BACH1 的缺失通过恢复自噬来改善糖皮质激素诱导的 hBMSCs 中的成骨分化。
Loss of BACH1 improves osteogenic differentiation in glucocorticoid-induced hBMSCs through restoring autophagy.
机构信息
Department of Endocrinology, The Affiliated Nanhua Hospital, Hengyang Medical School, University of South China, NO. 336, Dongfeng South Road, Zhuhui District, Hengyang, Hunan Province, 421002, China.
Department of Nutrition, The Second Affiliated Hospital, Hengyang Medical School, University of South China, Hengyang, China.
出版信息
BMC Musculoskelet Disord. 2024 Aug 24;25(1):665. doi: 10.1186/s12891-024-07761-y.
BACKGROUND
Glucocorticoid-induced osteoporosis (GIOP) is the most common type of secondary osteoporosis. Recently, autophagy has been found to be related with the development of various diseases, including osteoporosis and osteoblast differentiation regulations. BTB and CNC homology 1 (BACH1) was a previously confirmed regulator for osteoblast differentiation, but whether it's could involve in glucocorticoid-induced human bone mesenchymal stem cells (hBMSCs) differentiation and autophagy regulation remain not been elucidated.
METHODS
hBMSCs were identified by flow cytometry method, and its differentiation ability were measured by ARS staining, oil O red, and Alcian blue staining assays. Gene and proteins were quantified via qRT-PCR and western blot assays, respectively. Autophagy activity was determined using immunofluorescence. ChIP and dual luciferase assay validated the molecular interactions.
RESULTS
The data revealed that isolated hBMSCs exhibited positive of CD29/CD44 and negative CD45/CD34. Moreover, BACH1 was abated gradually during osteoblast differentiation of hBMSCs, while dexamethasone (Dex) treatment led to BACH1 upregulation. Loss of BACH1 improved osteoblast differentiation and activated autophagy activity in Dex-challenged hBMSCs. Autophagy-related proteins (ATG3, ATG4, ATG5, ATG7, ATG12) were repressed after Dex treatment, while ATG3, ATG7 and BECN1 could be elevated by BACH1 knockdown, especially ATG7. Moreover, BACH1 could interact ATG7 promoter region to inhibit its transcription. Co-inhibition of ATG7 greatly overturned the protective roles of BACH1 loss on osteoblast differentiation and autophagy in Dex-induced hBMSCs.
CONCLUSION
Taken together, our results demonstrated that silencing of BACH1 mitigated Dex-triggered osteogenic differentiation inhibition by transcriptionally activating ATG7-mediated autophagy, suggesting that BACH1 may be a therapeutic target for GIOP treatment.
背景
糖皮质激素诱导性骨质疏松症（GIOP）是最常见的继发性骨质疏松症。最近，自噬被发现与各种疾病的发展有关，包括骨质疏松症和成骨细胞分化调节。BTB 和 CNC 同源 1（BACH1）是先前证实的成骨细胞分化调节剂，但它是否参与糖皮质激素诱导的人骨髓间充质干细胞（hBMSCs）分化和自噬调节仍未阐明。
方法
通过流式细胞术鉴定 hBMSCs，通过 ARS 染色、油 O 红和阿利新蓝染色测定其分化能力。通过 qRT-PCR 和 Western blot 分别定量基因和蛋白质。通过免疫荧光测定自噬活性。ChIP 和双荧光素酶测定验证分子相互作用。
结果
数据显示，分离的 hBMSCs 呈 CD29/CD44 阳性和 CD45/CD34 阴性。此外，BACH1 在 hBMSCs 成骨分化过程中逐渐减少，而地塞米松（Dex）处理导致 BACH1 上调。BACH1 缺失可改善 Dex 刺激的 hBMSCs 中的成骨分化并激活自噬活性。自噬相关蛋白（ATG3、ATG4、ATG5、ATG7、ATG12）在 Dex 处理后受到抑制，而 BACH1 敲低可上调 ATG3、ATG7 和 BECN1，尤其是 ATG7。此外，BACH1 可以与 ATG7 启动子区域相互作用以抑制其转录。ATG7 的共同抑制极大地推翻了 BACH1 缺失对 Dex 诱导的 hBMSCs 中成骨分化和自噬的保护作用。
结论
总之，我们的研究结果表明，沉默 BACH1 通过转录激活 ATG7 介导的自噬来减轻 Dex 触发的成骨分化抑制，表明 BACH1 可能是 GIOP 治疗的一个治疗靶点。
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