ABSTRACT

Neohesperidin (NH) was reported to regulate osteoclastic differentiation, while LncRNA SNHG1 could inhibit osteogenic differentiation of bone marrow stromal cells (BMSCs). In this study, we aimed to explore whether SNHG1-mediated osteogenic differentiation could be regulated by NH. Osteonecrosis and adjacent tissues, as well as normal bone marrow samples were gathered. BMSCs were isolated from normal bone marrow samples by Ficoll density gradient and identified by flow cytometry. Histopathological changes of tissues were detected by hematoxylin-eosin staining. After the treatment with NH or transfection, cell viability, osteogenic differentiation, and the activity of alkaline phosphatase (ALP) in BMSCs were detected by MTT, alizarin red staining, and microplate method, respectively. The histone modification and expressions of SNHG1 and osteogenic marker genes in tissues or BMSCs were detected by q-PCR and Chromatin Immunoprecipitation (ChIp). SNHG1 was highly expressed in osteonecrosis tissues, and typical signs of empty lacunae appeared in the necrotic tissues zone. NH increased viability and osteogenic differentiation of BMSCs, activity of ALP, and expressions of RUNX2, OCN and ALP. NH decreased both SNHG1 expression and H3K4me3 (activating histone modification) occupancies and increased H3K27me3 (inhibiting histone modification) occupancies of SNHG1. Furthermore, siSNHG1 enhanced osteogenic differentiation of BMSCs and expressions of RUNX2, OCN and ALP, while SNHG1 overexpression did the opposite and reversed the effects of NH on the osteogenic differentiation of BMSCs. In a word, NH promotes the osteogenic differentiation of human BMSCs by inhibiting the histone modifications of lncRNA SNHG1.

摘要

据报道，新橙皮苷 (NH) 可调节破骨细胞分化，而 LncRNA SNHG1 可抑制骨髓基质细胞 (BMSCs) 的成骨分化。在本研究中，我们旨在探讨 SNHG1 介导的成骨分化是否受 NH 调节。收集了骨坏死和邻近组织，以及正常的骨髓样本。通过 Ficoll 密度梯度从正常骨髓样本中分离出 BMSCs，并通过流式细胞术进行鉴定。苏木精-伊红染色检测组织病理学变化。分别采用MTT、茜素红染色和微孔板法检测经NH或转染处理后的BMSCs细胞活力、成骨分化和碱性磷酸酶（ALP）活性。通过q-PCR和染色质免疫沉淀（ChIp）检测组织或骨髓间充质干细胞中组蛋白修饰和SNHG1和成骨标记基因的表达。SNHG1在骨坏死组织中高表达，坏死组织区出现典型的空洞征象。NH增加BMSCs的活力和成骨分化、ALP的活性以及RUNX2、OCN和ALP的表达。NH 降低了 SNHG1 的表达和 H3K4me3（激活组蛋白修饰）的占有率，并增加了 SN​​HG1 的 H3K27me3（抑制组蛋白修饰）的占有率。此外，siSNHG1 增强了 BMSCs 的成骨分化和 RUNX2、OCN 和 ALP 的表达，而 SNHG1 的过表达则相反，并逆转了 NH 对 BMSCs 成骨分化的影响。一句话，