Protective effect of Du-Zhong-Wan against osteoporotic fracture by targeting the osteoblastogenesis and angiogenesis couple factor SLIT3.

ETHNOPHARMACOLOGICAL RELEVANCE: Du-Zhong-Wan (DZW) is a traditional Chinese medicine (TCM) composed of Eucommia ulmoides Oliv. and Dipsacus asper Wall. ex C.B. Clarke in the ratio 1:1. Based on the TCM theory, DZW nourishes the kidney to strengthen the bones. The literature research revealed that DZW possesses anti-fatigue, anti-depressant, and anti-osteoporotic properties. However, the action and mechanism of DZW on osteoporotic fracture remains slightly unclear. AIM OF THE STUDY: To evaluate the pharmacological effect of DZW on ovariectomized mice with an open femoral fracture and reveal the underlying mechanism. MATERIALS AND METHODS: We conducted ovariectomy for 5 weeks, followed by unilateral open transverse femoral fracture for another 3 weeks in C57BL/6 mice; during this process, DZW was administrated. The femur bone and vertebra tissues were collected and analyzed by micro-computed tomography, histomorphometry, mechanical strength testing, immunohistochemistry staining, and qRT-PCR analyses. In addition, alkaline phosphatase (ALP) and Alizarin red S (ARS) staining were performed to determine the extent of osteoblastogenesis from bone marrow mesenchymal stem cells (BMSCs). Western blotting was performed to examine the protein expression. RESULTS: DZW treatment significantly improved the bone histomorphometric parameters in mice undergoing ovariectomy when combined with the femoral fracture, including an increase in the bone volume, trabecular number, and bone formation rate and a decrease in the bone erosion area. Simultaneously, DZW treatment histologically promoted fractured callus formation. Mechanical strength testing revealed significantly higher stiffness and an ultimate load after treatment with DZW. The angiogenesis of H-type vessels was enhanced by DZW, as evidenced by increased levels of CD31 and endomucin (EMCN), the H-type vessel endothelium markers, at the fractured endosteum and metaphysis regions. Relative to the osteoporotic fracture mice, the DZW treatment group showed an increased proangiogenic factor SLIT3 level. The increased level of SLIT3 was also recorded during the process of DZW-stimulated osteoblastogenesis from BMSCs. CONCLUSIONS: For the first time, we demonstrated that DZW promoted osteoporotic fracture healing by enhancing osteoblastogenesis and angiogenesis of the H-type vessels. This enhanced combination of osteoblastogenesis and angiogenesis was possibly related to the production of proangiogenic factor SLIT3 induced by DZW.

[Discussion about the divided methods and unification on the location of scalp acupuncture].

Two kinds of divided methods currently used in clinic and their unified new method on the scalp acupuncture were discussed. Through comparing with the nomination and selecting methods on the stimulating location, the characteristics of the two kinds of divided methods on the location of scalp acupuncture were pointed out respectively. The advantage of the method with scalp projection areas is that the nomination is easy to associate and remember, however its disadvantage is that the measurement on point-selecting is less reasonable. For the method with standard treatment lines, its measurement is more reasonable, but its nomination on the location is sophisticated and difficult to be associated and remembered, especially having more repeated lines. The new divided method of scalp acupuncture combines the advantages of the two methods in order to easily remember and freely apply through their unification, which is worth being popularized.