维拉帕米对大鼠创伤失血性休克合并内毒素血症模型干预机制的初步探讨

Effects of calcium antagonist verapamil on traumatic hemorrhagic shock with endotoxemia in rats

目的 观察钙通道阻滞剂维拉帕米对大鼠创伤失血性休克复合内毒素血症模型的干预作用 ,及其是否参与调控p38MAPK的激活。方法 将 18只雄性SD大鼠随机分为对照组、致伤组及维拉帕米干预组。分别于给予内毒素 (1 4mg/kg)后 90min ,检测各组大鼠血浆中TNF α及IL 10的含量 ,并检测各组大鼠腹腔巨噬细胞内p38MAPK磷酸化水平。另外取 6只大鼠分离其腹腔巨噬细胞 ,体外给予生理盐水、维拉帕米及SB2 0 35 80预处理 30min后 ,再予LPS (10 0ng/ml)刺激 ,90min后收集细胞上清液 ,检测TNF α及IL 10的含量 ,同时检测各组巨噬细胞内p38MAPK磷酸化水平。结果 致伤组血浆中TNF α、IL 10的含量明显较对照组高 (P <0 0 1) ,而维拉帕米组血浆中TNF α的含量则明显低于致伤组 (P <0 0 1) ,且IL 10的含量明显高于致伤组 (P <0 0 1)。维拉帕米对致伤组增高了的腹腔内p38MAPK的磷酸化水平无明显影响(P >0 0 5 )。SB2 0 35 80可明显抑制腹腔在内毒素诱导下生成的TNF α及IL 10 (P <0 0 1) ,并明显抑制p38MAPK的磷酸化 (P <0 0 1) ;而维拉帕米不仅可明显抑制腹腔在内毒素诱导下生成TNF α (Ρ <0 0 1) ,且使内毒素诱导的IL 10生成进一步增加 (P <0 0 1) ,但对P38MAPK的磷酸化水平无明显影响 (P >0 0 5 )。结论 在?

Objective To evaluate the effects of calcium antagonist verapamil on the murine model of traumatic hemorrhagic shock with endotoxemia in rats,and to investigate the changes of phosphorylated form of p38MAPK in macrophages.Methods Eighteen male Sprague Dawley rats were randomly divided into three groups with six in every group:control group,trauma group,and verapmil treatment group. Plasma concentrations of TNF-α and IL-10 were measured with ELISA in every group 90 minutes after LPS treatments. Peritoneal macrophages were collected at the same time, and cellular protein of peritoneal macrophages were extracted and analyzed by Western blot for the phosphorylated form of p38MAPK. Peritoneal macrophages were collected from another six male Sprague Dawley rats and separated randomly in six-well plates with 5 × 10 6 cells per well. Cells were exposed to LPS(100 ng/ml) in the presence of verapamil(50 μmol/L) or SB203580(10 μmol/L). TNF-α and IL-10 expressions were measured by ELISA on cellular supernatants after 90 minutes of LPS treatments. Cellular protein was extracted and analyzed by Western blot for the phosphorylated form of p38MAPK.Results Rats in trauma group had higher plasma concentrations of TNF-α and IL-10 and higher level of phosphorylated p38MAPK in the peritoneal macrophages when compared with the control group (P<0.01) . Rats in verapamil treatment group had lower plasma concentrations of TNF-α and higher plasma concentration of IL-10 when compared with trauma group (P<0.01) . Verapamil had no significant effect on trauma-induced p38MAPK phosphorylation in macrophages (P>0.05).Cells pretreated with SB203580 inhibited production of LPS-induced TNF-α,IL-10 and p38MAPK phosphorylation(P<0.01). Cells pretreated with verapamil inhibitted procluction of LPS-induced TNF-α,but enhanced production of IL-10. Verapamil had no significant effect on p38MAPK phosphorylation (P>0.05). Conclusion The expression of p38MAPK phosphorylation is higher in macrophages during traumatic hemorrhagic shock with endotoxemia.Verapamil can inhibit the excessive inflammation in the murine model of traumatic hemorrhagic shock with endotoxemia. But its inhibitory is probably independent of p38MAPK activation.