Human CD99L2 Regulates Leukocyte Transmigration at a Unique Step between those Regulated by PECAM-1 and CD99

Rutledge, Nakisha S

doi:https://doi.org/10.21985/n2-2va4-tq55

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Human CD99L2 Regulates Leukocyte Transmigration at a Unique Step between those Regulated by PECAM-1 and CD99

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CD99-Like 2 (CD99L2) is a 52 kDa type I glycoprotein expressed on leukocytes and endothelial cells as well as other cell types in mice. It is related to CD99, although it shows only 32% sequence identity. CD99L2 has been shown to play a role in leukocyte extravasation in mice under various inflammatory conditions using anti-CD99L2 antibodies and, in one case by targeted deletion of CD99L2. We report here studies on an independently made CD99L2 "knockout mouse" that extend our knowledge of the role of CD99L2 in inflammation. CD99L2 deficiency did not affect the total or relative numbers of circulating leukocyte subsets, red blood cells, or platelets. Neither did CD99L2 deficiency affect the expression of ICAM-1, PECAM-1, or CD99 on endothelial cells. Mice lacking CD99L2 had a defective inflammatory response in the thioglycollate peritonitis model with a greater than 80% block of neutrophil infiltration and a nearly complete block in monocyte emigration into the peritoneal cavity measured 16h after the inflammatory challenge. Additionally, live imaging of inflammation in the cremaster muscle showed that mice deficient in endothelial CD99L2 had similar responses in neutrophil recruitment to the vessel compared to control. Although there was no change in the overall recruitment specifically neutrophil rolling and adhesion, there was a substantial decrease in the number of extravasation events. TEM events were decreased nearly threefold compared to control. Using the croton oil dermatitis model we show that in the absence of CD99L2, neutrophils are primarily arrested at the step of migrating across the endothelium. These data show an important functional role for CD99L2 (L2) in leukocyte TEM. However, the role of L2 in inflammation has only been studied in mice. Furthermore, the mechanism by which it regulates TEM is not known. To explore its relevance to human inflammation, we studied the role of L2 on primary human cells in vitro. Our data show that like PECAM-1 and CD99, human L2 is constitutively expressed at the borders of endothelial cells and on the surface of leukocytes. Inhibiting L2 using antibody blockade or genetic knockdown significantly reduces transmigration of human neutrophils and monocytes across endothelial cells. Furthermore, our data also show that L2 regulates a specific, sequential step of TEM between PECAM-1 and CD99, rather than operating in parallel or redundantly with these molecules. Similar to PECAM and CD99, L2 promotes transmigration by recruiting the lateral border recycling compartment (LBRC) to sites of TEM, specifically downstream of PECAM initiation. Collectively, our data identify a novel functional role for human L2 in TEM and elucidate a mechanism that is distinct from PECAM and CD99.

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