Lambertini Elisabetta, Penolazzi Letizia, Angelozzi Marco, Grassi Francesco, Gambari Laura, Lisignoli Gina, De Bonis Pasquale, Cavallo Michele, Piva Roberta
Department of Biomedical and Specialty Surgical Sciences, University of Ferrara, Ferrara, Italy.
Ramses Laboratory, Rizzoli Orthopedic Institute, Bologna, Italy.
Oncotarget. 2017 Oct 4;8(60):101686-101696. doi: 10.18632/oncotarget.21514. eCollection 2017 Nov 24.
Hydrogen sulfide (HS), generated in the osteoblasts predominantly via cystathionine-γ-lyase (CSE), is bone protective. Previous studies suggested that the onset of bone loss due to estrogen deficiency is associated to decreased levels of HS and blunted gene expression of CSE. However, there are still a lot of unknowns on how HS levels influence bone cells function. The present study aims to explore the mechanisms by which estrogen may regulate CSE expression, in particular the role of estrogen receptor alpha (ERα) in human osteoblasts (hOBs). Vertebral lamina derived hOBs were characterized and then assessed for CSE expression by western blot analysis in the presence or absence of ERα overexpression. Bioinformatic analysis, luciferase reporter assay and ChIP assay were performed to investigate ERα recruitment and activity on hCSE gene promoter. Three putative half Estrogen Responsive Elements (EREs) were identified in the hCSE core promoter and were found to participate in the ERα - mediated positive regulation of CSE expression. All osteoblast samples responded to ERα over-expression increasing the levels of CSE protein in a comparable manner. Notably, the ERα recruitment on the regulatory regions of the CSE promoter occurred predominantly in female hOBs than in male hOBs. The obtained results suggest that CSE/HS system is in relation with estrogen signaling in bone in a gender specific manner.
硫化氢（HS）主要通过胱硫醚-γ-裂解酶（CSE）在成骨细胞中生成，具有骨保护作用。先前的研究表明，雌激素缺乏导致的骨质流失与HS水平降低和CSE基因表达减弱有关。然而，关于HS水平如何影响骨细胞功能仍有许多未知之处。本研究旨在探讨雌激素调节CSE表达的机制，特别是雌激素受体α（ERα）在人成骨细胞（hOBs）中的作用。对源自椎板的hOBs进行表征，然后在有或没有ERα过表达的情况下通过蛋白质印迹分析评估CSE表达。进行生物信息学分析、荧光素酶报告基因测定和染色质免疫沉淀测定以研究ERα对hCSE基因启动子的募集和活性。在hCSE核心启动子中鉴定出三个假定的半雌激素反应元件（ERE），并发现它们参与ERα介导的CSE表达的正调控。所有成骨细胞样本对ERα过表达均有反应，以类似方式增加CSE蛋白水平。值得注意的是，ERα在CSE启动子调控区域的募集在女性hOBs中比在男性hOBs中更常见。所得结果表明，CSE/HS系统以性别特异性方式与骨中的雌激素信号传导相关。
