The Role of Integrins a_Mb₂ (CD11b/CD18) and a_Db₂ (CD11d/CD18) in Macrophage Fusion.

Abstract 2138

Macrophage fusion leading to the formation of multinucleated giant cells (MGCs) is a hallmark of many chronic inflammatory reactions. MGCs are an invariable constituent of tuberculoid lesions and also found in a variety of conditions leading to granulomatous inflammation as well as the foreign body reaction. Despite the prominent phenotype, the molecular mechanisms underlying macrophage fusion are not well understood. MGCs originate from macrophages that are recruited to sites of chronic inflammation. The major myelo-monocytic integrin α_Mβ₂ (CD11b/CD18, Mac-1), together with a related integrin α_Dβ₂ (CD11d/CD18), mediate critical adhesive reactions of monocyte/macrophages. The function of β₂ integrins in macrophage fusion remains controversial. Some studies using function blocking antibodies implicated α_Mβ₂ in the cell/substrate adhesive interactions that are required for MGC formation, whereas one recent report indicated that this integrin plays a minor role. Moreover, the contribution of α_Dβ₂, a receptor with recognition specificity overlapping that of α_Mβ₂, to macrophage fusion is unknown. To evaluate the role of α_Mβ₂ and α_Dβ₂ in MGC formation, we examined fusion of inflammatory peritoneal macrophages isolated from wild-type mice and mice with deficiency of α_M or α_D integrin subunits. Macrophages were isolated at day 3 after thioglycollate injection and cultured for 24–72 hours in the presence of IL-4 to induce fusion. Percentage fusion was quantified as the number of giant cell nuclei (≥2 nuclei) to the number of total nuclei. The number of fused macrophages isolated from wild-type mice gradually increased and ∼40–50% macrophages formed MGCs after IL-4 stimulation by day 3. Analyses of fusion of α_Mβ₂-deficient macrophages demonstrated that fusion was significantly reduced. By day 3, macrophage fusion of α_Mβ₂-deficient macrophages was 23 ± 2% of wild-type macrophages. Fusion of α_Dβ₂-deficient macrophages was also decreased and the change was statistically significant, albeit to a smaller degree (75 ± 4%). Using a mouse model of sterile peritonitis induced by thioglycollate injection, we also examined the formation of MGCs in vivo. In wild-type mice, the number of MGCs gradually increased from 2.5 ± 0.3% at day 0 (resident macrophages) to 17 ± 2% at day 18 (the resolution phase of inflammation). Moreover, expression of α_Mβ₂ and α_Dβ₂ on peritoneal macrophages increased by ∼2 and 1.6-fold, respectively, on 18^th day after induction of inflammation. In α_Mβ₂-deficient mice, the number of MGCs was reduced by 2.1-fold compared to wild-type mice. Furthermore, while the size of wild-type and α_Mβ₂-deficient giant cells was the same, the number of cells with 3 and more nuclei in fused α_Mβ₂-deficient MGCs was 4-fold less than in wild-type cells. The results indicate that both α_Mβ₂ and α_Dβ₂ integrins support macrophage fusion with α_Mβ₂ playing a dominant role.