Objective:To investigate the regulatory effect of miR-146a overexpression on corneal inflammatory response in a mouse model of dry eye,and to analyze its relationship with the IRAK1/TRAF6/NF-кB signaling pathway.Meth...Objective:To investigate the regulatory effect of miR-146a overexpression on corneal inflammatory response in a mouse model of dry eye,and to analyze its relationship with the IRAK1/TRAF6/NF-кB signaling pathway.Methods:A total of 50 SPF-grade BALB/c mice were randomly divided into five groups,with10 mice in each group.Except for the control group,the other four groups were treated with 0.2%benzalkonium chloride(BAC)solution in both eyes to construct a dry eye model.After successful modeling,the control group and model group received NC agomir;the miR antagonist group received miR-146a antagomir;the miR agonist group received miR-146a agomir;and the pathway agonist group received miR-146a agomir+NF-κB activator 2.After four weeks of treatment,the expressions levels of miR-146a,inflammatory factors,and IRAK1/TRAF6/NF-κB signaling pathway proteins were observed and compared among the five groups.Results:After four weeks of treatment,there was a statistically significant difference in the relative expression of miR-146a in the five groups(F=61.058,P<0.001),which was significantly higher in the miR agonist group than in the other four groups.After4 weeks of treatment,there were statistically significant differences in the expression levels of IL-1β,IL-6,IL-8 and TNF-αin the five groups(F=84.757,103.658,55.477,46.762;P<0.001).After four weeks of treatment,there were statistically significant differences in the protein expression levels of IRAK1,TRAF6,NF-κB and IκBαin the five groups(F=62.975,77.173,67.108,29.381;P<0.001),except for the control group,the expression levels of IRAK1,TRAF6 and NF-κB proteins in the miR agonist group were significantly lower than those in the other three groups,and the expression levels of IκBαprotein were significantly higher than those in the other three groups.Conclusion:Overexpression of miR-146a can negatively regulate corneal inflammatory response in dry eye mice through the IRAK1/TRAF6/NF-κB signaling pathway,which can provide new insights for the clinical treatment of dry eye disease.展开更多
Interleukin I receptor associated kinase 1 (IRAK1) is a downstream signal molecule of activated MyD88 recruitment, which can activate Fas associated death domain protein (FADD) to induce apoptosis. IRAK1 can also acti...Interleukin I receptor associated kinase 1 (IRAK1) is a downstream signal molecule of activated MyD88 recruitment, which can activate Fas associated death domain protein (FADD) to induce apoptosis. IRAK1 can also activate tumor necrosis factor-related factor 6 (TRAF6) and induce the expression of a series of downstream specific genes. IRAK1 is an essential factor in the induction of mitochondrial division and necroptosis. In the current study, RNAi technique was used to silence IRAK1, and the apoptosis and necroptosis rate of SK-Hep1 cells were detected by flow cytometry. The apoptosis and the necroptosis pathway of hepatoma SK-Hep1 cells were blocked separately, and the expressions of FADD, RIP1 and TRAF6 genes were silenced separately. The results showed when the expression of IRAK1 was down-regulated, the apoptosis and necroptosis rate of SK-Hep1 cells were significantly increased. With silenced FADD, RIP1 and TRAF6, respectively, the expression of IRAK1 protein had no significant change. However, the expression of IRAK1 mRNA decreased significantly (p < 0.01) after the silencing of RIP1 and TRAF6 genes, while the IRAK1 mRNA did not change significantly after the silencing of FADD genes;when z-VAD-FMK was interfered, the expression of IRAK1 mRNA decreased significantly after the silencing of TRAF6 genes, while the IRAK1 mRNA did not change significantly after the silencing of FADD and RIP1genes. The study shows that RAK1 gene inhibits apoptosis and necroptosis in SK-Hep1 cells. TRAF6 gene affected the role of IRAK1 in apoptosis and necroptosis, RIP1 gene affected the role of IRAK1 in apoptosis, while FADD gene did not affect the role of IRAK1 in apoptosis and necroptosis.展开更多
Background Coronary microembolization （CME） is characterized by distal microvascular occlusion. However, the inflammatory mechanisms and therapeutic targets of CME are largely unknown. Methods A total of 11 Guangxi ...Background Coronary microembolization （CME） is characterized by distal microvascular occlusion. However, the inflammatory mechanisms and therapeutic targets of CME are largely unknown. Methods A total of 11 Guangxi Bama miniature swines were divided into two groups： sham （n = 5） and CME （n = 6）. Microspheres were injected into the left anterior descending artery of the CME group to make an animal model of CME. The expres- sions of microRNA-146a （miR-146a） and IRAK1, TRAF6, and AUF1 in the myocardium were detected by qPCR. Results In the CME group, microspheres, microinfarction, and inflammatory cell infiltration were found under an optical microscope. The expression levels of miR-146a were low in both groups. After CME, the expression levels of IRAK1, TRAF6, and AUF1 in the CME group were upregulated compared with those in the sham group （P 〈 0.01;P 〈 0.05;P 〈 0.05, respectively）. Conclusions AUF1, IRAK1 and TRAF6, but not miR-146a, could be involved, in myocardium inflammation following CME.展开更多
文摘Objective:To investigate the regulatory effect of miR-146a overexpression on corneal inflammatory response in a mouse model of dry eye,and to analyze its relationship with the IRAK1/TRAF6/NF-кB signaling pathway.Methods:A total of 50 SPF-grade BALB/c mice were randomly divided into five groups,with10 mice in each group.Except for the control group,the other four groups were treated with 0.2%benzalkonium chloride(BAC)solution in both eyes to construct a dry eye model.After successful modeling,the control group and model group received NC agomir;the miR antagonist group received miR-146a antagomir;the miR agonist group received miR-146a agomir;and the pathway agonist group received miR-146a agomir+NF-κB activator 2.After four weeks of treatment,the expressions levels of miR-146a,inflammatory factors,and IRAK1/TRAF6/NF-κB signaling pathway proteins were observed and compared among the five groups.Results:After four weeks of treatment,there was a statistically significant difference in the relative expression of miR-146a in the five groups(F=61.058,P<0.001),which was significantly higher in the miR agonist group than in the other four groups.After4 weeks of treatment,there were statistically significant differences in the expression levels of IL-1β,IL-6,IL-8 and TNF-αin the five groups(F=84.757,103.658,55.477,46.762;P<0.001).After four weeks of treatment,there were statistically significant differences in the protein expression levels of IRAK1,TRAF6,NF-κB and IκBαin the five groups(F=62.975,77.173,67.108,29.381;P<0.001),except for the control group,the expression levels of IRAK1,TRAF6 and NF-κB proteins in the miR agonist group were significantly lower than those in the other three groups,and the expression levels of IκBαprotein were significantly higher than those in the other three groups.Conclusion:Overexpression of miR-146a can negatively regulate corneal inflammatory response in dry eye mice through the IRAK1/TRAF6/NF-κB signaling pathway,which can provide new insights for the clinical treatment of dry eye disease.
文摘目的·扩增干扰素调节因子3(interferon regulator factor 3,IRF3)短发夹RNA(short hairpin RNA,sh RNA)腺病毒,并研究该病毒对脂多糖(lipopolysaccharide,LPS)刺激诱导Raw264.7细胞核内白介素受体相关激酶1结合蛋白1(interleukin-1 receptor associated kinase 1 binding protein 1,Irak1bp1)表达的影响。方法·IRF3 sh RNA腺病毒的扩增在人胚肾293 T(HEK293T)细胞中进行,并采用TCID 50法测定病毒滴度。Raw 264.7细胞随机分为4组,1组为腺病毒(-)LPS(-),2组为腺病毒(-)LPS(+),3组为腺病毒(+)LPS(-),4组为腺病毒(+)LPS(+)。细胞IRF3基因表达采用real-time PCR方法检测;核内IRF3及Irak1bp1的表达采用Western blotting方法检测。结果·经计算扩增腺病毒滴度为2.2×10^(11) PFU/m L,最佳MOI为300。LPS刺激后Raw 264.7细胞内IRF3 m RNA较对照组明显增加,核内IRF3蛋白及Irak1bp1表达也明显增加;IRF3 sh RNA腺病毒应用后,细胞对IRF3 m RNA的组成性表达及LPS刺激诱导的IRF3 m RNA和核内蛋白质表达均明显受抑,但未刺激状态下IRF3蛋白核内组成性表达无明显影响;IRF3 sh RNA腺病毒应用对细胞静息及LPS刺激诱导的核内Irak1bp1表达并无影响。结论·IRF3 sh RNA腺病毒能够有效抑制LPS刺激诱导的核内IRF3的表达,但并不影响核内Irak1bp1的表达。
文摘Interleukin I receptor associated kinase 1 (IRAK1) is a downstream signal molecule of activated MyD88 recruitment, which can activate Fas associated death domain protein (FADD) to induce apoptosis. IRAK1 can also activate tumor necrosis factor-related factor 6 (TRAF6) and induce the expression of a series of downstream specific genes. IRAK1 is an essential factor in the induction of mitochondrial division and necroptosis. In the current study, RNAi technique was used to silence IRAK1, and the apoptosis and necroptosis rate of SK-Hep1 cells were detected by flow cytometry. The apoptosis and the necroptosis pathway of hepatoma SK-Hep1 cells were blocked separately, and the expressions of FADD, RIP1 and TRAF6 genes were silenced separately. The results showed when the expression of IRAK1 was down-regulated, the apoptosis and necroptosis rate of SK-Hep1 cells were significantly increased. With silenced FADD, RIP1 and TRAF6, respectively, the expression of IRAK1 protein had no significant change. However, the expression of IRAK1 mRNA decreased significantly (p < 0.01) after the silencing of RIP1 and TRAF6 genes, while the IRAK1 mRNA did not change significantly after the silencing of FADD genes;when z-VAD-FMK was interfered, the expression of IRAK1 mRNA decreased significantly after the silencing of TRAF6 genes, while the IRAK1 mRNA did not change significantly after the silencing of FADD and RIP1genes. The study shows that RAK1 gene inhibits apoptosis and necroptosis in SK-Hep1 cells. TRAF6 gene affected the role of IRAK1 in apoptosis and necroptosis, RIP1 gene affected the role of IRAK1 in apoptosis, while FADD gene did not affect the role of IRAK1 in apoptosis and necroptosis.
基金supported by Guangxi Province Medical Technology Research and Development Project(Grant No.S201303_01)Youth Science Foundation of Guangxi Medical University(No.GXMUYSF201213)
文摘Background Coronary microembolization （CME） is characterized by distal microvascular occlusion. However, the inflammatory mechanisms and therapeutic targets of CME are largely unknown. Methods A total of 11 Guangxi Bama miniature swines were divided into two groups： sham （n = 5） and CME （n = 6）. Microspheres were injected into the left anterior descending artery of the CME group to make an animal model of CME. The expres- sions of microRNA-146a （miR-146a） and IRAK1, TRAF6, and AUF1 in the myocardium were detected by qPCR. Results In the CME group, microspheres, microinfarction, and inflammatory cell infiltration were found under an optical microscope. The expression levels of miR-146a were low in both groups. After CME, the expression levels of IRAK1, TRAF6, and AUF1 in the CME group were upregulated compared with those in the sham group （P 〈 0.01;P 〈 0.05;P 〈 0.05, respectively）. Conclusions AUF1, IRAK1 and TRAF6, but not miR-146a, could be involved, in myocardium inflammation following CME.
