Regulatory effect of traditional Chinese medicinal formula Zuo-Gui-Wan on the Th17/Treg paradigm in mice with bone loss induced by estrogen deficiency.

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE Bone loss is a common pathological condition induced by estrogen deficiency. The Th17/Treg paradigm, which can be skewed by estrogen, plays an important role in regulating bone metabolism AIM OF THE STUDY The purpose of this study was to determine the role of the Th17/Treg shift in estrogen deficiency-induced bone loss in mouse models and to elucidate the immunopharmacologic mechanism of Zuo-Gui-Wan (ZGW) in preventing bone loss in this process by regulating Th17/Treg paradigm. MATERIALS AND METHODS: Splenocytes of ovariectomized (Ovx) mice and naturally aged mice were isolated and Flow cytometry was used to detect the Th17/Treg subsets. In addition, serum estrodiol (E2) and serum C-terminal telopeptides of type Ι collagen (CTx) were detected by ELISA assay. Bone mineral density (BMD) of the left tibiae was measured by dual-energy X-ray absorptiometry. Moreover, Ovx mice were administrated with different doses of ZGW for 12 weeks, and BMD and Th17/Treg subsets were determined. Bone histomorphometry was observed by Hematoxylin and eosin (H&E) staining and serum protein levels of IL-6 were analyzed by ELISA assay. In addition, the mRNA and protein expression of RORγt and Foxp3 were detected by RT-PCR and Western blot respectively. RESULT: The Th17/Treg paradigm shifted to Th17 in estrogen-deficient mice both in the Ovx mice and the naturally aged mice. BMD and E2 levels negatively correlated with the Th17/Treg ratio. After ZGW administration, the BMD was enhanced markedly in the Ovx mice as well as in the naturally aged mice. Both the mRNA and protein expressions of IL-6 and RORγt were decreased, whereas those of Foxp3 were increased significantly after ZGW administration. CONCLUSION: Th17/Treg shift involved in the bone loss induced by estrogen deficiency. ZGW prevented bone loss efficiently and skewed Th17/Treg paradigm towards Treg without enhancing E2.