Expression of RNA Binding Motif Protein 15 in Myocardium of Septic Mice and Its Correlation with Neutrophil Extracellular Trapping Network
Objective To investigate the expression of RNA binding motif protein 15(RBM15)in septic myocardial injury mice and its correlation with neutrophil extracellular traps(NETs).Methods Lipopolysaccharides(LPS)were injected intraperitoneally to establish sepsis model in mice,and saline-treated mice were used as control group.Cardiac function was measured by Doppler echocardiography in mice 7 days after LPS injection.HE staining was used to observe myocardial injury in septic mice.Immunofluorescence staining was used to detect CitH3 expression levels in myocardial tissue sections.Enzymatic activities of citrullinated histone H3(CitH3)in myocardial tissue,myeloperoxidase(MPO)in plasma and neutrophil elastase(NE)in myocardial tissue were detected by ELISA.RT-qPCR was used to detect neutrophil-associated molecules inhibitor kappa B alpha(IκBα),nuclear factor-kappa B(NF-κB),B-cell lymphoma factor 10(Bcl10),peptidylarginine deiminase 4(PAD4)and RBM15,N6-methyladenine(m6A)methyltransferase methyltransferase-like protein 3(METTL3),methyltransferase-like 14(METTL14),wilms'1-binding protein(WTAP),KIAA1429.Western blot was used to detect neutrophil-related molecules IκBα,nuclear factor-kappa B/p65(NF-κB/p65),nuclear factor-kappa B/p50(NF-κB/p50),Bcl10,PAD4 and RBM15,m6A methyltransferase METTL3,METTL14,WTAP,KIAA1429.Results The results of cardiac function measured by cardiac Doppler ultrasound showed that LVEF and FS were decreased in the experimental group compared with the control group(P all＜0.001).The results of HE staining showed that the myocardial tissue structure of mice in the control group was normal,and the myocardium of mice in the experimental group was partially broken,showing inflammatory cell infiltration and cell edema.Immunofluorescence staining results showed that CitH3 fluorescence intensity was enhanced and expression was up-regulated in the experimental group(P＜0.05).ELISA results showed that NETs-related molecules CitH3,NE in myocardial tissue and MPO in plasma were increased in septic mice compared with controls(P all＜0.01).RT-qPCR results showed that there were no significant difference in Bcl10(P=0.960),PAD4(P=0.324),METTL14(P=0.592),and WTAP(P=0.505)mRNA expression levels in the experimental group compared with the control group.Increased IκBα(P＜0.01),NF-κB(p65/p50)(P＜0.05),and RBM15(P＜0.05)mRNA expression levels,while the METTL3(P＜0.05)and KIAA1429(P＜0.05)mRNA expression levels were decreased.Western blot results showed that there were no significant differences in IκBα(P=0.171),NF-κB/p65(P=0.191),NF-κB/p50(P=0.063),PAD4(P=0.687),RBM15(P=0.176),METTL3(P=0.430),METTL14(P=0.089),and KIAA1429(P=0.238)protein expression levels compared with controlgroup.WTAP protein expression level increased(P＜0.05)and Bcl10 protein expression level decreased(P＜0.05).RBM15 mRNA expression level was positively correlated with CitH3(r=0.631,P=0.011)expression level,but not with MPO(r=0.318,P=0.114)and NE(r=0.099,P=0.410)levels.Conclusion RBM15 and NETs were up-regulated in septic myocardial injury,and RBM15 was positively correlated with NETs-related molecule CitH3,and RBM15 may be involved in septic myocardial injury by regulating NETs-related molecules.
myocardial injury in sepsisneutrophil extracellular trapsRNA binding motif protein 15
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