新强谷银(N-QGY),在QGY公式的基础上添加沙棘,是在中国临床上广泛用于治疗骨质疏松症(OP)的草药配方,但其机制值得进一步探索。从破骨细胞-成骨细胞平衡的角度探讨QGY和N-QGY治疗OP的机制。30只Sprague-Dawley大鼠随机分为N-QGY组,QGY集团,和对照组。口服生理盐水的失控大鼠,其他大鼠每天两次口服等距N-QGY或QGY,持续3天。制备含药血清和对照血清,并通过二辛可宁酸测定法(BCA)确定其对破骨细胞来源的外泌体分泌的影响,纳米粒子跟踪分析,和Westernblot。采用GW4869和白细胞介素-1β(IL-1β)作为外泌体抑制剂和诱导剂,分别。外泌体摄取,细胞计数试剂盒-8,碱性磷酸酶(ALP)染色,茜素红染色,酶联免疫吸附测定,定量实时聚合酶链反应,进行和Western印迹以检查破骨细胞外泌体含量改变对间充质干细胞(MSC)成骨分化的影响。N-QGY,QGY,和GW4869抑制骨髓间充质干细胞的破骨细胞来源的外泌体分泌和外泌体摄取,而IL-1β发挥相反的作用(p<0.05)。与IL-1β不同,N-QGY,QGY,GW4869部分提高了MSC的活力,骨钙蛋白分泌,ALP,RUNX家族转录因子2(RUNX2)和骨桥蛋白(OPN)表达,和钙沉积在破骨细胞-MSCs共培养系统中(p<0.05)。机械上,破骨细胞增加了Notum蛋白水平,但降低了β-catenin水平,它被IL-1β增强,但被GW4869、QGY逆转,和N-QGY(p<0.05)。N-QGY的作用明显优于QGY(P<0.05)。含N-QGY的血清抑制破骨细胞外泌体水平,从而通过抑制Notum蛋白和促进β-catenin蛋白促进MSCs的成骨分化。
New-QiangGuYin (N-QGY), the addition of sea buckthorn on the basis of QGY formula, is herbal formula widely used clinically in China for the treatment of osteoporosis (OP), but its mechanism warrants further exploration. The mechanisms of QGY and N-QGY in the treatment of OP are probed from the perspective of
osteoclast-osteoblast balance. Thirty Sprague-Dawley rats are randomly divided into N-QGY group, QGY group, and Control group. Beyond control rats that orally took normal saline, other rats are orally administered with isometric N-QGY or QGY twice every day for 3 days. The drug-containing serum and control serum are prepared and their effects on
osteoclast-derived exosome secretion are determined by bicinchoninic acid assay (BCA), nanoparticle tracking analysis, and Western blot. GW4869 and Interleukin-1β (IL-1β) are adopted as the exosome inhibitor and inducer, respectively. Exosome uptake, cell counting kit-8, alkaline phosphatase (ALP) staining, alizarin red staining, enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, and Western blot are performed to examine the effects of altered
osteoclast exosome content on osteogenic differentiation of mesenchymal stem cells (MSCs). N-QGY, QGY, and GW4869 inhibit
osteoclast-derived exosome secretion and exosome uptake by MSCs, whereas IL-1β exerted the opposite effects (p < 0.05). Different from IL-1β, N-QGY, QGY, and GW4869 partially elevated MSC viability, osteocalcin secretion, ALP, RUNX Family Transcription Factor 2 (RUNX2) and Osteopontin (OPN) expressions, and calcium deposition in the
osteoclast-MSCs coculture system (p < 0.05). Mechanically, osteoclasts increased Notum protein level but decreased β-catenin level, which is enhanced by IL-1β but is reversed by GW4869, QGY, and N-QGY (p < 0.05). And the effect of N-QGY is more conspicuous than that of QGY (P<0.05). N-QGY-containing serum inhibits exosome levels in osteoclasts, thereby enhancing osteogenic differentiation of MSCs via inhibition of Notum protein and promotion of β-catenin protein.