The Cardioprotection Of RP105 In Myocardial Ischemia Reperfusion Injury By Regulating TLR4/TLR2 Signal Pathways

Objective The major aim of our present work was to study the cardioprotection of RP105 on myocardial ischemia reperfusion injury(MIRI)and expected to explore its molecular mechanism.Methods Seventy-fivemale Sprague-Dawley(SD)rats in specific pathogen free(SPF)grade(weight 220-250g)are randomly divided into 5 equal groups(n=15).(1)saline infection with sham operation(Sham group).(2)myocardial I/R with saline infection(I/R group)(3)myocardial IR with infection Ad-EGFP-RP105(Ad-R group).(4)myocardial I/R with infection Ad-EGFP-RP105-si RNA(Ad-R-si RNA group).(5)myocardial I/R with infection Ad-EGFP-si RNA(Ad-G-si RNA group).RP105 recombinant adenovirus vector(Ad-EGFP-RP105,Ad-EGFP-RP105-si RNA),virus with(Ad-EGFP)or saline are infected into the apex of heartrespectively.MIRI rats are established by ligatured the left anterior descending coronary artery for 30 min.The blood is pouced into artry 3 hours later.The expression of RP105 in cardiac tissue is observed by immunofluorescence.The levels ofmyocardial enzyme LDH,CK and CK-MB in serum were measured by blood biochemical analysis.Myocardial infarct area is measured by thetriphenyltetrazolium chloride(TTC)staining.HE is performed to observe rat myocardial tissue morphology.Western blot analysis were used to examine the protein and the gene level of TRL4/TLR2 signaling pathway respectively and the expression of interleukin-6(IL-6)and Tumor Necrosis Factor-α(TNF-α).Results(1)RP105 is delivered into rat cardiac tissue successfully.(2)I/R group,Ad-G-si RNA group LDH,CK and CK-MB were performed a significant increase in serum comparingwith the sham group and Ad-R group(P<0.05).After RP105 transduction,LDH,CK and CK-MB decreased obviouslyin Ad-R group(P<0.05).(3)Comparing with the Sham group,the myocardial infarction area of IR group increased apparently(P<0.05).The myocardium transduced with Ad-R exerted cardioprotective effect and the infarct areawas less than I/R group and Ad-G-si RNA(P<0.05).(4)HE staining was used to observe the rat myocardiummorphology.The cardial cells degenerated and motified apparently in I/R,Ad-R,Ad-G-si RNA and Ad-R-si RNA groups.And this phenomenon was more obvious in Ad-R-si RNA group than other groups.(5)We use the expression of TLR2/4proteins were detected by Western Blot.The expression of TLR2/4 increased by down regulated RP105 in Ad-R-si RNA,while the TLR2/4 expression decreased by up regulated RP105 in Ad-R.(6)the Western Blot was used todetecte the expression of inflammatory factors IL-6 and TNF-α.It is found that compared with I/R group and Ad-G-si RNA group,IL-6 and TNF-αincreased obviously after RP105 silence(P<0.05).(7)The expression of My D88 and NF-κB were detected by Western Blot.It is found that after RP105 silence,the expression of My D88 and NF-κB increased apparently compared with I/R and Ad-G-si RNA groups(P<0.05).(8)Detected Capcase-3 and Bcl-2 by Western Blot,we find that Capcase-3 expression level in Ad-R-si RNA group increased obviously(P<0.05),while the expression level of Bcl-2 decreased apparently(P<0.05).Conclusion: The silence of RP105 incardiocytes could increasethe expression of inflammatory factors,myocardial infarct size and serum myocardial enzyme viaregulating TLR4 and TLR2 signaling pathways.What we find may be a novel therapeutic approach for MIRI prevention.