mg/kg,ICS Ⅱ-L,n = 7),middle(8 mg/kg,ICS Ⅱ-M,n = 7)and high(16 mg/kg,ICS Ⅱ-H,n = 7)group,and 7 male WKY rats of the same age as control group(WKY group).The SHRs were treated with ICS Ⅱ(4,8,and 16 mg/kg)via daily gavage for 12 weeks,and WKY and SHR groups were given the equal volume of double-distilled water.After 12 weeks of administration,the blood pressure was measured by Scientific CODA Kent system;left ventricular function was detected using the Vevo2100 system,Then,the rats were sacrificed and the left ventricles were separated in order to calculate the left ventricular mass index.Left ventricular morphology was observed by HE staining.Masson staining was used to detect the occurrence of interstitial fibrosis in the cardiac tissues.Serum samples were collected for determination of IL-6 and TNF-α by ELISA.Real time PCR was used to observe the gene expression of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,Smad2,Smad3,NF-κB,IκB-α,IL-1β and TNF-α in the left ventricle in SHRs.The protein expression levels of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,p-Smad2,p-Smad3,p-NF-κBp65,IκB-α,IL-1β and TNF-α were measured by Western Blot.Results: Compared with the WKY group,the amount of collagen fibre was increased in SHRs myocardium.The blood pressure,left ventricular mass index,LVID: s,LVAD: d were significantly increased(P<0.05).the EF,FS were significantly decreased(P<0.05).The left ventricular wall thickness was increased,the chamber was markedly narrower.HE staining showed myocardial cells disorder,hypertrophy in SHR group.Masson’s trichrome staining myocardial interstitium increased to varying degrees,myocardial fibrosis can be seen in interstitial cells,accompanied by severe collagen deposition.Serum levels of IL-6and TNF-α were up-regulated(P<0.05).The expressions of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,Smad2,Smad3,NF-κB,IL-1β and TNF-αm RNA were increased in the left ventricular tissue(P<0.05),and the protein expressions of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,p-Smad2,p-Smad3,p-NF-κBp65,IL-1β and TNF-α were also increased(P<0.05),while the m RNA and protein expression of TIMP-1and IκB-α was significantly reduced(P<0.05).Compared with the SHR group,the myocardial fibrosis was reduced after ICS Ⅱ(8,16 mg/kg)treatment.The blood pressure,left ventricular mass index,LVID: s,LVAD: d were significantly decreased(P<0.05),the EF,FS were significantly increased(P<0.05).Serum levels of IL-6and TNF-α were down-regulated(P<0.05).The expressions of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,Smad2,Smad3,NF-κB,IL-1β and TNF-αm RNA were down-regulated in the left ventricular tissue(P<0.05),and the protein expressions of Collagen I,Collagen ⅡI,α-SMA,MMP-2,MMP-9,TIMP-1,TGF-β1,p-Smad2,p-Smad3,p-NF-κBp65,IL-1β and TNF-α were also down-regulated(P<0.05),while the m RNA and protein expression of TIMP-1and IκB-α was significantly up-regulated(P<0.05).Conclusion: ICS Ⅱ can significantly attenuate the myocardial fibrosis in SHR,which may be related to the reduction of SHR blood pressure,improve left ventricular cardiac function and reduce collagen deposition,the mechanism may involve:(1)ICS Ⅱ may regulate collagen synthesis and deposition by inhibiting the expression of α-SMA,type I and type ⅡI collagen,decreasing the ratio of type I/ⅡI collagen;(2)ICS Ⅱ may regulate the synthesis and degradation of ECM by down-regulation of MMP-2,MMP-9 expression and up-regulation of TIMP-1 expression and decreasing the ratio of MMP-9/TIMP-1;(3)ICS Ⅱ may suppresses myocardial fibrosis by inhibiting TGF-β/Smads pathway;(4)ICS Ⅱ may alleviate by myocardial fibrosis by inhibiting the NF-κB inflammatory pathway.