Effects of Targeting the Interleukin-1 Receptor in a Murine Model of Sickle Cell Disease
Authors: Venugopal Jessica , Wang Jintao , Guo Chiao , Eitzman Daniel 
Published: November 2019

Background: Sickle cell disease (SCD) is associated with chronic hemolytic anemia and a heightened inflammatory state. Multiple inflammatory pathways have been shown to be activated in SCD, however the causal role of these pathways in mediating organ damage and contributing to stroke is unclear.

Methods: Bone marrow transplantation (bmt) from SCD (Townes) to wild-type (WT) recipient mice was used to generate SCD mice (WTSCD). To generate mice with non-hematopoietic deficiency of the interleukin-1 receptor (IL1R), SCD marrow was transplanted into IL1R deficient recipients (IL1R-/- SCD). Other WTSCD mice were given the IL1R inhibitor, anakinra, or PBS for 2 or 4 weeks before sacrifice. Iron and collagen staining were performed in multiple formalin-fixed tissues with Prussian blue and Masson's Trichrome, respectively. Complete blood counts and reticulocyte percentages were analyzed 15 weeks following BMT. Plasma levels of soluble E-selectin (sE-sel) and soluble P-selectin (sP-sel) were measured 14 weeks following bmt with ELISA. Ischemic stroke was induced by middle cerebral artery (MCA) photothrombosis at 16 weeks of age. WTSCD mice were given anakinra following stroke induction. Seventy-two hours after MCA occlusion, stroke volume was assessed by staining brain sections with 2, 3, 5-triphenyltetrazolium chloride. Formalin-fixed brain sections were also stained for macrophages.

Results: All SCD mice were anemic and the severity of anemia was not different between WTSCD and IL1R-/- SCD mice. Increased circulating erythrocytes (9.01 ± 0.78 vs 6.77 ± 0.18 M/uL; p<0.05) and decreased reticulocytes (25.31 ± 1.79 vs 32.45 ± 2.74%; p<0.05) were observed in WTSCD mice treated with anakinra for 4 weeks. Two week anakinra treatment was sufficient to decrease organ iron deposition and lung collagen deposition. In vivo treatment with anakinra also decreased erythrocyte sickling in a whole blood ex vivo sickling assay (15.14 ± 0.94 vs 24.18 ± 1.95 % sickled; p<0.01). Further, in vitro treatment of WTSCD whole blood with IL-1β increased sickling (26.70 ± 0.39 vs 22.34 ± 0.95; p<0.05), and this was ameliorated with in vitro pretreatment with anakinra. Alterations to ex vivo sickling were not observed when conditions were repeated on erythrocytes alone. Stroke volume was significantly reduced in IL1R-/- SCD mice compared to WTSCD mice 3 days following MCA occlusion (8.08 ± 0.91 vs 11.10 ± 1.16 % hemisphere; p<0.05). Post-stroke treatment of WTSCD mice with anakinra also decreased stroke size compared to vehicle-treated mice (14.45 ± 1.44 vs 9.7 ± 3.23 % hemisphere; p=0.05). Non-hematopoietic deficiency of IL1R decreased sE-sel (vs 50.10 ± 2.31 vs 32.12 ± 2.08 pg/mL; p<0.05) and sP-sel (155.98 ± 24.44 vs 327.23 ± 18.31 ng/mL; p<0.05) levels in post-stroke animals and decreased the number of macrophages observed in the peri-stroke area (0.26 ± 0.05 vs 0.722 ± 0.15 % hemisphere; p<0.05).

Conclusions: Deficiency of non-hematopoietic IL1R or treatment with an IL1R antagonist was appears beneficial towards several phenotypes in a mouse model of SCD. Given the safety of IL1R inhibitors in various patient populations, this study reveals a potential therapeutic intervention for treatment of sickle cell patients.

Disclosures

No relevant conflicts of interest to declare.