Abstract
Monocyte differentiation to macrophages is triggered by migration across the endothelial barrier, which is constituted by both endothelial cells and their underlying basement membrane. We address here the role of the endothelial basement membrane laminins (laminins 411 and 511) in this monocyte to macrophage switch. Chimeric mice carrying CX3CR1-GFP bone marrow were employed to track CCL2-induced monocyte extravasation in a cremaster muscle model using intravital microscopy, revealing faster extravasation in mice lacking endothelial laminin 511 (Tek-cre::Lama5−/−) and slower extravasation in mice lacking laminin 411 (Lama4−/−). CX3CR1-GFPlow extravasating monocytes were found to have a higher motility at laminin 511 low sites and to preferentially exit vessels at these sites. However, in vitro experiments reveal that this is not due to effects of laminin 511 on monocyte migration mode nor on the tightness of the endothelial barrier. Rather, using an intestinal macrophage replenishment model and in vitro differentiation studies, we demonstrate that laminin 511, together with the attached endothelium, promote monocyte differentiation to macrophages. Macrophage differentiation is associated with a change in integrin profile, permitting differentiating macrophages to distinguish between laminin 511 high and low areas and to preferentially migrate across laminin 511 low sites. These studies highlight the endothelial basement membrane as a critical site for monocyte differentiation to macrophages, which may be relevant to the differentiation of other cells at vascular niches.
Highlights
Monocytes are a diverse population that varies in origin, in vivo localization, and function
CX3CR1GFP/+ mice have a high GFP expression on monocytes, and while GFP can be expressed by a subset of dendritic cells (DCs) and NK T cells, the GFP levels on these cells are significantly lower than that on monocytes [17, 25, 26]
To ensure that extravasating CX3CR1-GFP cells were predominantly myeloid cells and not DCs or NK T cells, cremaster muscles of WT chimeras were stained with anti-Gr-1 which detects both Ly6C+ monocytes and Ly6G+ neutrophils
Summary
Monocytes are a diverse population that varies in origin, in vivo localization, and function They represent an important early line of defense against infectious agents or reaction to tissue damage and in several tissues bone marrow derived monocytes play a critical role in the replenishment of resident monocytes, macrophages, and dendritic cells (DCs) required for tissue immunity [1]. Here use live imaging of CCL-2 induced monocyte extravasation in the cremaster muscle model to investigate whether the endothelial basement membrane represents an environmental factor that affects monocyte phenotype and/or infiltration into tissues. As reported for other leukocyte types, monocyte extravasation occurred preferentially at laminin 511 low sites; in vitro experiments revealed that this was not due to direct effects of laminin 511 on migration modes, nor effects on the tightness of the endothelial monolayer. Rather, using an intestinal macrophage replenishment model, we demonstrate that laminin 511 together with the attached endothelium provide a decisive cue, promoting monocyte differentiation to macrophages which, in turn, affects the extent of tissue infiltration
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