蒿鳖养阴软坚方对小鼠血吸虫病肝纤维化的疗效

目的 探讨蒿鳖养阴软坚方对小鼠血吸虫病肝纤维化的治疗作用.方法 随机取8只小鼠为正常对照组,其余小鼠每只用日本血吸虫尾蚴 (26±1) 条经腹壁皮肤感染;12周后再将感染小鼠按体质量随机区组法分为短期感染对照 (短模) 组和长期感染对照 (长模) 组、蒿鳖养阴软坚方高、中、低剂量(11.58、3.66,1.16 g/kg)组、阳性药物 (包括复方鳖甲软肝片和秋水仙碱)对照组.处死短模组小鼠,证实肝纤维化形成后给予治疗用药.蒿鳖养阴软坚方组给予蒿鳖养阴软坚方干粉状提取物混悬液灌胃,每日1次,共12周.光学显微镜观察肝组织病理学变化;盐酸水解法测定肝组织羟脯氨酸含量;放射免疫法测定血清透明质酸和Ⅲ型前胶原水平;明胶酶谱法测定肝组织基质金属蛋白酶 (MMP)-2和MMP-9水平.对正态分布计量资料多组间比较采用单因素方差分析,方差齐时用Tukey HSD检验,方差不齐时用Tamhane检验;对纤维化程度计分行非参数统计Mann-Whitney U检验;对纤维化程度分期行Ridit分析.结果 短模组和长模组小鼠肝组织均出现大量炎性细胞浸润及钙化的虫卵结节,大量胶原纤维分布于虫卵肉芽肿周围和汇管区,甚至形成假小叶;蒿鳖养阴软坚方可明显减轻血吸虫病肝纤维化小鼠肝脏的病理学改变,其各剂量均能显著降低肝纤维化分期 (u值分别为3.964、5.184、3.438,P值均<0.01) 及计分 (P值均<0.01),其中高剂量组的纤维化程度分期低于阳性对照药 (u值分别为32.204、2.451,P值均<0.05),且其计分低于秋水仙碱组 (P<0.05).短模组 (6.07±2.46) mg/g]和长模组 (7.84±4.12) mg/g]胶原蛋白含量显著高于正常对照组 (0.24±0.27) mg/g],P值均<0.01;蒿鳖养阴软坚方高剂量组 (1.88±1.62) mg/g]、中剂量组 (2.18±2.16) mg/g] 明显低于长模组 (P<0.05).短模组(1110.72±139.34) ng/ml] 和长模组 (1062.62±65.81) ng/ml] 透明质酸水平显著高于正常对照组 (647.52±360.29) ng/ml],P值均<0.05;蒿鳖养阴软坚方中剂量组 (677.04±256.93) ng/ml]、低剂量组 (733.96±273.14)ng/ml] 明显低于长模组 (P值均<0.05).短模组和长模组MMP-9水解条带的亮度分别为 (2.93±0.79)×10~5、(3.30±0.43)×10~5,显著高于正常对照组(1.18±0.30)×10~5],P<0.01;蒿鳖养阴软坚方高、中、低剂量组 (2.29±0.70)×10~5、(2.33±0.56)×10~5、(2.40±0.74)×10~5] 明显低于长模组 (P值均<0.05);短模组和长模组MMP-9水解条带的面积 (116.75±27.46,118.79±26.77) 显著高于正常对照组 (45.83±7.17),P值均<0.01.短模组和长模组MMP-2水解条带的亮度 (2.48±0.61)×10~5、(2.79±0.61)×10~5] 显著高于正常对照组 (1.17±0.31)×10~5,P值均<0.01];短模组和长模组MMP-2水解条带的面积(93.56±20.13、101.74±21.77) 显著高于正常对照组 (44.86±12.10,P值均<0.01),蒿鳖养阴软坚方高剂量组 (69.40±27.64) 明显低于长模组 (P<0.05).结论 蒿鳖养阴软坚方对小鼠血吸虫病肝纤维化有确切的治疗作用,其机制之一可能是降低肝组织MMP-2及MMP-9水平.

Therapeutic effect of Haobieyangyinruanjianfang on the mouse liver fibrosis caused by schistosomiasis

Objective To explore therapeutic effect of Haobieyangyinruanjianfang (HBYYRJ) on mouse liver fibrosis by schistosomiasis. Methods Mice except for normal control were infected with Japa-nese schistosome cercarias, after 12 weeks, infected mice were divided into 7 groups: low HBYYRJ group, middle HBYYRJ group, high HBYYRJ group, Fufangbiejiaruangan tablet (FFBJRG) group, colchicine group, 3 months infection group and 6 months infection group. Hepatic fibrosis was found in 3 months infection group. Liver hydroxyproline (Hyp) was determined, matrix metalloproteinase-2 and 9 (MMP-2, MMP-9) were detected with gelatin zymography, serum hyaluronic acid (HA) and precoilagen Ⅲ (PC-Ⅲ) were detected using RIA. Results HBYYRJ obviously reduced hepatic fibrosis (P < 0.01). Collagen and HA in 3 months infection group and 6 months infection group were higher than that in normal group (P < 0.01), collagen in high and middle HBYYRJ groups and HA in middle and low HBYYRJ groups were lower than that in 6 months infection group (P < 0.01, P < 0.05). The expression of MMP-9 and MMP-2 in 3 months infection group and 6 months infection group was higher than that in normal group(P<0.01), The expression of MMP-9 in three HBYYRJ groups and the expression of MMP-2 in high HBYYRJ group were lower than that in 6 months infection group (P < 0.05). Conclusion HBYYRJ can reduce liver fibrosis caused by schistosomiasis.