不同治法方药对大鼠脂肪肝Kupffer细胞ERK1/2蛋白活性的影响

Effects of different therapeutic methods and typical recipes on activation of ERK1/2 in Kupffer cells of rats with fatty liver

目的:观察疏肝、健脾、活血、祛湿、综合不同治法方药对大鼠脂肪肝Kupffer细胞ERK1/2蛋白活性的影响。方法:利用高脂饮食、白酒灌胃复制大鼠脂肪肝实验动物模型,同时给予疏肝、健脾、活血、祛湿、综合不同方药进行干预。12周后以Ⅳ型胶原酶、蛋白酶E联合灌注消化、梯度离心、选择性贴壁分离不同组别大鼠Kupffer细胞,采用Western blotting方法检测不同组别大鼠Kupffer细胞ERK1/2蛋白的表达及其磷酸化水平的变化。结果:模型组大鼠Kupffer细胞ERK1/2蛋白的表达和磷酸化ERK1/2蛋白的表达较正常组明显增加(P<0.01),各干预组ERK1/2表达和磷酸化ERK1/2蛋白的表达较模型组显著降低(P<0.01),其中疏肝组(柴胡疏肝散:柴胡、川芎、枳壳、陈皮、白芍、香附、炙甘草)、健脾组(参苓白术散:人参、白术、茯苓、薏苡仁、砂仁、山药、桔梗、白扁豆、莲子、炙甘草)ERK1/2蛋白表达和磷酸化ERK1/2蛋白的表达下降最为明显。结论:在大鼠脂肪肝的形成过程中Kupffer细胞ERK1/2蛋白高表达和磷酸化ERK1/2蛋白高表达可能起到了重要作用,Kupffer细胞ERK1/2蛋白的高表达和磷酸化ERK1/2蛋白高表达可能参与并促进了脂肪性肝损伤,抑制ERK1/2蛋白的表达及磷酸化可能是健脾疏肝等治法方药抗实验性大鼠脂肪肝的作用机制之一。

AIM： To investigate effects of different therapeutic methods and typical recipes on activation of ERK1/2 in Kupffer cells of rats with fatty liver. METHODS： The rat model of fatty liver was established by feeding high fat diet combinated with distillate spirit. Meanwhile Chinese medicines Shugan fang, Jianpi fang, Huoxue fang, Qushi fang, and Zonghe fang were given to treat different groups respectively. 12 weeks later,the Kupffer Cells were isolated from livers of control group, model group and different treatment groups by sequential in situ perfusion with collagenaseⅣ and pronase E, density gradient centrifugation,selective adherence. The expression of total ERK1/2 and phospho -ERK1/2 in Kupffer cells of control group, model group and different treatment groups were detected by Western blotting. RESULTS： The expressions of total ERK1/2 and phospho - ERK1/2 were higher in Kupffer cells from model group than those in control group （ P 〈 0. 01 ）. Phosphorylation of ERK1/2 was 12.4 - fold greater in model group than that in control group（ P 〈 0. 01 ）. The total ERK1/2 was 11.5 -fold greater in model group than that in control group（P 〈0. 01 ）. The expressions of total ERK1/2 and phospho -ERK1/2 were reduced in all treatment groups than that in model group（ P 〈 0. 01 ）, especially in Shugan fang group（ the ingredients of prescription： Bupleurum Chinese, Ligusticum chuanxiong HORT, Citrus aurantium, Citrus reticulata peel, Paeonia lactiflora PALL, Cyperus rotundus L, prepared Glycyrrhiza uralensis Fisch）and Jianpi fang treatment group（ the ingredients of prescription： Panax ginseng, Atractylodes, Poria cocos Wolf, Coix lacryma- jobi, Amomum villosum Lour, Dioscorea opposita Thunb, Platycodon grandiflorum A. DC, Dolichos lablab, Nelumbo nucifera Gaertn, prepared Glycyrrhiza uralensis Fisch）. CONCLUSION： The high expressions of ERK1/2 and phospho- ERK1/2 may be involved in fatty liver and contribute to liver injure. The recipes of Jianpi fang, Shugan fang, Qushi fang, Huoxue fang,and Zonghe fang may protect against liver injury by reducing the total ERK1/2 and activation of ERK1/2 in rats with fatty liver, especially the recipes of Jianpi fang and Shugan fang.