Jo Sungsin, Koo Bon San, Lee Bitnara, Kwon Eunji, Lee Young Lim, Chung Heekyoung, Sung Il-Hoon, Park Ye-Soo, Kim Tae-Hwan
Hanyang University Hospital for Rheumatic Diseases, Seoul, Republic of Korea.
Department of Rheumatology, Inje University Seoul Paik Hospital, Seoul, Republic of Korea.
Biochem Biophys Res Commun. 2017 Sep 23;491(3):787-793. doi: 10.1016/j.bbrc.2017.07.079. Epub 2017 Jul 18.
The main aim of this study are to explore the role of bone-derived cells (BdCs) in ankylosing spondylitis (AS) and determine the underlying molecular mechanisms of IL-23 production. Primary BdCs were isolated from diced bone of facet joints obtained during surgery from seven AS patients and seven disease control (Ct) patients. Osteoblastic activity of BdCs was assessed by measuring their alkaline phosphatase activity and by alizarin red staining. Osteoblast and endoplasmic reticulum (ER) stress-related genes were assessed by quantitative PCR, immunoblotting, immunofluorescence, and immunohistochemistry. In addition, expression of IL-23 in response to BIX (selective BIP inducer X)-induced ER stress was evaluated by qPCR and ELISA. Protein interaction and binding to IL-23 promoter were confirmed by Immunoprecipitation and Chromatin immunoprecipitation, respectively. Transcript levels of genes involved in osteoblast function, as well as of the ER stress marker were higher in the AS group than the Ct group, and elevated RUNX2, BiP and IL-23 expression were observed in the BdCs, serum, and bone biopsies from the AS group. BIX-induced ER stress stimulated osteoblastic activity and IL-23 secretion by upregulating RUNX2 expression. Furthermore, in AS BdCs, RUNX2 interacted with C/EBPβ to bind to IL-23 promoter and RUNX2 knockdown suppressed IL-23 secretion. These finding may provide a molecular mechanism involved in sustained ER stress in AS BdCs stimulates the activation of RUNX2 and C/EBPβ genes, leading to IL-23 production.
本研究的主要目的是探讨骨源性细胞（BdCs）在强直性脊柱炎（AS）中的作用，并确定白细胞介素-23（IL-23）产生的潜在分子机制。从7例AS患者和7例疾病对照（Ct）患者手术中获取的小关节骨切块中分离出原代BdCs。通过测量其碱性磷酸酶活性和茜素红染色来评估BdCs的成骨活性。通过定量PCR、免疫印迹、免疫荧光和免疫组织化学评估成骨细胞和内质网（ER）应激相关基因。此外，通过qPCR和ELISA评估对BIX（选择性BIP诱导剂X）诱导的ER应激反应中IL-23的表达。分别通过免疫沉淀和染色质免疫沉淀证实蛋白质与IL-23启动子的相互作用和结合。AS组中参与成骨细胞功能的基因以及ER应激标志物的转录水平高于Ct组，并且在AS组的BdCs、血清和骨活检中观察到RUNX2、BiP和IL-23表达升高。BIX诱导的ER应激通过上调RUNX2表达刺激成骨活性和IL-23分泌。此外，在AS BdCs中，RUNX2与C/EBPβ相互作用以结合IL-23启动子，并且RUNX2敲低抑制IL-23分泌。这些发现可能提供了一种分子机制，即AS BdCs中持续的ER应激刺激RUNX2和C/EBPβ基因的激活，导致IL-23产生。
