Department of Infections，Beijing Hospital of Traditional Chinese Medicine, Affiliated to the Capital Medical University, No. 23, Back Road of the Art Gallery, Dongcheng District, Beijing 100010, China.
Infections Department, Beijing Hospital of Traditional Chinese Medicine Shunyi Branch, Beijing 101300, China.
J Tradit Chin Med. 2019 Jun;39(3):315-323.
To investigate the therapeutic mechanism of compound Yindan decoction (CYD) in a rat model of acute intrahepatic cholestatic (AIC).
A total of 108 adult male rats were randomly divided into control (n = 18) and AIC groups (n = 90). AIC was induced in rats using alpha-naphthylisothiocyanate (ANIT) (75 mg/kg, 10 mL/kg in corn oil, p. o. ). Then, 90 AIC rats were randomly divided into five groups: a control group (n = 18), a CYD high dose group (n = 18), a CYD middle dose group (n = 18), a CYD low dose group (n = 18), and a ursodeoxycholic acid (UDCA) group (n = 18). According to sampling time, each group was subdivided into three subgroups: 24 h (n = 6), 48 h (n = 6), and 72 h groups (n = 6). The CYD-high, -middle and -low groups were orally administered 24.48, 12.24, and 6.12 g·kg－1·d－1 modified CYD, respectively, while the model group was given 20 mL/kg of body weight of distilled water once a day. The UDCA group was given 67. 5 mg·kg － 1·d － 1 UDCA once a day. Radioimmunity assay was used to detect the activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), gamma-glutamyl transpeptidase (GGT) and the levels of total bilirubin (TBil) and indirect biliruin (DBil) in rats. Reverse transcription quantitative polymerase chain reaction (qRT-PCR), Western blot analysis, and immunohistochemistry were used to detect multidrug resistance-associated protein 2 (MRP2) expression. In vitro, HepG2 hepatocellular carcinoma cells were treated with CYD medicated serum at a concentration of 15 mol/L. MRP2 and retinoid X receptor alpha (RXRα) expression was analyzed by qRT-PCR and Western blotting.
Serum levels of ALT, AST, GGT, ALP, TBil, and DBil were significantly reduced in the CYD and positive drug groups compared with the control group (P < 0. 05 and P < 0.01, respectively). Pathological changes in rat liver tissues at 72 h in the CYD-high and -medium dose groups and positive drug group were not significant compared with the control group. CYD and UDCA treatment ameliorated ANIT-induced biliary epithelial cell proliferation. Neutrophil infiltration was rare and little focal necrosis was observed in lobules in the CYD-high and -medium dose groups and UDCA group at 72 h. Compared with the control group, the expression of MRP2 mRNA and MRP2 protein in the liver tissue of the CYD groups was significantly increased (P < 0. 05 and P < 0. 01, respectively). MRP2 expression and RXRα nuclear receptor mRNA and protein levels in the CYD groups were significantly increased compared with the control and UDCA groups (P < 0. 01).
CYD can alleviate cholestasis in ANIT-induced AIC rats, and the mechanism underlying this action might involve increases in ALT, AST, GGT, ALP, TBil, and DBil and upregulation of MRP2 and RXRα mRNA and protein levels.
研究复方茵丹汤（CYD）在急性肝内胆汁淤积（AIC）大鼠模型中的治疗机制。
将 108 只成年雄性大鼠随机分为对照组（n = 18）和 AIC 组（n = 90）。采用α-萘基异硫氰酸酯（ANIT）（75mg/kg，10mL/kg玉米油，po）诱导大鼠 AIC。然后，将 90 只 AIC 大鼠随机分为五组：对照组（n = 18）、CYD 高剂量组（n = 18）、CYD 中剂量组（n = 18）、CYD 低剂量组（n = 18）和熊去氧胆酸（UDCA）组（n = 18）。根据采样时间，每组分为 24 h（n = 6）、48 h（n = 6）和 72 h 组（n = 6）。CYD 高、中、低剂量组分别口服 24.48、12.24 和 6.12 g·kg－1·d－1 改良 CYD，模型组给予 20mL/kg 体重蒸馏水，每天 1 次。UDCA 组每天给予 67.5mg·kg－1·d－1 UDCA。放射免疫法检测大鼠丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）、碱性磷酸酶（ALP）、γ-谷氨酰转肽酶（GGT）和总胆红素（TBil）、间接胆红素（DBil）水平。逆转录定量聚合酶链反应（qRT-PCR）、Western blot 分析和免疫组化检测多药耐药相关蛋白 2（MRP2）表达。体外，用浓度为 15mol/L 的 CYD 含药血清处理 HepG2 肝癌细胞，qRT-PCR 和 Western blot 分析 MRP2 和视黄醇 X 受体α（RXRα）表达。
与对照组相比，CYD 和阳性药物组大鼠血清 ALT、AST、GGT、ALP、TBil 和 DBil 水平明显降低（P<0.05 和 P<0.01）。72 h 时，CYD 高、中剂量组和阳性药物组大鼠肝组织病理变化与对照组相比无明显差异。与对照组相比，CYD 和 UDCA 治疗可改善 ANIT 诱导的胆管上皮细胞增殖。72 h 时，CYD 高、中剂量组和 UDCA 组肝组织中中性粒细胞浸润较少，小叶内偶见局灶性坏死。与对照组相比，CYD 组肝组织中 MRP2mRNA 和 MRP2 蛋白表达明显增加（P<0.05 和 P<0.01）。与对照组和 UDCA 组相比，CYD 组 MRP2 表达和 RXRα 核受体 mRNA 和蛋白水平明显增加（P<0.01）。
CYD 可减轻 ANIT 诱导的 AIC 大鼠胆汁淤积，其作用机制可能与 ALT、AST、GGT、ALP、TBil 和 DBil 升高以及 MRP2 和 RXRα mRNA 和蛋白水平上调有关。
