Osteoporosis (OP) can result in slower bone regeneration than the normal condition due to abnormal oxidative stress and high levels of reactive oxygen species (ROS), a condition detrimental for bone formation, making the OP‐related bone healing a significant clinical challenge. As the osteogenic differentiation ability of bone marrow mesenchymal stem cells (BMSCs) is closely related to bone regeneration; currently, this study assessed the effects of Picein on BMSCs in vitro and bone regeneration in osteoporotic bone defect in vivo. Cell viability was determined by CCK‐8 assay. The production of (ROS), malonaldehyde, superoxide dismutase activities, and glutathione was evaluated by using commercially available kits, and a flow cytometry analysis was adopted to detect macrophage polarization. Osteogenic capacity of BMSCs was evaluated by alkaline phosphatase (ALP) activity, ALP staining, and Alizarin red S staining. The expression of osteogenic‐related proteins (OPN, Runx‐2, OCN) and osteogenic‐related genes (ALP, BMP‐4, COL‐1, and Osterix) were evaluated by Western blotting and real‐time PCR (RT‐PCR). In addition, proliferation, migration ability, and angiogenic capacity of human umbilical vein endothelial cells (HUVECs) were evaluated by EdU staining, scratch test, transwell assay, and tube formation assay, respectively. Angiogenic‐related genes (VEGF, vWF, CD31) were also evaluated by RT‐PCR. Results showed that Picein alleviated erastin‐induced oxidative stress, enhanced osteogenic differentiation capacity of BMSCs, angiogenesis of HUVECs, and protects cells against ferroptosis through Nrf2/HO‐1/GPX4 axis. Moreover, Picein regulate immune microenvironment by promoting the polarization of M2 macrophages in vitro. In addition, Picein also reduce the inflammation levels and promotes bone regeneration in osteoporotic bone defect in OP rat models in vivo. Altogether, these results suggested that Picein can promote bone regeneration and alleviate oxidative stress via Nrf2/HO‐1/GPX4 pathway, offering Picein as a novel antioxidant agent for treating osteoporotic bone defect.

中文翻译：

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云杉素通过 Nrf2/HO-1/GPX4 通路抑制铁死亡，减轻骨质疏松性骨缺损的氧化应激并促进骨再生


由于异常的氧化应激和高水平的活性氧（ROS），骨质疏松症（OP）可导致骨再生速度比正常情况慢，这种情况不利于骨形成，使得OP相关的骨愈合成为重大的临床挑战。由于骨髓间充质干细胞（BMSCs）的成骨分化能力与骨再生密切相关；目前，本研究评估了Picein对体外骨髓间充质干细胞和体内骨质疏松性骨缺损骨再生的影响。通过 CCK-8 测定测定细胞活力。使用市售试剂盒评估（ROS）、丙二醛、超氧化物歧化酶活性和谷胱甘肽的产生，并采用流式细胞术分析检测巨噬细胞极化。通过碱性磷酸酶（ALP）活性、ALP染色和茜素红S染色评估BMSCs的成骨能力。通过蛋白质印迹和实时 PCR (RT-PCR) 评估成骨相关蛋白（OPN、Runx-2、OCN）和成骨相关基因（ALP、BMP-4、COL-1 和 Osterix）的表达。此外，分别通过EdU染色、划痕试验、Transwell实验和成管实验评估人脐静脉内皮细胞（HUVEC）的增殖能力、迁移能力和血管生成能力。还通过 RT-PCR 评估了血管生成相关基因（VEGF、vWF、CD31）。结果表明，Picein 可减轻erastin 诱导的氧化应激，增强 BMSC 的成骨分化能力、HUVEC 的血管生成，并通过 Nrf2/HO-1/GPX4 轴保护细胞免于铁死亡。此外，Picein通过促进体外M2巨噬细胞的极化来调节免疫微环境。 此外，Picein还能降低体内OP大鼠模型骨质疏松性骨缺损的炎症水平并促进骨再生。总之，这些结果表明Picein可以通过Nrf2/HO-1/GPX4途径促进骨再生并减轻氧化应激，为Picein提供作为治疗骨质疏松性骨缺损的新型抗氧化剂。