摘要目的 探讨抑制核因子-κB(NF-κB)后热休克蛋白70(HSP70)在创伤失血性休克肝组织中的变化及其对肝脏结构和功能的影响.方法 雄性健康Wistar大鼠66只,采用双侧股骨骨折伴失血性休克创伤模型,随机分成正常对照组6只,创伤休克组30只,NF-κB抑制伴创伤休克组30只,NF-κB抑制采用致伤前1 h腹腔注射二硫代氨基甲酸吡咯烷(PDTC)200mg/kg.动态观察伤后0.5、2、4、6、8h大鼠肝组织NF-κB、HSP70、肝脏病理、肝功能、TNF-α、IL-6等变化.NF-κB采用EMSA法测定结合活性,HSP70采用免疫印迹法测定其蛋白含量,并进行计算机图像分析.数据采用SPSS 12.0软件分析,两组间比较采用成组资料的t检验.结果 NF-κB的活性伤后迅速升高,伤后6 h达到高峰;HSP70伤后2 h较正常对照相比[(10.8±1.1)、vs.(4.7±0.5),P＜0.01],伤后6 h达到高峰,和正常组相比[(23.0±1.7)vs.(4.7±0.5),P＜0.01].TNFα、IL-6伤后逐渐升高,并于伤后6 h达到高峰,和正常组相比[TNF-α(173.7±12.1)vs.(30.8±1.8)pg/ml,P＜0.01;IL-6(175.5±12.5)vs.(10.4±0.7)pg/ml,P＜0.01];伤后8 h光镜下可见肝窦内淤血明显,有大量炎性细胞浸润;血清ALT、TB伤后4 h开始增高,8 h达到峰值,和创伤组相比[ALT(640.6±80.2)vs.(536.8±60.0)nmol-1·L-1,P＜0.01;TB(4.7±1.1)vs.(1.6±0.2)mol/L,P＜0.01].抑制NF-κB再致伤后,HSP70在肝组织中表达仍然较高,但在伤后各个时相点的表达均较未抑制NF-κB创伤性休克伤组明显回落;伤后6 h和创伤组相比[(16.9±4.4)vs.(23.0±1.7),P＜0.05].TNF-α、IL-6伤后各个时相点均迅速回落,伤后6 h,和创伤组相比[TNF-α(135.2±10.2)vs.(173.7±12.1)pg/ml,P＜0.05;IL-6(113.0 4±10.8)vs.(175.5±12.5)pg/ml,P＜0.05];肝脏大体淤血、肿胀明显减轻;伤后8 h光镜下可见肝细胞变性明显好转,肝窦内淤血减轻,仅见少许淋巴细胞及中性粒细胞浸润;伤后4 h,血清ALT、TB即明显下降,和未抑制组相比[ALT(540.8±66.2)vs.(640.6±80.2)nmol-1·L-1,P＜0.05;TB(2.3±0.3)vs.(4.7±1.1)mol/L,P＜0.05].结论 NF-κB、HSP70参与了严重创伤失血性休克后肝损伤与抗损伤的发生,抑制NF-κB的活性有助于减轻创伤失血性休克后肝脏的急性损害,NF-κB、HSP70可作为反映创伤休克后肝脏损害程度的重要应激指标.

abstractsObjective To study the effects of the inhibition of nuclear factor kappa-B ( NF-κB) , on the hepatic heat shock protein 70 (HSP70) expression as well as on the changes of hepatic function and ultrastructure in a rodent model of hemonhageic shock. Method Hemorrhagic shock was produced by inducing bilateral femoral fractures in male Wistar rats. Intraperitoneal injection of pyrrolidine dithiocarbamate(PDTC)was used to inhibit NF-κB activation 1 hour before induction of shock. A total of 66 adult male Wistar rats were randomly divided into 3 groups: control group (Control, n = 6), trauma shock (TS, n = 30), and NF-κB inhibition followed by trauma shock (NF-κB inhibition, n =30). Measurements of hepatic NF-KB and HSP70, hepatic function bio-markers, TNF-α and IL-6 were obtained 0.5, 2, 4, 6, 8 hours after trauma. Histopathological changes in liver tissues were also noted. Hepatic expression of NF-κB was determined by using electrophoretic mobility shift assay, while HSP70 was assayed by western blot and analyzed with computer imaging. Results In rats with trauma shock, both hepatic NF-κB activity and HSP70 expression increased significantly compared to the control group, reaching peaks at 6 hour post injury. Serum alanine transferase (ALT) and total bilirubin (TB) also rose significantly,reaching peaks at 8 hours post trauma. Light microscopy revealed hepatic congestion with infiltration of inflammatory cells into hepatic sinusoid in the TS group at 8 hours. Inhibiting the activity of NF-κB one hour before trauma significantly decreased expression of HSP70 at 6 hours post trauma [16.9±4.4 (NF-κB inhibition) vs. 23.0±1.7 (TS), P ＜ 0.05]. In addition,levels TNF-α and IL-6 in the liver tissue also decreased, and hepatic congestion as well as hepatic cell degeneration were ameliorated, showing minimal inflammatory infiltrates in the hepatic sinusoids. NF-κB inhibition also significantly lowered the levels of ALT and TB at 4 hours post trauma [ALT, 540.8 ±66.2 nmol/L (NF-KB inhibition) vs. 640.6±80.2 nmol/L (TS), P ＜ 0.05; TB,2.3±0.3 mol/L (NF-κB inhibition) vs. 4.7 ±1.1 mol/L (TS), P ＜ 0.05]. Conclusions NF-κB and HSP70 are involved in the pathogenesis of hepatic injury during hemorrhagic shock, and the degree of NF-κB activity and HSP70 expression may be consistent with the extent of hepatocellular damage. Inhibition of NF-κB helps ameliorate liver injury due to trauma shock.