磷脂型二十二碳六烯酸和二十碳五烯酸对脂多糖所致小鼠急性肝损伤的保护作用及机制

Protective Effect and Mechanism of Docosahexaenoic Acid-Enriched Phospholipids and Eicosapentaenoic Acid-Enriched Phospholipids on Lipopolysaccharide-Induced Acute Liver Injury in Mice

目的:探讨磷脂型二十二碳六烯酸(docosahexaenoic acid-enriched phospholipids,DHA-PL)和磷脂型二十碳五烯酸(eicosapentaenoic acid-enriched phospholipids,EPA-PL)对脂多糖(lipopolysaccharide,LPS)所致小鼠急性肝损伤的保护作用及其机制。方法:以南海鸢乌贼(Sthenoteuthis oualaniensis)卵和冰岛刺参(Cucumaria frondosa)体壁为原料,分离提取DHA-PL和EPA-PL。将C57BL/6J雄性小鼠随机分为Control组、模型组、LPS+DHA-PL组和LPS+EPA-PL组,Control组和模型组小鼠每天灌胃20 mL/kg mb生理盐水,其他组以400 mg/kg mb剂量分别每天灌胃受试物,灌胃体积为10 mL/kg mb,连续干预28 d;第29天Control组腹腔注射无菌生理盐水,其他组注射LPS(10 mg/kg mb),注射体积为10 mL/kg mb,建立急性肝损伤模型;称量小鼠体质量和肝质量,计算肝指数;测定血清中丙氨酸氨基转移酶(alanine aminotransferase,ALT)和天冬氨酸氨基转移酶(aspartate aminotransferase,AST)活力;苏木精-伊红染色观察肝组织病理改变情况;采用实时荧光定量聚合酶链式反应和酶联免疫吸附试剂盒测定肝脏中肿瘤坏死因子(tumor necrosis factor,TNF)-α、白细胞介素(interleukin,IL)-1β和IL-6的mRNA相对表达量和含量;采用蛋白免疫印迹法考察肝脏中丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)如细胞外调节蛋白激酶(extracellular regulated protein kinases 1/2,ERK1/2)、c-Jun氨基末端激酶(c-Jun N-terminal kinase,JNK)、p38以及核因子(nuclear factor,NF)-κB?p65蛋白磷酸化水平。结果:DHA-PL和EPA-PL预防性干预能有效保护LPS所致小鼠急性肝损伤,降低肝指数以及血清ALT和AST活力,下调肝脏中TNF-α、IL-1β和IL-6含量,降低MAPK和NF-κB p65蛋白磷酸化水平。结论:DHA-PL和EPA-PL预防性干预可保护LPS所致小鼠急性肝损伤,其作用机制可能依赖于其对MAPK和NF-κB信号通路的调控。

Purpose: To investigate the protective effect and mechanism of docosahexaenoic acid-enriched phospholipids(DHA-PL) and eicosapentaenoic acid-enriched phospholipids(EPA-PL) on lipopolysaccharide(LPS)-induced acute liver injury. Methods: DHA-PL and DHA-PL were obtained from the eggs of Sthenoteuthis oualaniensis and the body wall of Cucumaria frondosa, respectively. C57BL/6 male mice were randomly divided into four groups, i.e. control group,LPS-induced model group, LPS + DHA-PL group and LPS + EPA-PL group. The mice in the LPS + DHA-PL and LPS + EPA-PL groups were given DHA-PL and EPA-PL via the intragastric route at 400 mg/kg mb on a daily basis for 28 days, respectively, while those in the control and model groups were gavaged with normal saline. The mice in the control group were injected with normal saline intraperitoneally at day 29, while those in the other groups were injected with LPS(10 mg/kg mb) at an injection volume of 10 mL/kg mb to establish a model of acute liver injury. The pathological changes of liver tissues were observed by hematoxylin-eosin(HE) staining. Serum alanine aminotransferase(ALT) and aspartate aminotransferase(AST) activities were measured. The transcriptional levels and contents of tumor necrosis factor(TNF)-α,interleukin(IL)-1β and IL-6 in liver tissues were detected by real-time quantitative polymerase chain reaction(PCR) and enzyme linked immunosorbent assay(ELISA), respectively. The phosphorylation levels of mitogen-activated protein kinases(MAPKs) such as extracellular regulated protein kinases 1/2(ERK1/2), c-Jun N-terminal kinase(JNK), p38, and nuclear factor(NF)-κB p65 were determined by Western blotting. Results: DHA-PL and EPA-PL intervention effectively alleviated LPS-induced acute liver injury and reduced hepatic index and serum ALT and AST activities, decreased the hepatic mRNA expression and contents of TNF-α, IL-1β and IL-6, and down-regulated the phosphorylation of ERK1/2, JNK, p38 and NF-κB p65. Conclusion: DHA-PL and EPA-PL can prevent LPS-induced acute liver injury in mice, and the mechanism may depend on regulation of the MAPKs and NF-κB signaling pathways.