淫羊藿苷常用于股骨头坏死(ONFH)的临床治疗。miR-23a-3p在调节骨髓间充质干细胞(BMSCs)成骨分化中发挥重要作用。本研究旨在探讨淫羊藿苷和miR-23a-3p在BMSCs成骨分化中的作用及建立ONFH模型。用不同浓度的含淫羊藿苷的血清体外分离培养BMSCs,和BMSCs也用miR-23a抑制剂转染。在BMSCs中测量碱性磷酸酶(ALP)活性和细胞活力以及BMP-2/Smad5/Runx2和WNT/β-catenin途径相关的mRNA和蛋白表达。此外,双荧光素酶报告基因试验和通路抑制剂用于验证淫羊藿苷治疗/miR-23a与上述通路的关系.建立体内ONFH大鼠模型,进行28天的灌胃处理和miR-23a-3p抑制剂的慢病毒转染。然后,血清中的骨生化标志物(ELISA试剂盒),股骨头(HE染色和数字X线摄影,DR)和上述通路相关蛋白进行检测。我们的结果表明,淫羊藿苷处理/miR-23a敲低促进BMSC的活力和成骨分化,以及增加BMP-2,BMP-4,Runx2,p-Smad5,Wnt1和β-catenin在BMSCs和ONFH模型大鼠中的mRNA和蛋白表达。此外,淫羊藿苷治疗/miR-23a敲低增加骨生化标志物(ACP-5,BAP,NTXI,CTXI和OC)和改良ONFH模子年夜鼠的ONFH。此外,双荧光素酶报告基因试验证实Runx2是miR-23a-3p的直接靶标.这些数据表明淫羊藿苷通过降低miR-23a-3p水平和调节BMP-2/Smad5/Runx2和WNT/β-联蛋白途径来促进BMSC活力和成骨分化以及改善ONFH。
Icariin is commonly used for the clinical treatment of osteonecrosis of the femoral head (ONFH). miR-23a-3p plays a vital role in regulating the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). The present study aimed to investigate the roles of icariin and miR-23a-3p in the osteogenic differentiation of BMSCs and an ONFH model. BMSCs were isolated and cultured in vitro using icariin-containing serum at various concentrations, and BMSCs were also transfected with a miR-23a inhibitor. The alkaline phosphatase (ALP) activity and cell viability as well as BMP-2/Smad5/Runx2 and WNT/β-catenin pathway-related mRNA and protein expression were measured in BMSCs. Additionally, a dual-luciferase reporter assay and pathway inhibitors were used to verify the relationship of icariin treatment/miR-23a and the above pathways. An ONFH rat model was established in vivo, and a 28-day gavage treatment and lentivirus transfection of miR-23a-3p inhibitor were performed. Then, bone biochemical markers (ELISA kits) in serum, femoral head (HE staining and Digital Radiography, DR) and the above pathway-related proteins were detected. Our results revealed that icariin treatment/miR-23a knockdown promoted BMSC viability and osteogenic differentiation as well as increased the mRNA and protein expression of BMP-2, BMP-4, Runx2, p-Smad5, Wnt1 and β-catenin in BMSCs and ONFH model rats. In addition, icariin treatment/miR-23a knockdown increased bone biochemical markers (ACP-5, BAP, NTXI, CTXI and OC) and improved ONFH in ONFH model rats. In addition, a dual-luciferase reporter assay verified that Runx2 was a direct target of miR-23a-3p. These data indicated that icariin promotes BMSC viability and osteogenic differentiation as well as improves ONFH by decreasing miR-23a-3p levels and regulating the BMP-2/Smad5/Runx2 and WNT/β-catenin pathways.