重组纤维连接蛋白N端肝素结合域多肽抗内毒素所致小鼠肝衰竭的作用

目的 通过动物实验研究重组纤维连接蛋白N端肝素结合域多肽(rhFNHN-29多肽)对内毒素血症肝衰竭的治疗作用.方法 用内毒素脂多糖(LPS,Sigma)和半乳糖胺(Galactosamine GalN,Sigma)注射小鼠腹腔,建立内毒素血症肝衰竭小鼠模型.实验小鼠随机分两组,1组为rhFNHN-29多肽治疗组,1组为生理盐水对照组,另设正常对照组.治疗组分别于注射(LPS和GalN)前半小时,后1、2、3 h尾静脉注射rhFNHN-29多肽10 mg/kg,生理盐水对照组注射等体积生理盐水,正常对照组注射等体积生理盐水.腹腔注射药物6 h,眶静脉采血检测小鼠血浆TNFα,72 h后计算小鼠死亡率,处死活鼠,取其肝、肾、脾、肺、心、脑组织进行病理组织学观察,肝组织做电镜超微结构观察,取肝组织对其TNFα、IL-1β、IL-6的表达进行PCR分析.结果 生理盐水对照组小鼠72 h死亡率为70%,多肽治疗组死亡率仅为15%.病理组织学观察显示生理盐水对照组肝组织呈广泛变性与严重坏死,多肽治疗组肝组织呈小面积变性和轻微坏死.电镜超微结构显示生理盐水埘照组肝细胞呈严重的变性和坏死,多肽治疗组轻微变性.肝细胞因子表达分析显示多肽治疗组小鼠的TNFα、IL-1β、IL-6 mRNA表达水平及血浆TNFα水平(0.86±0.43,0.86±0.31,0.27±0.13和23.6±5.8,P<0.01)显著低于生理盐水对照组小鼠的水平(1.26±0.37,0.98±0.21,0.43±0.17和87.43±16.7,P<0.01).结论 rhFNHN-29多肽对内毒素血症肝衰竭具有明显的预防和治疗作用,其作用机制可能与多肽的抗炎等作用有关.

Recombinant polypeptide of N-terminal heparin-binding domain of fibronectin antagonizes hepatic failure induced by endotoxin in mice

Objective To study the preventive effect of recombinant polypeptide of N-terminal heparin-binding domain of fibronectin on hepatic failure induced by endotoxin in mice. Methods The 40 hepatic failure Balb/C mice were established by intraperitoneal injection of lipopolysaccharide ( LPS) and d-galactosamine(GalN). The mice were randomly divided into two groups,one for polypeptide treatment, the othe for saline treatment. Another 20 mice were used as normal control. Half hour prior to , 1,2 ,and 3 hours after injection of LPS and GalN, the rhFNHN-29 polypeptide (10 mg/kg) was injected through the tail vein of mice. The same volume of saline was given to the saline treated group and the normal control group. Six hours after the injection of LPS and GalN, 250 μl blood was taken from the eye vein of each mouse for plasma TNFα testing, and 72 hours after the injection, mortality rates of the mice of different groups were observated. The liver, lung, heart, kidney, and brain tissues of the survival mice were examined for histopathology after 72 hours . The Liver tissue was also examined for electron micrograph and for mRNA expression of TNFα, IL-1 β, IL-6 by RT-PCR. Results The 72 hours mortality rates in saline-treated and polypeptide treated-mice were 70% and 15% respectively (P < 0. 01 ) . The histopathology showed that necrosis occurred less on the hepatocytes of polypeptide treated mice than on the saline treated ones. The ultrastructure of hepatocyte under the electron microscope showed that cell apparatus of saline treated mice were destroyed and cytoplasm become loose. The expression level of TNFα,IL-l β,IL-6 mRNA on hepatocytes in polypeptide treated mice was significantly lower( 1. 26 ± 0. 37,0. 98 ± 0. 21,0. 43 ± 0. 17,87. 43 ± 16. 7 respectively) than that in the saline treated ones(1.98 ±0. 56,1. 24 ± 0. 35,0. 64 ± 0. 25 and 236. 11 ± 32.7, respectively) (P<0. 01). Similarly, the plasm TNFα level(87.43 ±16.7 ) in polypeptide treated group was significantly lower than that(236. 11 ±32. 7 ) in the saline treated group(P <0. 01). Conclusion The rhFNHN-29 polypeptide can prevent and treat hepatic failure induced by endotoxin . The mechanism by which the polypeptide takes the effect may involve its ability to down-regulate expression of those inflammative factors such as TNFα, IL-1β, IL-6.