Immunomodulatory mechanism of Bushen Huoxue Recipe alleviates cyclophosphamide-induced diminished ovarian reserve in mouse model.

ETHNOPHARMACOLOGICAL RELEVANCE

Bushen Huoxue recipe (BHR) is a Chinese herbal prescription composed of ten herbs and it is widely used for the treatment of diminished ovarian reserve (DOR). This study investigates the potentially beneficial effects and underlying mechanism of BHR on a cyclophosphamide (CTX) induced model of DOR in mice.

MATERIALS AND METHODS

Granules of BHR were first subjected to high performance liquid chromatography (HPLC) to determine the exact ingredients within the mixture. We then induced DOR in mice by injecting them with 90mg/kg of CTX. Following the single intraperitoneal injection, mice then received either saline or BHR for 21 days. To assess the effects of BHR on DOR, we examined splenic and ovarian morphology, estrous cycle duration, ovarian index, follicle number, body weight, and concentration of serum E and FSH. To explore the immunological mechanism behind the effects, mouse splenocytes were isolated in order to analyze the proportion of CD4, CD8 T cells and Th1, Th17 and Treg subsets by flow cytometry. The serum levels of IFN-γ, TNF-α, IL-4, IL-17A, IL-6 and IL-10 were detected using Cytometric Bead Array (CBA). Additionally, the mRNA expression levels of T-bet, RORγt and Foxp3 were measured with quantitative real-time PCR.

RESULTS

Our results show that following treatment with BHR in DOR mice, several measures showed significant improvement. The morphology of the ovary and spleen, estrous cycle duration, body weight, ovarian index, and serum levels of E and FSH recovered to approximately normal levels and the loss of follicles at all stages was significantly attenuated. Furthermore, the elevated proportions of CD4 T cells, Th1, Th17, Treg subsets and the increased serum levels of IFN-γ, TNF-α, IL-17A, IL-6 and IL-10 as well as the mRNA expressions of T-bet, RORγt and Foxp3 in DOR mice were significantly decreased.

CONCLUSIONS

Our results show that BHR is a promising candidate to treat DOR mice and this beneficial effect may be mediated through the downregulation of augmented autoimmunity.