Picein alleviates oxidative stress and promotes bone regeneration in osteoporotic bone defect by inhibiting ferroptosis via Nrf2/HO‐1/GPX4 pathway

Abstract 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.