长期酒精暴露对SMS2^-/-小鼠肝脑的损伤及机制

Molecular mechanism of the chronic alcohol exposure induced injury of the liver-brain in SMS_2knockout mice

目的观察长期酒精暴露引起的肝脑损伤与氧化应激的关系,探讨酒精引起多系统损伤的机制,神经酰胺在酒精暴露诱导海马应激损伤中的调节作用。方法建立C57BL6J野生小鼠和神经鞘磷脂合成酶2基因敲除(SMS-/-2)小鼠酒精暴露模型,利用HE和Masson染色观察肝脏细胞和结构变化,透射电子显微镜观察肝脏超微结构变化,免疫荧光染色法观察对照组与模型组小鼠海马CA1区氧化应激引起的阳性细胞数量变化,免疫印迹技术检测海马组织c-Fos、核因子-κB(NF-κB)激活蛋白的相对表达量。结果酒精暴露导致野生型和SMS-/-2小鼠肝细胞脂肪变性和肝纤维化,并有Mallory小体形成和炎症细胞浸润。免疫组织化学染色显示,酒精暴露后野生型和SMS-/-2小鼠海马c-Fos、NF-κB阳性细胞表达增多(P<0.01)且具有剂量依赖性;与相同处理条件的野生型小鼠相比,SMS-/-2小鼠海马c-Fos、NF-κB阳性细胞表达较多(P<0.05)。免疫印迹技术检测各组间小鼠海马组织cFos、NF-κB蛋白相对表达量与上述结果一致。结论长期酒精暴露引起肝脏和神经系统损伤的病理基础是氧化应激和炎症反应;神经酰胺参与酒精暴露诱导与促进肝脑细胞损伤的过程;酒精、胰岛素抵抗和神经酰胺相互作用造成肝脑损伤。

Objective To investigate the relationships among the chronic alcohol exposure, stress injury of liverbrain and ceramide＇ s effect. Methods The chronic alcohol exposure models in both wide type （WT） and sphingomyelin synthetase 2 knockout （SMS2-/-） mice were established. HE staining, Masson staining, c-Fos and NF-κB immunolabeling, Western blotting analysis and trranssmission electron microscopy were carried out. Results Alcohol exposure led to the wild-type and SMS2-/- mice hepatocytic steatosis and hepatic fibrosis. Mallory bodies and inflammatory cell infiltration were found in liver. After alcohol exposure, c-Fos and NF-κB positive cells increased in hippocampal CA1 area with dose dependency （P 〈 0. 01 ）. However, comparing with the WT pups, the number of positive c-Fos and NF-κB cells in SMS2-/- mice increased much more than WT mice （P 〈 0. 05）. Immunoblotting evidence of c-Fos and NF-κB protein supported the data of immunohistochemistry in both in WT and SMS2 -/- mice （P 〈 O. O1 ）. Conclusion Oxidative stress and inflammatory responses in liver and CNS are the main pathological alterations after chronic alcohol exposure. The alcohol-inducing oxidative stress probably is the cause of insulin resistance in organism. Ceramide is involved in the processes above through c-Fos and NF-κB pathway. Alcohol exposure induces the oxidative stress injury in liver-brain axis, such as inflammatory responses and the expression of oxidative stress cues, c-Fos and NF-κB. Ceramide increases the alterations above.