Abstract T P237: Effect of Danger Associated Molecular Pattern Receptor Complex Proteins CD24 and Siglec-G on Stroke Outcome and Microglial Responses

Abstract

Background: Both microglia and Toll-like receptors (TLRs) are critical in stroke pathophysiology. In ischemic brain, microglia sense endogenous TLR agonists (danger associated molecular patterns or DAMPs) and respond with varied immune reactions. CD24 and Siglec-G form a receptor complex that modulates TLR4 function and controls responses to DAMPs. The role of CD24 and Siglec-G in stroke is unknown. Methods: We performed 45 min middle cerebral artery occlusion (MCAO) on 12 - 14 week old wild-type, TLR4-/-, CD24-/- and Siglec-G-/- male mice and assessed total and regional adjusted infarct volumes at 48 hours with 2,3,5-triphenyltetrazolium staining. Number of mice per group was determined by power analysis. Cerebral blood flow was assessed with laser doppler flowmetry. In vitro, we examined the effects of endogenous TLR4 agonists heat shock protein-70 and high mobility group box 1 on cytokine (TNFα, IL-6) and chemokine (CXCL10, CCL5) release from microglia derived from wild-type, TLR4-/-, CD24-/- and Siglec-G-/- mice. Results: Following exclusions for weight, temperature and sub-optimal vessel occlusion/reperfusion, total infarct volumes (mean±SEM) were 51±8 mm3 (n = 21), 51±10 mm3 (n = 8), 28±8 mm3 (n = 13) and 54±8 mm3 (n = 19) in wild-type, TLR4-/-, CD24-/- and Siglec-G-/- mice, respectively (p>0.05, one-way ANOVA). Release of cytokines and chemokines was absent (as expected) in microglia from TLR4-/- mice and differentially regulated in microglia from CD24-/- and Siglec-G-/- mice. Conclusions: Genetic deficiency in TLR4, CD24 or Siglec-G modulated microglial response to endogenous TLR4 agonists but did not significantly alter post-stroke infarct volume.