| Objective: To investigate the effect of human α1-antitrypsin(hAAT)on myocardial ischemia-reperfusion injury(MIRI)in mice and its mechanism.Methods : SPF grade healthy male C57BL/6 wild-type mice and hAAT transgenic overexpression mice were randomly divided into four groups: Wild type sham operation group(Sham group),wild type ischemia-reperfusion group(I/R group),hAAT-Tg sham operation group(hAAT group),hAAT-Tg ischemia-reperfusion group(hAAT-Tg+I/R group),sham operation group(n=18),operation group(n=18).The model of MIRI in mice was prepared by ligating the left anterior descending coronary artery for 60 min and unting the ligation line.The sham operation group only opened the chest without ligation.After 24 hours of reperfusion,five mice in each group were randomly selected,and the myocardial infarction area of each group was evaluated by Evans blue and 2,3,5-triphenyltetrazolium chloride(TTC)double staining.After 7 days of reperfusion,the indexes of cardiac function of mice in each group were measured by small animal ultrasound system,the pathological changes of myocardial tissue were observed by HE staining,and the degree of myocardial fibrosis was observed by Masson staining.The concentration changes of serum markers cardiac troponin I(TNI),creatine kinase isoenzyme(CK-MB)and interleukin-1β(IL-1β)were detected by enzyme-linked immunosorbent assay(ELISA).Real-time fluorescence quantitative PCR was used to detect the level of pro-inflammatory factor TNFɑ,IL-6 and IL-1β m RNA in myocardial tissue of each group.Nucleotide Oligomerization Domain Like Receptor Protein 3(NLRP3),Cysteine proteinase-1(caspase-1)and IL-1β in myocardial tissue of each group were detected by Western blot and immunohistochemical staining Protein expression.Result:(1)Compared with sham group,the myocardial infarction area in I/R group and hAAT-Tg+I/R group increased significantly(P<0.01),and compared with I/R group,the myocardial infarction area in hAAT+I/R group decreased significantly [(23.4 ± 2.36)% vs.(39.4 ± 2.06)%P<0.05].(2)The left ventricular end diastolic diameter(LVEDD)and left ventricular end systolic diameter(LVESD)in the hAAT-Tg+I/R group were significantly lower than those in the I/R Group [3.58 ± 0.38 mm vs.(4.12 ± 0.61)mm and(4.17 ± 0.20)mm vs.(4.54 ±0.55)mm,respectively,P <0.05],and the left ventricular ejection fraction(LVEF)and left ventricular short axis shortening(LVFS)were significantly higher than those in the i/r Group [(32.75 ± 10.72)% vs.(20.92 ± 8.50)% and(15.53 ± 8.50)% respectively.5.56)% vs.(9.56 ± 4.07)% P <0.05].(3)HE staining results showed that compared with sham group,inflammatory cell infiltration was obvious and myocardial fibers were disordered in I/R group.Compared with I/R group,inflammatory cell infiltration was reduced and myocardial fibers were relatively orderly in hAAT-Tg+I/R group.Masson staining results showed that compared with sham group,there was a large amount of collagen deposition in the infarct area of I/R group,and the degree of fibrosis was significantly increased.Compared with I/R group,the collagen deposition was significantly reduced and the degree of fibrosis was mild in hAAT-Tg+I/R group.(4)ELISA results showed that compared with sham group,the serum levels of Tn I,CK-MB and IL-1β in I/R group were significantly higher,and the levels of Tn I,CK-MB and IL-1β in hAAT-Tg+I/R group were significantly lower than those in I/R group(P<0.05).The results of fluorescence quantitative PCR showed that the level of TNFɑ,IL-6 and IL-1β m RNA in myocardial tissue of I/R group was significantly higher than that of Sham group(P<0.05),and the level of TNFɑ,IL-6 and IL-1β m RNA in hAAT-Tg+I/R group was significantly lower than that of I/R group(P<0.05).(5)Compared with sham group,the expression of NLRP3,caspase-1 and IL-1β in myocardial tissue of I/R group was significantly up-regulated,while the expression levels of NLRP3,caspase-1 and IL-1β in hAAT-Tg+I/R group were significantly lower than those in I/R group(P<0.01).Conclusion: HAAT can alleviate MIRI in mice,and its mechanism may be related to the inhibition of inflammatory response by inhibiting the activation of NLRP3/caspase-1/IL-1β signaling pathway. |
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