Although chemokines are sufficient for chemotaxis of various cells, increasing evidence exists for their fine-tuning by selective proteolytic processing. Using a model of immune cell chemotaxis into the CNS (experimental autoimmune encephalomyelitis [EAE]) that permits precise localization of immigrating leukocytes at the blood-brain barrier, we show that, whereas chemokines are required for leukocyte migration into the CNS, additional MMP-2/9 activities specifically at the border of the CNS parenchyma strongly enhance this transmigration process. Cytokines derived from infiltrating leukocytes regulate MMP-2/9 activity at the parenchymal border, which in turn promotes astrocyte secretion of chemokines and differentially modulates the activity of different chemokines at the CNS border, thereby promoting leukocyte migration out of the cuff. Hence, cytokines, chemokines, and cytokine-induced MMP-2/9 activity specifically at the inflammatory border collectively act to accelerate leukocyte chemotaxis across the parenchymal border.

Chemotaxis, the movement of cells along chemical gradients, is critical for the recruitment of immune cells to sites of inflammation; however, how cells navigate in chemotactic gradients is poorly understood. Here, we show that macrophages navigate in a gradient of the chemoattractant C5a through the release of adenosine triphosphate (ATP) and autocrine "purinergic feedback loops" that involve receptors for ATP (P2Y₂), adenosine diphosphate (ADP) (P2Y₁₂), and adenosine (A2a, A2b, and A3). Whereas macrophages from mice deficient in pannexin-1 (which is part of a putative ATP release pathway), P2Y₂, or P2Y₁₂ exhibited efficient chemotactic navigation, chemotaxis was blocked by apyrase, which degrades ATP and ADP, and by the inhibition of multiple purinergic receptors. Furthermore, apyrase impaired the recruitment of monocytes in a mouse model of C5a-induced peritonitis. In addition, we found that stimulation of P2Y₂, P2Y₁₂, or adenosine receptors induced the formation of lamellipodial membrane protrusions, causing cell spreading. We propose a model in which autocrine purinergic receptor signaling amplifies and translates chemotactic cues into directional motility.

Endothelial basement membranes constitute barriers to extravasating leukocytes during inflammation, a process where laminin isoforms define sites of leukocyte exit; however, how this occurs is poorly understood. In addition to a direct effect on leukocyte transmigration, we show that laminin 511 affects endothelial barrier function by stabilizing VE-cadherin at junctions and downregulating expression of CD99L2, correlating with reduced neutrophil extravasation. Binding of endothelial cells to laminin 511, but not laminin 411 or non-endothelial laminin 111, enhanced transendothelial cell electrical resistance (TEER) and inhibited neutrophil transmigration. Data suggest that endothelial adhesion to laminin 511 via β1 and β3 integrins mediates RhoA-induced VE-cadherin localization to cell-cell borders, and while CD99L2 downregulation requires integrin β1, it is RhoA-independent. Our data demonstrate that molecular information provided by basement membrane laminin 511 affects leukocyte extravasation both directly and indirectly by modulating endothelial barrier properties.

We describe a mouse strain in which B cell development relies either on the expression of membrane-bound immunoglobulin (Ig) γ1 or μ heavy chains. Progenitor cells expressing γ1 chains from the beginning generate a peripheral B cell compartment of normal size with all subsets, but a partial block is seen at the pro– to pre–B cell transition. Accordingly, γ1-driven B cell development is disfavored in competition with developing B cells expressing a wild-type (WT) IgH locus. However, the mutant B cells display a long half-life and accumulate in the mature B cell compartment, and even though partial truncation of the Igα cytoplasmic tail compromises their development, it does not affect their maintenance, as it does in WT cells. IgG1-expressing B cells showed an enhanced Ca²⁺ response upon B cell receptor cross-linking, which was not due to a lack of inhibition by CD22. The enhanced Ca²⁺ response was also observed in mature B cells that had been switched from IgM to IgG1 expression in vivo. Collectively, these results suggest that the γ1 chain can exert a unique signaling function that can partially replace that of the Igα/β heterodimer in B cell maintenance and may contribute to memory B cell physiology.