Cytokine dynamics following joint injury in hemophilia A mice

Introduction Hemarthrosis – including subclinical episodes – in hemophilia A (HA) provokes synovial inflammation and progressive arthropathy [ 1 ]. While the structural joint consequences are well described, the underlying inflammatory processes following joint bleeding and their modulation by FVIII prophylaxis remain poorly understood [ 2 ]. Using HA mice lacking anti-FVIII antibody formation (F8 -/- Ighm -/- ), we aimed to characterize cytokine dynamics after joint injury and evaluate their modulation by secondary FVIII prophylaxis. Method Mice were randomized into three groups: untreated (Gr.1), single FVIII injection (200 IU/kg, Gr.2), and FVIII prophylaxis (200 IU/kg on day 0, followed by 100 IU/kg on days 1–4 and three times per week thereafter, Gr.3). A unilateral knee injury was induced on day 1, and physical performance was monitored. Mice were sacrificed on days 4, 11, and 21 for sample collection. Plasma cytokines (IL-6, IL-10, IL-12p70, IFN-γ, MCP-1, TNF), transcriptomic profiling of splenocytes (RNA-seq), and histological analyses of hind limbs (Valentino scoring [ 3 ] and OARSI grading [ 4 ]) were performed. Results Gr.1 (untreated) and Gr.2 (single dose) showed knee swelling, reduced physical activity, and histological signs of overt joint bleeding. In Gr.3 (secondary prophylaxis), overt bleeding was largely prevented, with minimal or no knee swelling, preserved physical activity, and very little or no blood or hemosiderin deposition in joint histology. However, Gr.3 displayed synovial hyperplasia and cartilage damage similar to Gr.1 and Gr.2. IL-6 increased in Gr.1 and Gr.2, and IL-10 peaked in Gr.1, but neither was observed in Gr.3, consistent with an acute inflammatory response to bleeding rather than synovial inflammation and in line with recent findings [ 5 ]. Notably, IL-6 mRNA expression in splenocytes contrasted plasma levels, with a trend toward higher expression in Gr.2 and Gr.3, probably reflecting an immune response against human FVIII. Plasma IL-12p70 was increased in Gr.3 on day 11, possibly also reflecting FVIII-driven immune activation. MCP-1 and TNF-alpha did not differ between groups. There was a trend toward higher expression of Il1rl1 (ST2, the IL-33 receptor) in splenocytes of Gr.1 and Gr.2 compared with Gr.3, consistent with recent data implicating the IL-33/ST2 axis in hemophilic arthropathy [ 6 ]. Conclusion Systemic cytokine responses paralleled physical performance and joint pathology after hemarthrosis. Secondary FVIII prophylaxis normalized systemic profiles and prevented major bleeding-related changes, though synovial hyperplasia and cartilage damage persisted. These findings demonstrate that hemarthrosis is accompanied by systemic inflammatory activation and identify IL-6 as a marker of acute bleeding rather than chronic joint pathology. In addition, our data are consistent with recent reports implicating the IL-33/ST2 axis in hemophilic arthropathy. Publication History Article published online: 17 February 2026 © 2026. Thieme. All rights reserved. Georg Thieme Verlag KG Oswald-Hesse-Straße 50, 70469 Stuttgart, Germany