Experimental Study Of Function Of WISP1Protein And Effects Of Puerarin In Viral Myocarditis

Background: Viral myocarditis (VMC) is a common diseases of the cardiovascularsystem in pediatric. The disease’s clinical manifestations is diverse from mildly toseverely, and no special treatment. Its prognosis is significantly different.Pathogenesis of VMC has not completely cleared yet and the myocardial fibrosis (MF)which lead to is closely related to its prognosis.WISP1is Wnt-inducible protein1.Some studies found that WISP1exerts prohypertrophic and mitogenic effectsstimulating myocyte hypertrophy, cardiac fibroblast (CF) proliferation, and collagenexpression.It is closely related with MF and may be induced by TNF-α in theTNF-α/WISP1pathway. Puerarin is a isoflavonoids extract from pueraria. It canprotect ischemic myocardium and improve myocardial metabolism and also has thefunction of oxidation resistance, antiapoptotic and anti-myocardial fibrosis, and so on.Objective: To establish the models of VMC through Balb/c mice infected withcoxsackie virus B3, we detect the expression of WISP1protein in myocardial cellsand study the role of WISP1in the process of the occurrence and development ofVMC and be clear of the intervention effect of Puerarin on VMC.Methods: The experiment selected120male Balb/c mice,weighting14~15g,dividedinto three groups randomly：control group(n=30),VMC group(n=45)，treatmentgroup(n=45). The VMC group and the treatment group establish VMC model byintraperitoneal injection CVB30.2ml for3days.The treatment group were injectedpuerarin for28days from the date of the virus inoculation.We recorded the status anddeaths of the mice everyday.We weighted the mice on the4th,7th,14th,28th day,killed6mice randomly everytime,detected the value of CK-MB in blood serum,examinedthe score for myocardial necrosis and cellular infiltration by HE staining, observedmyocardial collagen fiber distribution and measured collagen volume fraction (CVF)in mice, tested WISP1and TNF-α protein by immunohistochemical method. Finallywe use software SSPS17.0to analyse the results. Results:(1) VMC mice model was successfully build after intraperitioneal injectionof CVB33days.Compared with the control group and treatment group,mice of theVMC group is as follows: bad general state and poor quality of life, light weight andhigh mortality; high peak value of CK-MB and declined slowly; HE staining show thesevere pathological changes and high scores of myocardial necrosis and cellularinfiltration.There’s significant differences compared with the treatment group;Massonstaining show that compared with the treatment group, myocardial fibers increased onthe7th day, collgen area increased significantly and the staining deepening,CVFincreased,too. Immunohisochemical method show that the expression of WISP1andTNF-α increased significantly.The above-mentioned show that puerarin injection hastherapeutic effect on VMC which maybe in protecting the myocardium and improvinginflammation and anti-fibrotic sides and maybe related to the mechanism of action ofWISP1.(2) Immunohisochemical method also show the negative mice has a littleexpression of WISP1and TNF-α, meanwhile more expression on VMC mice,suggestthat WISP1and TNF-α indeed involved in the pathogenesis of VMC. The mice of theVMC group begin to express WISP1and TNF-α on the4th day and more on thefollowing days.WISP1was expressed up to the peak level on the28th day, whileTNF-α peaked on the7th day and decreased slightly on the14th day, reached thelowest value on the28th day.WISP1and TNF-α express up on the place of theinflammation around the necrotic lesions and fibrosis,and WISP1express earlier thancollagen fibers increased, suggest that inflammatory cytokines TNF-α maybestimulates fibroblast and then mediated the abnormal expression of WISP1.(3) Linearrelevance analysis shows that:WISP1is correlated positively with CVF remarkablyand no relevance with CK-MB or the scores of myocardial necrosis and cellularinfiltration,which suggest that WISP1is closely related to myocardial fibrosis andthus involved in the progress and development of VMC. TNF-α is correlatedpositively with CK-MB and the scores of myocardial necrosis and cellular infiltrationremarkably but no relevance with CVF,which suggest that TNF-α is aproinflammatory cytokines but not a sign of factor of myocardial fibrosis on VMC.The linear relevance analysis also shows that,WISP1has no relevance with TNF-α,which inconsistent with the speculated,supposed that because of multiple rolesof TNF-α—sensitive inflammtory cytokines and be correlate with fibrosis and alsoinvolve in immune response,but we can’t deny that it maybe mediated the expressionof WISP1in myocardial fibrosis on VMC, and we need more study to confirm.Conclusion:1. WISP1participate in the occurrence and development of VMC.2.WISP1is closely related to the degree of myocardial fibrosis,and the process maybemediated by the upregulation of TNF-α.3. Puerarin can improve the livability ofVMC mice,protect myocardium,improve myocardial inflammation,Which may beassociated with its function of apoptosis resistance,oxidation resistance andscavenging oxygen free radicals.4. Puerarin can reduce the VMC myocardialfibrosis,Which may be achieved by inhibiting the expression of TNF-α anddown-regulated the expression of WISP1.