AIM:To investigate lipopolysaccharide(LPS) related signal transduction in interstitial cells of Cajal(ICCs) from mouse small intestine.METHODS:For this study,primary culture of ICCs was prepared from the small intesti...AIM:To investigate lipopolysaccharide(LPS) related signal transduction in interstitial cells of Cajal(ICCs) from mouse small intestine.METHODS:For this study,primary culture of ICCs was prepared from the small intestine of the mouse.LPS was treated to the cells prior to measurement of the membrane currents by using whole-cell patch clamp technique.Immunocytochemistry was used to examine the expression of the proteins in ICCs.RESULTS:LPS suppressed the pacemaker currents of ICCs and this could be blocked by AH6809,a prostaglandin E2-EP2 receptor antagonist or NG-Nitro-Larginine Methyl Ester,an inhibitor of nitric oxide(NO) synthase.Toll-like receptor 4,inducible NO synthase or cyclooxygenase-2 immunoreactivity by specific antibodies was detected on ICCs.Catalase(antioxidant agent) had no action on LPS-induced action in ICCs.LPS actions were blocked by nuclear factor kB(NF-kB) inhibitor,actinomycin D(a gene transcription inhibitor),PD 98059(a p42/44 mitogen-activated protein kinases inhibitor) or SB 203580 [a p38 mitogen-activated protein kinases(MAPK) inhibitor].SB 203580 also blocked the prostaglandin E2-induced action on pacemaker currents in ICCs but not NO.CONCLUSION:LPS inhibit the pacemaker currents in ICCs via prostaglandin E2-and NO-dependent mechanism through toll-like receptor 4 and suggest that MAPK and NF-kB are implicated in these actions.展开更多
本试验旨在考察牛膝多糖(ABPS)对脂多糖(LPS)免疫应激下仔猪空肠上皮细胞(IPEC-J2)促炎细胞因子分泌和表达的影响,并探讨ABPS调控IPEC-J2免疫应激可能的作用机制。选用4~5代的IPEC-J2,培养基中分别添加0(对照)、300、600、900、1 200μg...本试验旨在考察牛膝多糖(ABPS)对脂多糖(LPS)免疫应激下仔猪空肠上皮细胞(IPEC-J2)促炎细胞因子分泌和表达的影响,并探讨ABPS调控IPEC-J2免疫应激可能的作用机制。选用4~5代的IPEC-J2,培养基中分别添加0(对照)、300、600、900、1 200μg/m L ABPS和10μg/m L LPS,每组12个重复,每孔为1个重复。培养72 h后,采用酶联免疫吸附测定(ELISA)方法检测ABPS对促炎细胞因子白细胞介素1(IL-1)、白细胞介素6(IL-6)、白细胞介素8(IL-8)和肿瘤坏死因子α(TNF-α)分泌量的影响,采用实时定量PCR测定Toll样受体4(TLR4)、核转录因子κB(NF-κB)的mRNA表达量,采用Western blot法测定TLR4、NF-κB、磷酸化核转录因子κB(p-NF-κB)蛋白表达量。结果显示:与对照组相比,300、600、900和1 200μg/m L ABPS组能显著减少IL-1、IL-6、IL-8和TNF-α的分泌量(P<0.05);300μg/m L ABPS组能显著减少p-NF-κB蛋白的表达量(P<0.05),900和1 200μg/m L ABPS组能显著减少TLR4、NF-κB的mRNA和NF-κB蛋白的表达量(P<0.05)。由此可见,ABPS通过TLR4/NF-κB信号转导途径来调控促炎细胞因子的分泌,从而缓解免疫应激,低浓度ABPS通过直接抑制NF-κB磷酸化过程来降低免疫应激,高浓度ABPS则是通过抑制TLR4 mRNA、NF-κB mRNA和NF-κB蛋白的表达量来缓解免疫应激。展开更多
基金Supported by Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Education,Science and Technology,No.2012-0001335
文摘AIM:To investigate lipopolysaccharide(LPS) related signal transduction in interstitial cells of Cajal(ICCs) from mouse small intestine.METHODS:For this study,primary culture of ICCs was prepared from the small intestine of the mouse.LPS was treated to the cells prior to measurement of the membrane currents by using whole-cell patch clamp technique.Immunocytochemistry was used to examine the expression of the proteins in ICCs.RESULTS:LPS suppressed the pacemaker currents of ICCs and this could be blocked by AH6809,a prostaglandin E2-EP2 receptor antagonist or NG-Nitro-Larginine Methyl Ester,an inhibitor of nitric oxide(NO) synthase.Toll-like receptor 4,inducible NO synthase or cyclooxygenase-2 immunoreactivity by specific antibodies was detected on ICCs.Catalase(antioxidant agent) had no action on LPS-induced action in ICCs.LPS actions were blocked by nuclear factor kB(NF-kB) inhibitor,actinomycin D(a gene transcription inhibitor),PD 98059(a p42/44 mitogen-activated protein kinases inhibitor) or SB 203580 [a p38 mitogen-activated protein kinases(MAPK) inhibitor].SB 203580 also blocked the prostaglandin E2-induced action on pacemaker currents in ICCs but not NO.CONCLUSION:LPS inhibit the pacemaker currents in ICCs via prostaglandin E2-and NO-dependent mechanism through toll-like receptor 4 and suggest that MAPK and NF-kB are implicated in these actions.
文摘本试验旨在考察牛膝多糖(ABPS)对脂多糖(LPS)免疫应激下仔猪空肠上皮细胞(IPEC-J2)促炎细胞因子分泌和表达的影响,并探讨ABPS调控IPEC-J2免疫应激可能的作用机制。选用4~5代的IPEC-J2,培养基中分别添加0(对照)、300、600、900、1 200μg/m L ABPS和10μg/m L LPS,每组12个重复,每孔为1个重复。培养72 h后,采用酶联免疫吸附测定(ELISA)方法检测ABPS对促炎细胞因子白细胞介素1(IL-1)、白细胞介素6(IL-6)、白细胞介素8(IL-8)和肿瘤坏死因子α(TNF-α)分泌量的影响,采用实时定量PCR测定Toll样受体4(TLR4)、核转录因子κB(NF-κB)的mRNA表达量,采用Western blot法测定TLR4、NF-κB、磷酸化核转录因子κB(p-NF-κB)蛋白表达量。结果显示:与对照组相比,300、600、900和1 200μg/m L ABPS组能显著减少IL-1、IL-6、IL-8和TNF-α的分泌量(P<0.05);300μg/m L ABPS组能显著减少p-NF-κB蛋白的表达量(P<0.05),900和1 200μg/m L ABPS组能显著减少TLR4、NF-κB的mRNA和NF-κB蛋白的表达量(P<0.05)。由此可见,ABPS通过TLR4/NF-κB信号转导途径来调控促炎细胞因子的分泌,从而缓解免疫应激,低浓度ABPS通过直接抑制NF-κB磷酸化过程来降低免疫应激,高浓度ABPS则是通过抑制TLR4 mRNA、NF-κB mRNA和NF-κB蛋白的表达量来缓解免疫应激。
