鳖甲煎改良方对大鼠肝纤维化的防治作用

A modified recipe of Turtle Shell Decoction reduces experimental hepatic fibrosis in rats

目的:研究鳖甲煎改良方对大鼠肝纤维化(HF)的防治作用及对TGF-β1和Sman3/7蛋白表达的影响,探讨其机制.方法:SD♂大鼠90只,随机取10只作为正常对照组(A组),其余大鼠用皮下注射40%CCl4橄榄油油剂3mL/kg诱导大鼠肝纤维化模型8wk,于第2周时随机处死5只大鼠证实HF形成后,将剩下的大鼠随机分为肝纤维化模型组(B组)、鳖甲煎改良方高剂量组[C组,28.4g/(kg·d)]、中剂量组[D组,14.2g/(kg·d)]、低剂量组[E组,7.1g/(kg·d)]、复方鳖甲软肝片组[F组,0.6g/(kg·d)],每组15只.C、D、E、F组给予10mL/(kg·d)相应药液灌胃治疗,A、B组同时给予等剂量的生理盐水灌胃处理.8wk后采血测定血清丙氨酸氨基转移酶(ALT)、草氨酸氨基转移酶(AST)、白蛋白和球蛋白含量;取肝组织作HE染色观察肝纤维化程度变化;Westernblot方法检测TGF-β1、Smad3和Smad7蛋白的表达.结果:8wk后,与B组比较,C、D、E组和F组肝小叶结构破坏明显减轻,肝纤维化程度分级较B组明显好转;ALT和AST含量显著降低(P<0.01),白蛋白含量显著增加(37.85g/L±3.08g/L,38.25g/L±1.90g/L,38.13g/L±1.50g/L,38.09g/L±3.27g/Lvs27.05g/L±4.81g/L,均P<0.01);TGF-β1、Smad3蛋白表达显著减少(TGF-β1:0.127±0.014,0.122±0.051,0.126±0.027,0.119±0.020vs0.332±0.074,均P<0.01;Smad3:0.415±0.057,0.427±0.074,0.425±0.080,0.432±0.075vs0.527±0.054,均P<0.01),Smad7蛋白的表达明显增加(0.308±0.077,0.326±0.086,0.315±0.071,0.348±0.065vs0.185±0.059,均P<0.01),且C、D、E组和F组间疗效比较无明显差异.结论:鳖甲煎改良方能够显著减轻CCl4导致的大鼠肝纤维化程度,其作用机制可能与鳖甲煎改良方调控TGF-β1和Sman3/7信号转导蛋白表达有关.

AIM： To investigate the effects of treatment with a modified recipe of Turtle Shell Decoction （MRTSD） on the expression of TGF-β1 and Smad 3/7 in experimental hepatic fibrosis in rats, and to explore its anti-fibrotic mechanism. METHODS： Ninety Sprague-Dawley adult rats were used in this study, 10 of which were randomly selected as normal controls （A）, and the rest were used to induce hepatic fibrosis by multiple subcutaneous injections of 40% carbon tetrachloride （CCl4, 3 mL/kg） for 8 wk. At week 2, five rats were executed to confirm the formation of liver fibrosis, and the rest of rats were randomly and equally divided into pathological model group （B）, high-dose [28.4 g/（kg·d）] MRTSD group （C）, medium-dose [14.2 g/（kg·d）] MRTSD group （D）, low-dose [7.1 g/（kg·d）] MRTSD group, and Fufang Biejia Ruangan Tablet-treated group [F, 0.6 g/（kg·d）]. Drugs were intragastrically administrated at a volume of 10 mL/（kg·d） in groups C-E, whereas the rats of groups A and B were given the same volume of physiological saline. At week 8, serum levels of ALT, AST, albumin, and globulin were measured. The formation of hepatic fibrosis was confirmed by HE staining. The protein expres- sion of TGF-β1 and Smad 3/7 in liver tissue was detected by Western blot. RESULTS： In groups C-F at week 8, the structure of liver lobules was almost restored to normal; HF score was obviously reduced; the contents of ALT and AST were significantly decreased （P 0.01）; the content of albumin was significantly increased （37.85 g/L ± 3.08 g/L, 38.25 g/L ± 1.90 g/L, 38.13 g/L ± 1.50 g/L, 38.09 g/L ± 3.27 g/L vs 27.05 g/L ± 4.81 g/L, all P 0.01）; the protein expression of TGF-β1 and Smad 3 was remarkably down-regulated （TGF-β1： 0.127 ± 0.014, 0.122 ± 0.051, 0.126 ± 0.027, 0.119 ± 0.020 vs 0.332 ± 0.074, all P 0.01; Smad 3： 0.415 ± 0.057, 0.427 ± 0.074, 0.425 ± 0.080, 0.432 ± 0.075 vs 0.527 ± 0.054, all P 0.01）; and the expression of Smad 7 was significantly upregulated （0.308 ± 0.077, 0.326 ± 0.086, 0.315 ± 0.071, 0.348 ± 0.065 vs 0.185 ± 0.059, all P 0.01） when compared with group B. There were no significant differences in the above parameters between MRTSD-and Fufang Biejia Ruangan Tablet-treated groups. CONCLUSION： MRTSD can reverse CCl4-induced liver fibrosis in rats possibly by regulating the protein expression of TGF-β1 and Smad 3/7.