CHARACTERIZATION OF RAT LECAM-1 (L-SELECTIN) BY THE USE OF MONOCLONAL-ANTIBODIES AND EVIDENCE FOR THE PRESENCE OF SOLUBLE LECAM-1 IN RAT SERA

We have characterized the rat LECAM-1 (L-selectin) by the use of newly generated hamster anti-rat LECAM-1 monoclonal antibodies (mAb) (HRL1, HRL2, HRL3, HRL4), with respect to the biochemistry, cellular distrib ution and function, and developed an ELISA system to detect the solubl e form of rat LECAM-1. In the rat, lymphocyte and neutrophil LECAM-1 h ave apparent molecular masses of 65 and 62 kDa, respectively, and diff erential glycosylation may account for the molecular heterogeneity. Re adily detectable levels of LECAM-1 are expressed on peripheral blood l ymphocytes and neutrophils, but not on thymocytes. Lymphocyte LECAM-1 is rapidly shed from the cell surface upon cell activation with PMA, b ut not with interleukin (IL)-8. In contrast, neutrophil LECAM-1 showed rapid shedding upon stimulation with phorbol 12-myristate 13-acetate (PMA) or IL-8. Concomitantly there is up-regulated expression of Mac-1 in PMA- and IL-8-stimulated neutrophils. Neutrophil rolling in mesent eric venules was significantly inhibited by administration of function -blocking anti-rat LECAM-1 mAb HRL3, but not by non-blocking HRL4, ind icating that LECAM-1 plays a significant role in leukocyte rolling. Gi ven that LECAM-1 is rapidly shed from the cell surface, we attempted t o develop an ELISA system for detecting LECAM-I is soluble form, and m easured the levels in experimental autoimmune uveitis. The circulating levels of LECAM-1 increased from day 4, which preceded the appearance of clinical signs of uveitis and remained high until uveitis subsided , suggesting that soluble LECAM-1 is potentially a useful parameter to monitor certain types of inflammatory or immune disorders.