Overexpression of Circ_0004496 is Associated with Pathological Bone Formation in Ankylosing Spondylitis via the miR-145/ACTG1 Axis.

IF 4.5 4区医学 Q2 MEDICINE, GENERAL & INTERNAL

Balkan Medical Journal Pub Date : 2026-04-30 Epub Date: 2026-03-10 DOI:10.4274/balkanmedj.galenos.2026.2025-12-249

Yu-Cong Zou, Yong-Sheng Wu, Ting Hu, Hao-Bin Zeng

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引用次数: 0

Abstract

Background: The functional role of circular ribonucleic acids in ankylosing spondylitis (AS) remains poorly understood. In our previous study, circ_0004496 and actin gamma 1 (ACTG1) were found to be upregulated in ossified tissues from patients with AS.

Aims: To investiagate the regulatory role of circ_0004496 in pathological bone formation through the miR-145/ACTG1 axis.

Study design: Combined in vitro and in vivo experimental study.

Methods: Tissue samples were obtained from 15 patients with AS and hip ankylosis and 15 control patients with femoral neck fractures. Quantitative real-time polymerase chain reaction was performed to measure circ_0004496 and miR-145 expression. Western blot analysis was used to determine the protein levels of alkaline phosphatase (ALP), osteocalcin (OCN), runt-related transcription factor 2 (Runx2), and ACTG1. Cell proliferation was evaluated using Cell Counting Kit-8 and 5-ethynyl-2'-deoxyuridine assays. Osteogenic differentiation was assessed by measuring ALP activity and performing Alizarin Red S staining. Interactions among circ_0004496, miR-145, and ACTG1 were examined using RNA immunoprecipitation (RIP), RNA pull-down, dual-luciferase reporter, and fluorescence in situ hybridization (FISH) assays.

Results: Circ_0004496 and ACTG1 expression levels were significantly elevated in AS hip capsule tissues, whereas miR-145 expression was reduced. Overexpression of circ_0004496 in AS-derived fibroblasts enhanced cell proliferation, accelerated cell cycle progression, increased calcium deposition, and upregulated ALP, OCN, and Runx2 protein levels. Mechanistically, circ_0004496 functioned as a molecular sponge for miR-145, which directly targets ACTG1. These regulatory interactions were confirmed by dual-luciferase reporter, RIP, RNA pulldown, and FISH assays. In vivo, circ_0004496 overexpression was associated with sacroiliac joint fusion in proteoglycan-induced arthritis mice.

Conclusion: Circ_0004496 overexpression promotes pathological bone formation in AS by regulating the miR-145/ACTG1 axis both in vitro and in vivo.

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Circ_0004496过表达通过miR-145/ACTG1轴与强直性脊柱炎的病理性骨形成相关。

背景：环状核糖核酸在强直性脊柱炎（AS）中的功能作用仍然知之甚少。在我们之前的研究中，circ_0004496和actin γ - 1 （ACTG1）在AS患者的骨化组织中被发现上调。目的：本研究通过miR-145/ACTG1轴探讨circ_0004496在病理性骨形成中的调节作用。研究设计：体内体外联合实验研究。方法：选取15例AS合并髋关节强直患者和15例股骨颈骨折患者作为对照。采用实时定量聚合酶链反应检测circ_0004496和miR-145的表达。Western blot检测碱性磷酸酶（ALP）、骨钙素（OCN）、矮子相关转录因子2 （Runx2）、ACTG1蛋白水平。使用细胞计数试剂盒-8和5-乙基-2'-脱氧尿苷测定法评估细胞增殖。通过测定ALP活性和茜素红S染色评估成骨分化。circ_0004496、miR-145和ACTG1之间的相互作用采用RNA免疫沉淀（RIP）、RNA下拉、双荧光素酶报告基因和荧光原位杂交（FISH）检测。结果：Circ_0004496和ACTG1在AS髋关节囊组织中的表达水平显著升高，而miR-145的表达水平降低。as衍生成纤维细胞中circ_0004496的过表达增强了细胞增殖，加速了细胞周期进程，增加了钙沉积，上调了ALP、OCN和Runx2蛋白水平。在机制上，circ_0004496作为miR-145的分子海绵，直接靶向ACTG1。通过双荧光素酶报告基因、RIP、RNA拉下和FISH检测证实了这些调节相互作用。在体内，circ_0004496过表达与蛋白多糖诱导的关节炎小鼠的骶髂关节融合有关。结论：Circ_0004496过表达通过调节miR-145/ACTG1轴在体外和体内促进AS病理性骨形成。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Balkan Medical Journal MEDICINE, GENERAL & INTERNAL-

CiteScore

4.10

自引率

6.70%

发文量

审稿时长

6-12 weeks

期刊介绍： The Balkan Medical Journal (Balkan Med J) is a peer-reviewed open-access international journal that publishes interesting clinical and experimental research conducted in all fields of medicine, interesting case reports and clinical images, invited reviews, editorials, letters, comments and letters to the Editor including reports on publication and research ethics. The journal is the official scientific publication of the Trakya University Faculty of Medicine, Edirne, Turkey and is printed six times a year, in January, March, May, July, September and November. The language of the journal is English. The journal is based on independent and unbiased double-blinded peer-reviewed principles. Only unpublished papers that are not under review for publication elsewhere can be submitted. Balkan Medical Journal does not accept multiple submission and duplicate submission even though the previous one was published in a different language. The authors are responsible for the scientific content of the material to be published. The Balkan Medical Journal reserves the right to request any research materials on which the paper is based. The Balkan Medical Journal encourages and enables academicians, researchers, specialists and primary care physicians of Balkan countries to publish their valuable research in all branches of medicine. The primary aim of the journal is to publish original articles with high scientific and ethical quality and serve as a good example of medical publications in the Balkans as well as in the World.