石见穿多糖对脂多糖和D-氨基半乳糖胺联合诱导小鼠急性肝衰竭的保护作用

Protective effect of Salvia chinensis Benth. polysaccharides on lipopolysaccharide and D-galactosamine induced acute liver failure in mice

目的研究石见穿多糖(PSSC)对脂多糖(LPS)/D-氨基半乳糖胺(Ga IN)诱导小鼠急性肝衰竭(ALF)的保护作用及可能机制。方法将昆明小鼠随机分为正常组、模型组、PSSC 30和100 mg·kg^(-1)给药组。给药组每日1次,连续给药1周。给药结束后,除正常组外,其余各组ip给予LPS 10μg·kg^(-1)和Gal N700 mg·kg^(-1),制备小鼠ALF模型。HE染色法检测肝组织病理变化;用试剂盒法检测血清谷草转氨酶(GOT)和谷丙转氨酶(GPT)及肝过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、丙二醛(MDA)和谷胱甘肽(GSH)水平;DCFH-DA荧光探针法检测肝组织活性氧(ROS)的相对含量;ELISA法测定血清及肝组织中肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL^(-1)β)和IL-6的含量;胱天蛋白酶3活性测试试剂盒检测小鼠肝组织匀浆中胱天蛋白酶3的活性。结果与正常对照组相比,模型组小鼠肝细胞排列杂乱,胞质皱缩,细胞边界模糊,可见较大量的炎症细胞浸润和明显的肝组织内出血,病理评分明显升高(P<0.01);MDA和ROS含量分别升高至正常对照组的2.2倍和4.3倍(P<0.01),GSH含量下降至51%(P<0.01),抗氧化酶SOD,CAT和GSH-Px的活性分别降低至74%,36%和42%(P<0.01),TNF-α,IL^(-1)β和IL-6的水平有明显提高(P<0.01),胱天蛋白酶3活性升高至正常对照组的5.3倍(P<0.01)。与模型组相比,PSSC组小鼠存活比例明显升高(P<0.01);小鼠肝组织病理评分降低(P<0.01);MDA和ROS含量升高(P<0.01),GSH含量下降(P<0.01);TNF-α,IL^(-1)β和IL-6的含量降低(P<0.01);胱天蛋白酶3活性降低(P<0.01)。结论 PSSC对LPS和Gal N联合诱导的小鼠ALF具有良好的缓解作用,该作用可能与降低肝氧化应激、抑制肝炎症反应和细胞凋亡相关。

OBJECTIVE To explore the protective effect of polysaccharides from Salvia Chinensis Benth. （PSSC） on lipopolysaccharide （LPS） and D-galactosamine （GalN）-provoked mouse acute liver failure （ALF） and the possible molecular mechanism. METHODS Kunming mice were randomly divided into four groups： normal control, model, model＋PSSC 30 and 100 mg-kg-1 groups. PSCC was given once a day and for a week. To establish an ALF model, mice of model and PSSC groups were ip injected with LPS 10 ug-kg-1 and GaIN 700 mg-kg-1 at the end of PSSC treatment. The microscopic structure of the liver was detected by HE staining. Serum and hepatic biochemical parameters of glutamic-oxalacetic transaminase （GOT）, glutamic-pyruvic transaminase （GPT）, catalase （CAT）, superoxide dismutase （SOD）, glutathione peroxidase （GSH-Px）, malondialdehyde （MDA）, and glutathione （GSH） were detected by colorimetric methods. The relative content of hepatic reactive oxygen species （ROS） was measured by DCFH-DA fluorescent probes. Levels of cytokines tumor necrosis factor-α （TNF-α）, interleukin 1β（IL-1β） and IL-6 in the serum and liver were detected by ELISA. Activity of caspase 3 in liver homogenates was detected by aspase 3 activity assay kit. RESULTS Compared with normal control group, in the liver of model group, hepatocytes were arrayed in disorder, cytoplasm of hepatocytes shrank, and boundaries between cells were fuzzy, the infiltration of a large number of inflammatory cells and tissue hemorrhage could be detected, pathological scores were elevated significantly （P〈0.01）, levels of MDA and ROS in the liver of AI_F model mice were elevated to 2.2 and 4.3 times that of the normal control, respectively （P〈0.01）, the level of GSH decreased to 51% （P〈0.01）, and the activities of SOD, CAT and GSH-Px declined to 74%, 36% and 42%, respectively （P〈0.01）. Levels of TNF-α, IL-1β and IL-6 in the serum and liver of model group were increased （P〈0.01）, and caspase 3 activity was increased to 5.3 times that of the normal control （P〈0.01）. Compared with the model group, the number of surviving mice in PSSC groups increased, liver pathological scores declined （P〈0.01）, levels of MDA and ROS increased （P〈0.01）, levels of GSH, TNF-α, IL-1β and IL-6 in the liver and serum declined （P〈0.01）, and caspase 3 activity decreased （P〈0.01）. CONCLUSION PSSC is able to alleviate LPS and GaiN-induced ALF in mice. Inhibition of hepatic oxidative stress, inflammatory response, and cell apoptosis is possibly implicated in the protective effect of PSSC.