Tongtong Li¹ , Olaf Dekkers¹ , Ramon Arens² , Liesbeth Winter^1,3

¹Department of Medicine, Division of Endocrinology, Leiden University Medical Center

²Department of Immunology, Leiden University Medical Center, Leiden, The Netherlands.

³Department of Medicine, Center for Bone Quality, Leiden University Medical Center

Background

Adult chronic nonbacterial osteitis (CNO) is an autoinflammatory bone disorder characterized by localized bone pain and increased bone turnover. The underlying pathophysiology remains unclear, and systemic cytokine levels are largely unremarkable despite local inflammation. To refine our understanding of T-cell involvement, we profiled cytokine production at the single-cell level.

Methods

Peripheral blood mononuclear cells from adults with CNO (n=18) and healthy controls (n=17) were stimulated with PMA/ionomycin and assessed by multiparameter intracellular cytokine staining for IFN-γ, TNF, IL-17, IL-2 and IL-10 in CD4⁺ and CD8⁺ T cells.

Results

Compared with healthy controls, CNO patients exhibited increased proportions of CD3⁺ T cells (73.1% vs 82.0%, p = 0.009) and CD4⁺ T cells (65.5% vs 76.7%, p = 0.015), while CD8⁺ T cell frequencies were similar (25.0% vs 19.9%, p = 0.096). In contrast, CD4⁺IL-2⁺ (39.5% vs 33.2%, p = 0.031) and CD4⁺IL-10⁺ (0.29% vs 0.12%, p = 0.011) T cells, IL-2⁺/IL-10⁺ CD4⁺ T cells co-expressing IFN-γ or TNF-α, and IL-10⁺ CD8⁺ T cells were significantly reduced in adult CNO. Trends toward lower frequencies of multifunctional IFN-γ– and TNF-α–producing CD4⁺ and CD8⁺ subsets were observed. Geometric mean fluorescence intensity data indicated that cytokine production per cell was comparable between groups.

Conclusions

From a T-cell perspective, persistent bone inflammation in adult CNO does not seem to be driven by systemic overproduction of pro-inflammatory cytokines. Instead, our results indicate a chronically activated but functionally dysregulated T-cell compartment with a reduced pool of cytokine-producing subsets, particularly IL-10–producing cells, consistent with impaired IL-10–mediated regulation.