葛根素对胎鼠骺板Ⅰ型胶原蛋白表达的影响

Effect of puerarin on typeⅠcollagen expression in epiphyseal plates in fetal mice

取怀孕16d昆明种小鼠的胎鼠，以胚胎长骨为靶器官，设17β-雌二醇1umol／L阳性对照组、空白对照组及葛根素1，10，100umol／L3个剂量组。剥离出胎鼠尺骨，置于BGJb培养基中，葛根素和雌二醇作用48h后，将长骨固定，脱钙后作石蜡切片，进行免疫组织化学染色，光镜下观察骺板静止区、增殖区和肥大区阳性细胞定位，图像分析系统下测定免疫反应阳性细胞面积。每组培养长骨8根，重复3次。葛根素10，100umol／L组胎鼠骺板各区Ⅰ型胶原蛋白免疫组织化学染色阳性细胞面积显著大于对照组（P〈0．01）。葛根素1，10umol／L组各区Ⅰ型胶原蛋白免疫组织化学染色阳性细胞面积均低于17β雌二醇组（P〈0．05～0．01）；葛根素100umol／L组阳性细胞面积与17β-雌二醇组差异无显著性意义（P〉0．05）。随葛根素剂量升高，各区阳性细胞面积基本呈逐渐增加的趋势。在体外培养条件下，葛根素可上调胎鼠骺板静止区、增殖区和肥大区Ⅰ型胶原蛋白表达水平，并呈剂量依赖性。

Regarding long bone of 16-day Kunming mouse fetuses as target organ, and 1 u mol/L 17 β -estradiol as positive control group, at the same time established blank control group, 1, 10, 100 u mol/L puerarin groups. The ulnas of fetal mice were cultured for 48 hours in BGJb medium treated respectively with estradiol and purearin. After fixing long bones and decalcification, paraffin sections were made, and then performed immunohistochemical staining for determining the level of type Ⅰ collagen protein in growth plates. Positive cell locations in the resting zone, proliferative zone and hypertrophic zone of epiphyseal plate were observed under light microscope. Positive cell area was determined with image analysis system. Eight long bones were cultured in each group and dulplicated 3 times. The areas of type I collagen protein immunohistochemical staining-positive cells in every zone in the 10, 100 u mol/L puerarin groups were significantly bigger than that in the control group （P 〈 0.01）. The areas of positive cells in every zone in the 1, 10 u mol/L puerarin groups were smaller than that in the 17 β -estradiol group （P 〈 0.05-0.01）; there were no significant differences between 100 u mol/L puerarin group and 17 β -estradiol group （P 〉 0.05）. The areas of positive cells in every zone increased gradually with concentration of puerarin in medium increasing. Under in vitro culture condition, puerarin can upregulate fetal mouse level of type I collagen in resting zone, proliferative zone and hypertrophic zone of epiphyseal plate in a dose-dependent manner.