Metalloproteinases Atroxlysin-I (Atr-I) and Atroxlysin-III (Atr-III) from Bothrops atrox snake venom bind to murine macrophage CD11b altering cytokine production

Atroxlysin-I (Atr-I) and atroxlysin-III (Atr-III) are snake venom metalloproteinases (SVMPs) isolated from Peruvian Bothrops atrox. These proteins were previously characterized as hemorrhagic, fibrin(ogen)olytic and platelet aggregation inhibitors. In this work, we evaluated the response of murine macrophages treated with Atr-I (23 kDa, P-I class), Atr-III (56 kDa, P-III class), and B. atrox venom. Neither SVMPs nor crude venom decreased cell viability, whereas treatment with a higher concentration of Atr-I and B. atrox venom increased cellular metabolism. The SVMPs and B. atrox venom differentially induce mRNA expression of inflammatory cytokines, but only IL-1β was detected in the cell supernatants. Furthermore, SVMPs and crude venom modulate IL-6, TNF-α, and IL-1β production by macrophages previously stimulated with TLR2 and TLR4 agonists. The SVMPs differ in proteolytic activity on cytokines. Atr-I cleaves IL-6 and TNF-α, while Atr-III cleaves TNF-α and IL-1β. Also, Atr-I induced ERK1/2 and p38 pathway activation. Surprisingly, we show here for the first time that SVMPs are internalized and colocalize with CD11b, and that CD11b partially regulates cytokine production. Bioinformatics analyses identified two putative CD11b-interaction sites for Atr-I and one for Atr-III. The common region identified for both metalloproteinases encompasses the catalytic site. Altogether, these results demonstrate that SVMPs are recognized by macrophages and may modulate the inflammatory response in snakebite victims.