Department of Periodontology, School and Hospital of Stomatology, Shandong University & Shandong Key Laboratory of Oral Tissue Regeneration & Shandong Engineering Laboratory for Dental Materials and Oral Tissue Regeneration, Cheeloo College of Medicine, No .44-1 Wenhua Road West, Jinan, 250012, Shandong, China.
Department of Endodontics, Jinan Stomatological Hospital, No. 101 Jingliu Road, Jinan, 250001, Shandong, China.
J Mol Histol. 2021 Dec;52(6):1245-1255. doi: 10.1007/s10735-020-09941-1. Epub 2021 Feb 10.
After periodontal treatment, the local inflammatory environment surrounding periodontal tissues cannot be entirely eliminated. The means by which alveolar bone repair and regeneration are promoted in inflammatory environments have important clinical significance. As a powerful protein that promotes the differentiation of osteocytes, semaphorin 3A (Sema3A) shows potential for bone regeneration therapy. However, the effect of Sema3A on osteogenic differentiation in an inflammatory environment, as well as the underlying mechanism, have not yet been explored. We used lentivirus to transduce rat bone marrow-derived mesenchymal stem cells (rBMSCs) to stably overexpress Sema3A. Lipopolysaccharide from Escherichia coli (E. coli LPS) was used to stimulate rBMSCs to establish an inflammatory environment. ALP staining, Alizarin red staining, ALP activity tests, quantitative RT-PCR (qRT-PCR), and Western blotting were used to elucidate the effect of Sema3A on the osteogenesis of rBMSCs in inflammatory environments. XAV939 and LiCl were used to determine whether the Wnt/β-catenin signaling pathway was involved in attenuating the inhibition of Sema3A-induced osteogenic differentiation by LPS. The qRT-PCR and Western blot results demonstrated that the lentiviral vector (LV-NC) and lentiviral-Sema3A (LV-Sema3A) were successfully transduced into rBMSCs. An inflammatory environment could be established by stimulating rBMSCs with 1 μg/ml E. coli LPS. After Sema3A overexpression, mineral deposition was exacerbated, and the BSP and Runx2 gene and protein expression levels were increased. Furthermore, E. coli LPS activated the Wnt/β-catenin signaling pathway and decreased rBMSC osteogenesis, but these effects were attenuated by Sema3A. In conclusion, Sema3A could protect BMSCs from LPS-mediated inhibition of osteogenic differentiation in inflammatory environments by suppressing the Wnt/β-catenin pathway.
牙周治疗后，牙周组织周围的局部炎症环境不能完全消除。在炎症环境中促进牙槽骨修复和再生的方法具有重要的临床意义。作为一种促进成骨细胞分化的强大蛋白，神经信号素 3A（Sema3A）显示出了骨再生治疗的潜力。然而，Sema3A 对炎症环境中成骨分化的影响及其潜在机制尚未得到探索。我们使用慢病毒转导大鼠骨髓间充质干细胞（rBMSCs），以稳定过表达 Sema3A。使用大肠杆菌脂多糖（E. coli LPS）刺激 rBMSCs 以建立炎症环境。碱性磷酸酶（ALP）染色、茜素红染色、ALP 活性试验、定量 RT-PCR（qRT-PCR）和 Western blot 用于阐明 Sema3A 对炎症环境中 rBMSCs 成骨的影响。XAV939 和 LiCl 用于确定 Wnt/β-catenin 信号通路是否参与减弱 LPS 对 Sema3A 诱导的成骨分化的抑制作用。qRT-PCR 和 Western blot 结果表明，慢病毒载体（LV-NC）和慢病毒-Sema3A（LV-Sema3A）成功转导到 rBMSCs 中。通过用 1μg/ml 的大肠杆菌 LPS 刺激 rBMSCs 可以建立炎症环境。过表达 Sema3A 后，矿化沉积加剧，BSP 和 Runx2 基因和蛋白表达水平增加。此外，E. coli LPS 激活了 Wnt/β-catenin 信号通路并降低了 rBMSC 成骨能力，但这些作用被 Sema3A 减弱。总之，Sema3A 可以通过抑制 Wnt/β-catenin 通路来保护 BMSCs 免受 LPS 介导的炎症环境中成骨分化的抑制。
