Circulating levels of microRNA-539-5p, also designated as miR-539-5p, are significantly downregulated in serum collected from patients diagnosed with ankylosing spondylitis compared to control cohorts, demonstrating notable diagnostic utility. As highlighted by the investigators, reliable biomarkers for the early diagnosis of ankylosing spondylitis and for monitoring its characteristic pathological bone remodeling imbalance remain insufficient in clinical settings. Addressing this diagnostic shortfall, the study was designed to determine whether circulating miR-539-5p can serve as an effective diagnostic biomarker and whether it actively modulates osteogenic and osteoclast differentiation phenotypes in laboratory models by directly targeting secreted phosphoprotein 1, or SPP1.
To evaluate the clinical potential of the biomarker, the researchers collected and analyzed serum samples from three distinct participant groups: patients confirmed with ankylosing spondylitis, a comparative Lumbar Muscle Strain group, and healthy controls. Quantitative measurements showed that serum miR-539-5p was significantly downregulated among individuals in the ankylosing spondylitis cohort relative to the comparison groups. In receiver operating characteristic curve evaluations, serum miR-539-5p exhibited high diagnostic accuracy in distinguishing ankylosing spondylitis patients from individuals with lumbar muscle strain, achieving an area under the curve value of 0.881. Statistical analyses further identified serum miR-539-5p as an independent protective factor, supporting its clinical relevance as a candidate indicator for the condition.
Alongside patient serum profiling, the investigators conducted a series of controlled laboratory experiments to clarify how miR-539-5p affects cellular functions during bone remodeling in vitro. The team utilized two induced cell lines representing distinct bone cell lineages: MC3T3-E1 cells, which model osteoblasts, and RAW264.7 cells, which serve as models for osteoclasts. Both cell types were transfected with miR-539-5p mimics to elevate microRNA levels or miR-539-5p inhibitors to suppress expression. To comprehensively assess the resulting biological functions and underlying molecular pathways, the researchers performed Cell Counting Kit-8 assays for viability, alkaline phosphatase and tartrate-resistant acid phosphatase activity assays for enzymatic behavior, quantitative reverse transcription polymerase chain reaction for gene transcription, Western blotting for protein abundance, and dual-luciferase reporter assays for target binding.
The functional cell assays revealed that miR-539-5p overexpression significantly influenced both osteogenic and osteoclastogenic phenotypes. In the induced MC3T3-E1 cells, transfection with the miR-539-5p mimic suppressed the dexamethasone-induced increase in osteoblast viability. Dual-luciferase assays demonstrated that miR-539-5p directly targeted the SPP1 gene, and quantitative analyses of patient sera confirmed that SPP1 messenger RNA expression was negatively correlated with circulating miR-539-5p concentrations. Furthermore, rescue experiments using an SPP1 overexpression plasmid showed that restoring SPP1 substantially attenuated the inhibitory and regulatory effects exerted by the miR-539-5p mimic on differentiation phenotypes.
In their conclusion, the authors noted that serum miR-539-5p may serve as a promising diagnostic biomarker for ankylosing spondylitis, particularly given its capacity to distinguish cases from lumbar muscle strain. Additionally, the experimental findings indicate that miR-539-5p influences cellular behavior in in vitro bone remodeling models by targeting SPP1 and partially modulating both osteogenic and osteoclast differentiation.
Subject of Research: Serum miR-539-5p as a diagnostic biomarker for ankylosing spondylitis and its in vitro regulation of bone remodeling via SPP1
Article Title: Serum miR-539-5p decreases in ankylosing spondylitis and regulates osteogenic and osteoclastogenic phenotypes in vitro via SPP1
Article References: Lin, M., Dai, B., Cheng, Z., & Deng, R. (2026). Serum miR-539-5p decreases in ankylosing spondylitis and regulates osteogenic and osteoclastogenic phenotypes in vitro via SPP1. BMC Musculoskeletal Disorders. https://doi.org/10.1186/s12891-026-10632-3
Image Credits: AI Generated
DOI: 10.1186/s12891-026-10632-3
Keywords: miR-539-5p, ankylosing spondylitis, SPP1, bone remodeling, osteoblast differentiation, osteoclastogenesis, MC3T3-E1 cells, RAW264.7 cells, lumbar muscle strain, alkaline phosphatase, tartrate-resistant acid phosphatase, dual-luciferase reporter assay, qRT-PCR, Western blot, CCK-8 assay, dexamethasone, biomarkers, AUC analysis
