目的 研究青蒿琥酯对日本血吸虫诱导的早期肝纤维化小鼠血清和肝组织中热休克蛋白47（HSP47）表达的影响,探讨青蒿琥酯抑制日本血吸虫病肝纤维化作用的可能机制。 方法 30只ICR小鼠随机分成感染模型组、感染治疗组和健康对照组等3组,每组10只。感染组每鼠经腹部皮肤贴壁感染日本血吸虫尾蚴（20 ± 1）条,健康对照组不感染。感染后第6周,用吡喹酮300 mg/kg 2日疗法杀虫,同时感染治疗组小鼠按60 mg/kg剂量每天腹腔注射0.3 ml青蒿琥酯,连续2周;感染模型组和健康对照组腹腔注射灭菌生理盐水0.3 ml。治疗结束次日,各组小鼠用戊巴比妥钠麻醉后采血,应用ELISA检测血清HSP47和转化生长因子β1（TGF-β1）含量,取小鼠肝、脾称重,计算脏器指数。取部分肝组织,应用碱水解法检测肝羟脯氨酸含量。取部分肝组织,用10%多聚甲醛固定,行苏木素-伊红（HE）和Masson染色,观察肝组织病理学变化和胶原增生情况。取部分肝组织,应用实时荧光定量PCR检测肝脏HSP47 mRNA、Ⅰ型胶原α1链mRNA表达水平。 结果 健康对照组小鼠肝脏指数为（4.72 ± 0.52）,低于感染模型组（10.50 ± 0.57）和感染治疗组（8.31 ± 0.52）（P < 0.01）,感染治疗组小鼠肝脏系数与感染模型组差异有统计学意义（P < 0.01);健康对照组小鼠脾脏系数为（0.38 ± 0.04）,低于感染模型组（2.41 ± 0.44）和感染治疗组（2.26 ± 0.06）（P < 0.01）,感染治疗组小鼠脾脏系数与感染模型组差异无统计学意义（P > 0.05)。HE染色结果显示,感染治疗组小鼠肝脏肿大程度和表面虫卵结节较感染模型对照组轻、小。Masson染色结果显示,感染治疗组小鼠肝脏肉芽肿大小和胶原纤维面积均较感染模型组小,感染模型组胶原增生面积为（25.78 ± 2.61）%,高于感染治疗组的（6.87 ± 3.54）%和健康对照组的（1.19 ± 0.18）%（P < 0.01）。ELISA检测结果显示,感染治疗组小鼠血清HSP47、TGF-β1含量分别为（169.81 ± 20.94）pg/ml、（20.82 ± 1.90）ng/ml,均低于感染模型组[（203.14 ± 46.29）pg/ml、（27.49 ± 6.81）ng/ml]（P < 0.05）,二者HSP47、TGF-β1含量均高于健康对照组（P < 0.01）。羟脯氨酸含量和肝组织HSP47 mRNA、Ⅰ型胶原α1链mRNA表达量与血清HSP47、TGF-β1含量的趋势一致：感染治疗组小鼠肝羟脯氨酸含量为（0.27 ± 0.08）mg/g肝湿重,低于感染模型组的（0.69 ± 0.07）mg/g肝湿重（P < 0.01）,二者肝羟脯氨酸含量均高于健康对照组[（0.11 ± 0.04）mg/g肝湿重]（P < 0.05）;感染治疗组小鼠肝脏HSP47 mRNA、Ⅰ型胶原α1链mRNA表达水平分别为（0.49 ± 0.27）、（0.67 ± 0.09）,均低于感染模型组的（0.84 ± 0.17）、（0.91 ± 0.11）（P < 0.05）,二者HSP47 mRNA、Ⅰ型胶原α1链mRNA表达水平均高于健康对照组（0.24 ± 0.04、0.24 ± 0.05）（P < 0.01）。血清HSP47水平与TGF-β1水平呈正相关关系（r = 0.928 0）;血清HSP47水平、TGF-β1水平与肝组织Ⅰ型胶原α1链mRNA表达水平间均呈正相关关系（r = 0.926 2、0.872 8）。 结论 青蒿琥酯对日本血吸虫诱导的小鼠早期肝纤维化具有较好的干预效果,其机制可能系通过下调HSP47的表达水平抑制胶原合成,进而减轻肝纤维化程度。

Objective To understand the possible mechanism underlying the artesunate’s inhibition on Schistosoma japonicum-induced hepatic fibrosis, the effect of artesunate on the expression of heat shock protein 47 (HSP47) in serum and liver during the early stage of mouse hepatic fibrosis induced by S. japonicum infection was studied. Methods Total 30 female ICR mice were randomly divided into 3 groups: the un-infected control group, the infection group and the treatment group (infection + artesunate treatment), with 10 mice in each group. Each mouse in the infection and treatment groups was infected with 20 ± 1 cercariae of S. japonicum via abdomen skin penetration. All infected mice were treated with 300 mg/kg praziquantel for 2 days to eliminate the adult worms and the mice in treatment group were additionally treated with 60 mg/kg artesunate in 0.3 ml volume by intraperitoneal injection daily for 2 weeks. For the control group, mice were given the same volume of saline. After the artesunate treatment, all mice from 3 groups were sacrificed, blood samples were taken for serum separation, and the levels of HSP47 and transforming growth factor-β1(TGF-β1) in sera were detected by ELISA. The mouse livers and spleens were collected and weighed to calculate their organ index(organ weight/body weight). The livers were fixed with paraformaldehyde and sectioned for hematoxylin eosin (HE) and Masson-trichrome staining to observe the liver pathological changes and collagen proliferation. The hydroxyl-proline level in liver was measured using the alkaline hydrolysis. The mRNA expression levels of liver HSP47 and type Ⅰ collagen (COL1) were determined by real-time quantitative PCR (RT-PCR). Results The liver index in infection group (10.50 ± 0.57) was significantly higher than that in treatment group (8.31 ± 0.52) and in un-infected control group (4.72 ± 0.52) with statistical difference (P < 0.01). The spleen index was significantly higher in infection group (2.41 ± 0.44) than that in un-infected control group (0.38 ± 0.04) (P < 0.01), however it was not significantly different compared to that in treatment group(P > 0.05). Liver pathological damage was significantly reduced in treatment group compared to the infection group, including reduced liver size and egg granuloma. Masson-trichrom staining showed less liver fibrosis in treatment group than in infection group. The area of collagen proliferation in liver in treatment group was (6.87 ± 3.54)%, that is significantly smaller than that in infection group [(25.78 ± 2.61)%] (P < 0.01). The levels of HSP47 and TGF-β1 in sera of mice in the treatment group [(169.81 ± 20.94) pg/ml and (20.82 ± 1.90) ng/ml, respectively] were significantly lower than that in infection group [(203.14 ± 46.29) pg/ml and (27.49 ± 6.81) ng/ml] (P < 0.05), even though the levels of HSP47 and TGF-β1 in both infection and treatment groups were significantly higher than that in the un-infected control group (P < 0.01). The similar results were also observed in the hydroxyproline level and the expression of HSP47 and COL1 in liver tissues. The level of liver hydroxyproline in the treatment group was (0.27 ± 0.08) mg/g, which was significantly lower than that of the infection group [(0.69 ± 0.07) mg/g] (P < 0.01), however, all higher than that in un-infected control group [(0.11 ± 0.04) mg/g] (P < 0.05). The mRNA expression levels of HSP47 and COL1 in the liver of mice of treatment group were 0.49 ± 0.27 and 0.67 ± 0.09, respectively, which are significantly lower than those in the infection group (0.84 ± 0.17 and 0.91 ± 0.11, respectively) (P < 0.05). The expression levels of HSP47 and COL1 in both infection and treatment groups were significantly higher than those in the un-infected control group (0.24 ± 0.04 and 0.24 ± 0.05) (P < 0.01). The serum HSP47 level was positively correlated with the level of TGF-β1 (r = 0.928 0), and serum HSP47 level and TGF-β1 level were positively correlated with the level of hepatic COL1 mRNA expression(r = 0.926 2, 0.872 8). Conclusion Artesunate has protective effects on the early hepatic fibrosis caused by S. japonicum infection and the protection is possibly achieved by down-regulating the expression of HSP47, further suppressing the collagen synthesis so as to alleviate the hepatic fibrosis. HSP47 is expected to be one of the new prognostic markers and treatment targets in S. japonicum infection.