Shandong University of Traditional Chinese Medicine, Jinan, China.
The First Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, China.
J Ethnopharmacol. 2022 Sep 15;295:115399. doi: 10.1016/j.jep.2022.115399. Epub 2022 May 29.
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.
To evaluate the pharmacological effect of DZW on ovariectomized mice with an open femoral fracture and reveal the underlying mechanism.
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.
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.
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.
杜仲丸（DZW）是一种中药，由 1:1 的杜仲（Eucommia ulmoides Oliv.）和续断（Dipsacus asper Wall. ex C.B. Clarke）组成。根据中医理论，DZW 具有补肾壮骨的作用。文献研究表明，DZW 具有抗疲劳、抗抑郁和抗骨质疏松的特性。然而，DZW 对骨质疏松性骨折的作用和机制仍不太清楚。
评估 DZW 对去卵巢伴有开放性股骨骨折小鼠的药理作用，并揭示其潜在机制。
我们对 C57BL/6 小鼠进行了 5 周的卵巢切除术，然后再进行 3 周的单侧开放性横断股骨骨折；在此过程中，给予 DZW 治疗。通过微计算机断层扫描、组织形态计量学、力学强度测试、免疫组织化学染色和 qRT-PCR 分析，收集和分析股骨和椎骨组织。此外，进行碱性磷酸酶（ALP）和茜素红 S（ARS）染色以确定骨髓间充质干细胞（BMSCs）成骨的程度。进行 Western blot 以检测蛋白表达。
DZW 治疗显著改善了卵巢切除合并股骨骨折小鼠的骨组织形态计量学参数，包括骨体积、小梁数量和骨形成率增加，骨侵蚀面积减少。同时，DZW 治疗在组织学上促进了骨折愈合。经 DZW 治疗后，力学强度测试显示刚度和极限载荷显著增加。DZW 增强了 H 型血管的血管生成，表现为骨折端和骺端区域的 H 型血管内皮细胞标志物 CD31 和内皮质蛋白（EMCN）水平升高。与骨质疏松性骨折小鼠相比，DZW 治疗组 SLIT3 等促血管生成因子水平升高。在 DZW 刺激 BMSCs 成骨过程中也记录到 SLIT3 水平升高。
我们首次证明 DZW 通过增强成骨细胞和成骨细胞的血管生成来促进骨质疏松性骨折愈合。这种成骨细胞和成骨细胞的增强结合可能与 DZW 诱导的促血管生成因子 SLIT3 的产生有关。
