[目的]丙泊酚改善重症脓毒症小鼠脑保护作用的研究。[方法]将48只雄性小鼠随机分为对照组、LPS组、PF组、PF+ML385组,每组12只。通过苏木素-伊红染色评估各组小鼠海马组织的病理变化;通过原位末端标记法分析海马组织中神经元的凋亡率;...[目的]丙泊酚改善重症脓毒症小鼠脑保护作用的研究。[方法]将48只雄性小鼠随机分为对照组、LPS组、PF组、PF+ML385组,每组12只。通过苏木素-伊红染色评估各组小鼠海马组织的病理变化;通过原位末端标记法分析海马组织中神经元的凋亡率;通过酶联免疫吸附实验分析血清中TNF-α、IL-6、SOD和CAT的水平;通过蛋白免疫印迹实验分析海马组织中TLR4/MD-2的蛋白表达水平。[结果]与LPS组比较,PF组与PF+ML385组小鼠海马组织区域炎症细胞聚集减少,神经元损伤减轻,神经元凋亡率降低,血清炎症因子(TNF-α和IL-6)水平降低,血清抗氧化酶(SOD和CAT)活性水平增加,海马组织的TLR4、MD-2蛋白表达水平降低。与PF组比较,PF+ML385组小鼠海马组织神经元凋亡率降低(13.26%±2.19%vs 9.28%±1.58%;P<0.05),血清炎症因子(TNF-α、IL-6)水平降低(15.61±1.29 vs 10.28±1.46 pg/mL;30.28±1.11 vs 23.56±2.13 pg/mL;P<0.05),血清抗氧化酶(SOD、CAT)活性水平增加(13.69±1.72 vs 19.23±2.39 U/mg protein;12.97±2.26 vs 17.38±1.47 U/mg protein;P<0.05),海马组织的TLR4、MD-2蛋白表达水平降低(0.65±0.03 vs 0.33±0.09;0.72±0.08 vs 0.41±0.04;P<0.05)。[结论]丙泊酚能够改善脓毒症小鼠海马组织的神经元损伤,减少神经元凋亡率和炎症因子水平,增加抗氧化酶水平。丙泊酚的这一作用与抑制TLR4/MD-2通路密切相关。展开更多
AIM: To investigate the expression of toll-like receptor 4(TLR4) and MD-2 gene and protein in Kupffer cells (KCs)and their role in ischemia-reperfusion (IR) injury of ratliver graft.METHODS: KCs were isolated at 0 (co...AIM: To investigate the expression of toll-like receptor 4(TLR4) and MD-2 gene and protein in Kupffer cells (KCs)and their role in ischemia-reperfusion (IR) injury of ratliver graft.METHODS: KCs were isolated at 0 (control group), 2, 12,24 h (IR group) following IR in rat liver graft, mRNA expressionof TLR4 and MD-2 was detected by RT-PCR analysis, proteinexpression of TLR4/MD-2 was detected by flow cytometric(FCM) analysis, and tumor necrosis factor-(~ (TNF-(~) levelin supernatant was measured by ELISA. Then isolated KCswere incubated with anti-TLR4 polyclonal antibody (anti-TLR4 group), and TNF-(~ level was measured again.RESULTS: The mRNA and protein expression of TLR4/MD-2 and the level of TNF-(~ in IR group increased significantlyat 2 h following IR, and reached the maximum at 12 h, andslightly decreased at 24 h, but were still significantly higherthan those in the control group (P<O.01). The expression ofthese factors was markedly decreased after anti-TLR4antibody treatment as compared with the IR group (P<O.01).CONCLUSION: Lipopolysaccharide (LPS) following IR canup-regulate TLR4/MD-2 gene and protein expression inKCs, and synthesize cytokine TNF-(~. Anti TLR4 antibodycan inhibit the production of TNF-(~ induced by LPS. TLR4and its partner molecule MD-2 may play an important rolein Kupffer cell activation and IR injury.展开更多
文摘[目的]丙泊酚改善重症脓毒症小鼠脑保护作用的研究。[方法]将48只雄性小鼠随机分为对照组、LPS组、PF组、PF+ML385组,每组12只。通过苏木素-伊红染色评估各组小鼠海马组织的病理变化;通过原位末端标记法分析海马组织中神经元的凋亡率;通过酶联免疫吸附实验分析血清中TNF-α、IL-6、SOD和CAT的水平;通过蛋白免疫印迹实验分析海马组织中TLR4/MD-2的蛋白表达水平。[结果]与LPS组比较,PF组与PF+ML385组小鼠海马组织区域炎症细胞聚集减少,神经元损伤减轻,神经元凋亡率降低,血清炎症因子(TNF-α和IL-6)水平降低,血清抗氧化酶(SOD和CAT)活性水平增加,海马组织的TLR4、MD-2蛋白表达水平降低。与PF组比较,PF+ML385组小鼠海马组织神经元凋亡率降低(13.26%±2.19%vs 9.28%±1.58%;P<0.05),血清炎症因子(TNF-α、IL-6)水平降低(15.61±1.29 vs 10.28±1.46 pg/mL;30.28±1.11 vs 23.56±2.13 pg/mL;P<0.05),血清抗氧化酶(SOD、CAT)活性水平增加(13.69±1.72 vs 19.23±2.39 U/mg protein;12.97±2.26 vs 17.38±1.47 U/mg protein;P<0.05),海马组织的TLR4、MD-2蛋白表达水平降低(0.65±0.03 vs 0.33±0.09;0.72±0.08 vs 0.41±0.04;P<0.05)。[结论]丙泊酚能够改善脓毒症小鼠海马组织的神经元损伤,减少神经元凋亡率和炎症因子水平,增加抗氧化酶水平。丙泊酚的这一作用与抑制TLR4/MD-2通路密切相关。
文摘目的探讨脂多糖作用小鼠RAW264.7巨噬细胞24 h内细胞表面TLR4/CD14/MD-2受体复合物表达的变化特点。方法分别用低剂量(100 ng/ml)和高剂量(1 000 ng/ml)脂多糖刺激RAW264.7,应用RT-PCR检测TLR4、CD14、MD-2m RNA水平的变化,ELISA检测细胞上清TNF-α含量的变化。结果低剂量脂多糖下调TLR4 m RNA表达,14 h降至最低,24 h维持在低水平表达;CD14和MD-2 m RNA表达增加。高剂量脂多糖上调TLR4 m RNA表达,1 h上升至高峰,此后维持在高水平表达;CD14和MD-2 m RNA表达也明显增强。高剂量脂多糖作用后TNF-α含量显著高于低剂量脂多糖(P<0.05)。结论高剂量脂多糖逆转低剂量脂多糖下调的TLR4表达,提高TLR4/CD14/MD-2受体复合物的表达水平,放大下游信号转导通路的炎症效应。
基金Supported by the National Natural Science Foundation of China,No.30300337,30200278,30170919
文摘AIM: To investigate the expression of toll-like receptor 4(TLR4) and MD-2 gene and protein in Kupffer cells (KCs)and their role in ischemia-reperfusion (IR) injury of ratliver graft.METHODS: KCs were isolated at 0 (control group), 2, 12,24 h (IR group) following IR in rat liver graft, mRNA expressionof TLR4 and MD-2 was detected by RT-PCR analysis, proteinexpression of TLR4/MD-2 was detected by flow cytometric(FCM) analysis, and tumor necrosis factor-(~ (TNF-(~) levelin supernatant was measured by ELISA. Then isolated KCswere incubated with anti-TLR4 polyclonal antibody (anti-TLR4 group), and TNF-(~ level was measured again.RESULTS: The mRNA and protein expression of TLR4/MD-2 and the level of TNF-(~ in IR group increased significantlyat 2 h following IR, and reached the maximum at 12 h, andslightly decreased at 24 h, but were still significantly higherthan those in the control group (P<O.01). The expression ofthese factors was markedly decreased after anti-TLR4antibody treatment as compared with the IR group (P<O.01).CONCLUSION: Lipopolysaccharide (LPS) following IR canup-regulate TLR4/MD-2 gene and protein expression inKCs, and synthesize cytokine TNF-(~. Anti TLR4 antibodycan inhibit the production of TNF-(~ induced by LPS. TLR4and its partner molecule MD-2 may play an important rolein Kupffer cell activation and IR injury.
