壮骨止痛胶囊对去卵巢骨质疏松大鼠骨组织Treg/Th17平衡的调节作用

Regulatory effect of Zhuanggu Zhitong Capsule on Treg/Th17 balance in rats with osteoporosis induced by ovariectomy

目的从骨组织调节性T细胞/辅助性T细胞(Treg/Th17)平衡角度探讨壮骨止痛胶囊抗绝经后骨质疏松作用机理。方法 72只8月龄SD雌鼠随机分为假手术组,模型组,戊酸雌二醇组(0.21 mg/kg),壮骨止痛胶囊高剂量组(1.28 g/kg)、中剂量组(0.64 g/kg)和低剂量组(0.32 g/kg),每组12只。除假手术组外,其余5组去卵巢复制绝经后骨质疏松动物模型。术后1周开始给药,连续给药18周,最后一次给药1 h后处死,快速取左股骨检测雌二醇(E2)、白细胞介素17A(IL-17A)、白细胞介素10(IL-10),取右股骨RT-PCR检测维甲酸相关孤体受体γt(RORγt)、叉状头转录因子(Foxp3)、IL-17A、IL-10 mRNA的表达,取左胫骨病理切片免疫组化染色检测表达RORγt、Foxp3的细胞百分数。结果壮骨止痛胶囊能显著升高去卵巢雌鼠骨组织E2、IL-10水平(P<0.01),降低骨组织IL-17A水平(P<0.05);显著增强骨组织Foxp3、IL-10 mRNA的表达(P<0.05),显著抑制骨组织RORγt、IL-17A mRNA的表达(P<0.01或P<0.05);显著增加骨组织内表达Foxp3的细胞百分数(P<0.05),下调表达RORγt的细胞百分数(P<0.05)。Foxp3阳性细胞百分数与RORγt阳性细胞百分数比值与E2水平呈正相关(r=0.990 5,P<0.01)。结论壮骨止痛胶囊通过抑制去卵巢雌鼠骨组织Th17细胞数量和功能活动,增加骨组织Treg细胞数量并增强其活动而调节骨组织Treg/Th17平衡,该调节作用可能部分通过提高去卵巢雌鼠体内E2水平而实现。

Objective To investigate the anti-postmenopausal osteoporosis mechanism of Zhuanggu Zhi- tong Capsule （Bone-strengthening Pain-relieving Capsule, ZGZTC ） in regulating Treg/Thl7 balance in bone tissue. Methods Seventy-two 8-month-old SD female rats were allocated randomly into sham oper- ation group, model group, estradiol group （0.21 mg/kg）, ZGZTC high-dose group （1.28 g/kg）, ZGZ- TC medium-dose group （0.64 g/kg） and ZGZTC low-dose group （0.32 g/kg）. There were 12 rats in each group. Bilateral ovariectomy was performed on rats in all groups except for the sham operation group. After administration for 18 weeks, all rats were anaesthetized intraperitoneally with hydrazine hy-drate. The content of E2, IL-10 and IL-17A was measured with ELISA in left femur. Gene expression of Foxp3, RORγt, IL-17A and IL-10 in right femur was detected by using RT-PCR, and the number of cell with RORγt, or Foxp3 was counted with immunohistochemistry in left tibia. Results Zhuanggu Zhitong Capsule significantly increased E2 and IL-10 levels in bone tissue （P 〈 0.01 ） , reduced the level of IL- 17A in bone tissue （P 〈0.05 or P 〈0.01 ）. It significantly increased the Foxp3 and IL-10 mRNA ex- pression and inhibited the RORγt mRNA expression in bone tissue （P 〈 0.05）. The percentage of cell with Foxp3 was significantly increased by ZGZTC in bone tissue （P 〈 0.05 ）, and the percentage of cell with RORγt was significantly reduced （P 〈 0.05 or P 〈 0.01 ）. The ratio of Foxp3 to RORγt presented a positive correlation with the E2 level （ r = 0. 990 5, P 〈 0.01 ）. Conclusion Zhuanggu Zhitong Capsule could downregulate the number and function of Thl7, upregulate the number and function of Treg in bone tissue in ovariectomized rats partly via improving the E2 level.