Hemarthrosis in FVIII-deficient Mice Causes Altered Expression of Local and Systemic Iron Regulators and Defective Iron Clearance from the Joint
ISTH Academy. von Drygalski A. Jul 10, 2019; 274128; OC 75.4 Topic: Hemophilia - Basic
Annette von Drygalski
Annette von Drygalski
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OC 75.4

Hemarthrosis in FVIII-deficient Mice Causes Altered Expression of Local and Systemic Iron Regulators and Defective Iron Clearance from the Joint

E. Cooke1,2, C. Nasamran3, K. Fisch3, A. von Drygalski1,2
1University of California San Diego, Department of Medicine, La Jolla, United States, 2The Scripps Research Institute, Department of Molecular Medicine, La Jolla, United States, 3University of California San Diego, Center for Computational Biology & Bioinformatics, La Jolla, United States

Main Topic: Hemophilia and Bleeding (including Transfusion)
Category: Hemophilia - Basic

Background: Hemarthrosis in hemophilia causes toxic iron accumulation in the joint, which contributes to synovitis.
Aims: To explore mechanisms of iron clearance after hemarthrosis, and whether these are defective in hemophilia.
Methods: Hemarthrosis was induced by sub-patellar needle puncture in FVIII-deficient mice and hypocoagulable BALB/c (HypoBALB/c) mice treated with 10µg/ml warfarin for 7 days and 0.25mg/kg anti-FVIII 2 hours before injury. Warfarin was reversed with 100IU/kg 4‑factor prothrombinase complex concentrate on day 2 post-injury +/- twice weekly anti-FVIII treatment. Ferric iron was quantified in joint tissue at 2-4 weeks post-injury by Prussian Blue staining. Splenic and synovial tissue were harvested from FVIII-deficient mice on day 3 and 2 weeks post-injury for gene expression studies by RNA-sequencing.
Results: Knee injury caused substantial hemarthrosis in all mice. A 5.3-fold increase (p=0.003) in ferric iron, as found in the iron-storage complex, hemosiderin, was detected in FVIII-deficient mice at week 4 only. No ferric iron was detected in HypoBALB/c mice despite comparable bleeding and anti-FVIII treatment. 10/56 genes relating to iron transport were differentially expressed (DE) in synovium from FVIII-deficient mice on day 3, persisting to some extent at 2 weeks. These included up-regulation of heme oxygenase-1 (31-fold; p=3x10-6) and lipocalin-2 (10-fold; p=0.001), and down-regulation of ceruloplasmin (17-fold; p=5x10-5), CD163 (hemoglobin scavenging receptor) (5-fold; p=0.03), and hephaestin (5-fold; p=0.002). In the spleen, there were 12 DE genes on day 3 only, including up-regulation of Steap-3 (4-fold; p=0.0008) and transferrin receptor-1 (3-fold; p=0.005), and down-regulation of lipocalin-2 (4-fold; p=0.01). These changes may influence iron scavenging/retention and transport in synovium, and cellular uptake of iron in the spleen in response to anemia.
Conclusions: Iron accumulation after hemarthrosis appears unique to hemophilia, suggesting impaired clearance. RNA-sequencing created insights into mechanisms of local and systemic iron transport, prompting further investigations to elucidate detrimental synovial iron accumulation.

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