CCl4诱导的肝纤维化大鼠模型肝小叶内卵圆细胞总体积与轮廓数密度变化的体视学分析

Changes in the total volume and contour density of oval cells in hepatic lobules of rats with carbon tetrachloride-induced hepatic fibrosis:A stereological study

目的 定量研究CCl4诱导的肝纤维化大鼠肝小叶内卵圆细胞(HOC)总体积和轮廓数密度的变化。方法 将11只雄性健康SD大鼠随机分为对照组(n=5)、肝纤维化组(n=6),每周2次皮下注射CCl4与橄榄油混悬液,每次3 mL/kg。在肝纤维化造模5周后取材,从每只大鼠肝脏随机抽选5个大小约1 mm3的肝组织块分别制作1张Epon812环氧树脂包埋超薄切片,运用体视学方法结合透射电子显微镜技术,对大鼠肝小叶内的HOC总体积和轮廓数密度进行定量研究,另从每只大鼠剩余肝脏等距随机抽选4个2 mm厚的肝组织块并分别制作2张石蜡包埋的Masson染色切片,按照肝纤维化Metavir分期标准定性评估每只大鼠的肝纤维化程度。计量资料两组间比较采用成组t检验。结果 体视学定量研究显示,对照组肝小叶内HOC总体积为(15.40±7.63) mm3,肝纤维化组肝小叶内HOC总体积为(146.80±114.00) mm3,与对照组比较,肝纤维化组大鼠肝小叶内HOC总体积显著增加了8.53倍(t=-2.551,P=0.031);对照组肝小叶内HOC轮廓数密度为(56.20±40.40),肝纤维化组肝小叶内HOC轮廓数密度为(566.50±317.00),与对照组比较,肝纤维化组大鼠肝小叶内轮廓数密度显著增加了9.08倍(t=-3.539,P=0.006);定性观察研究结果显示,肝纤维化大鼠肝纤维化分期按照Metavir评分标准达到Ⅱ~Ⅲ期,大鼠窦周隙内肝星状细胞增生部位周围伴随着HOC的大量增生。结论 CCl4诱导肝纤维化大鼠肝小叶内HOC显著增生。

Objective To quantitatively investigate the changes in the total volume and contour density of hepatic oval cells(HOC)in hepatic lobules of rats with carbon tetrachloride(CCl4)-induced hepatic fibrosis.Methods A total of 11 healthy male Sprague-Dawley rats were randomly divided into control group with 5 rats and hepatic fibrosis group with 6 rats,and CCl4 and olive oil suspension were injected subcutaneously twice a week,3 mL/kg each time.After five weeks of hepatic fibrosis modeling,five liver tissue blocks with a size of about 1 mm3 were randomly selected from the liver of each rat to prepare one Epon812 epoxy resinembedded ultrathin section,and the stereological method and transmission electron microscopy were used for the quantitative analysis of the total volume and contour density of HOC in the hepatic lobules of rats. In addition, four liver tissue blocks with a thickness of 2 mm were randomly selected from the remaining liver of each rat to prepare two paraffin-embedded Masson staining sections, and the degree of liver fibrosis in each rat was qualitatively evaluated according to the Metavir staging criteria for liver fibrosis. The independent-samples t test was used for comparison of continuous data between groups. Results The quantitative stereological analysis showed that the total volume of HOC in hepatic lobules was 15.40±7.63 mm3 in the control group and 146.80± 114.00 mm3 in the liver fibrosis group, and compared with the control group, the total volume of HOC in hepatic lobules of rats in the liver fibrosis group was significantly increased by 8.53 times (t=−2.551, P=0.031);the contour density of HOC in hepatic lobules was 56.20±40.40 in the control group and 566.50±317.00 in the liver fibrosis group, and compared with the control group, the contour density of HOC in hepatic lobules of rats in the liver fibrosis group was significantly increased by 9.08 times (t=−3.539, P=0.006). Qualitative observation showed that liver fibrosis stage of rats reached stage Ⅱ- Ⅲ according to the Metavir scoring criteria, and massive proliferation of HOC was observed around the proliferation site of hepatic stellate cells in the perisinusoidal space of rats. Conclusion CCl4 induces significant proliferation of HOC in hepatic lobules of rats with liver fibrosis.