Hanyang University Institute for Rheumatology Research (HYIRR), Hanyang University, Seoul 04763, Republic of Korea; Deparment of Translational Medicine, Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul 04763, Republic of Korea.
Hanyang University Institute for Rheumatology Research (HYIRR), Hanyang University, Seoul 04763, Republic of Korea.
Int Immunopharmacol. 2023 May;118:110132. doi: 10.1016/j.intimp.2023.110132. Epub 2023 Apr 4.
Ankylosing spondylitis (AS) exhibits paradoxical bone features typically characterized by new bone formation and systemic bone loss. Although abnormal kynurenine (Kyn), a tryptophan metabolite, has been closely linked to the disease activity of AS, the distinct role of its pathological bone features remains unknown.
Kynurenine sera level was collected from healthy control (HC; n = 22) and AS (n = 87) patients and measured by ELISA. In the AS group, we analyzed and compared the Kyn level based on the modified stoke ankylosing spondylitis spinal score (mSASSS), MMP13, and OCN. Under osteoblast differentiation, the treatment with Kyn in AS-osteoprogenitors conducted cell proliferation, alkaline phosphatase activity, bone mineralization-related alizarin red s (ARS), von kossa (VON), hydroxyapatite (HA) staining, and mRNA expression markers (ALP, RUNX2, OCN, and OPG) for bone formation. TRAP and F-actin staining was used for osteoclast formation of mouse osteoclast precursors.
Kyn sera level was significantly elevated in the AS group compared to the HC. In addition, Kyn sera level was correlated with mSASSS (r = 0.03888, p = 0.067), MMP13 (r = 0.0327, p = 0.093), and OCN (r = 0.0436, p = 0.052). During osteoblast differentiation, treatment with Kyn exhibited no difference in cell proliferation and alkaline phosphate (ALP) activity for bone matrix maturation but promoted ARS, VON, and HA staining for bone mineralization. Interestingly, osteoprotegerin (OPG) and OCN expressions of AS-osteoprogenitors were augmented in the Kyn treatment during differentiation. In growth medium, Kyn treatment of AS-osteoprogenitors resulted in induction of OPG mRNA, protein expression, and Kyn-response genes (AhRR, CYP1b1, and TIPARP). Secreted OPG proteins were observed in the supernatant of AS-osteoprogenitors treated with Kyn. Notably, the supernatant of Kyn-treated AS-osteoprogenitors interrupted the RANKL-mediated osteoclastogenesis of mouse osteoclast precursor such as TRAP-positive osteoclast formation, NFATc1 expression, and osteoclast differentiation markers.
Our results revealed that elevated Kyn level increased the bone mineralization of osteoblast differentiation in AS and decreased RANKL-mediated osteoclast differentiation by inducing OPG expression. Out study have implication for potential coupling factors linking osteoclast and osteoblast where abnormal Kyn level could be involved in pathological bone features of AS.
强直性脊柱炎（AS）表现出矛盾的骨骼特征，通常表现为新骨形成和全身骨质流失。尽管异常的犬尿氨酸（Kyn），一种色氨酸代谢物，与 AS 的疾病活动密切相关，但它在病理性骨骼特征中的独特作用尚不清楚。
从健康对照组（HC；n=22）和 AS 患者（n=87）中采集犬尿氨酸血清水平，并通过 ELISA 进行测量。在 AS 组中，我们根据改良的斯托克强直性脊柱炎脊柱评分（mSASSS）、MMP13 和 OCN 分析和比较了 Kyn 水平。在成骨细胞分化过程中，用 Kyn 处理 AS-成骨前体细胞，进行细胞增殖、碱性磷酸酶活性、骨矿化相关茜素红 s（ARS）、Von Kossa（VON）、羟基磷灰石（HA）染色以及骨形成的 mRNA 表达标志物（ALP、RUNX2、OCN 和 OPG）。TRAP 和 F-肌动蛋白染色用于检测小鼠破骨细胞前体的破骨细胞形成。
与 HC 相比，AS 组的犬尿氨酸血清水平显著升高。此外，犬尿氨酸血清水平与 mSASSS（r=0.03888，p=0.067）、MMP13（r=0.0327，p=0.093）和 OCN（r=0.0436，p=0.052）相关。在成骨细胞分化过程中，用 Kyn 处理对细胞增殖和碱性磷酸酶（ALP）活性无影响，但促进了 ARS、VON 和 HA 染色的骨矿化。有趣的是，在分化过程中，Kyn 处理增加了 AS-成骨前体细胞的骨保护素（OPG）和 OCN 的表达。在生长培养基中，Kyn 处理 AS-成骨前体细胞诱导 OPG mRNA、蛋白表达和 Kyn 反应基因（AhRR、CYP1b1 和 TIPARP）。用 Kyn 处理的 AS-成骨前体细胞的上清液中观察到分泌的 OPG 蛋白。值得注意的是，Kyn 处理的 AS-成骨前体细胞的上清液通过诱导 OPG 表达，中断了 RANKL 介导的小鼠破骨细胞前体的破骨细胞生成，如 TRAP 阳性破骨细胞形成、NFATc1 表达和破骨细胞分化标志物。
我们的结果表明，AS 中犬尿氨酸水平升高增加了成骨细胞分化的骨矿化，通过诱导 OPG 表达减少了 RANKL 介导的破骨细胞分化。我们的研究为潜在的连接破骨细胞和成骨细胞的偶联因子提供了依据，其中异常的犬尿氨酸水平可能参与了 AS 的病理性骨骼特征。
