title: Time-dependent loss of Mac-1 from infiltrating neutrophils in the reperfused myocardium.
abstract: Numerous studies have shown that polymorphonuclear neutrophils (PMNs) infiltrate the myocardium immediately after reperfusion of infarcted tissue. Studies with mAbs in vivo and cellular studies in vitro suggest that PMN-induced injury of the cardiac myocyte involve Mac-1 adhesion to myocyte ICAM-1. In this study we demonstrate that PMNs that have infiltrated the ischemic area begin to lose Mac-1 within the first 3 h. By the fifth hour of reperfusion, minimal CD11b staining is seen on PMNs using immunostaining, whereas CD11a remained unchanged. Immunoreactivity of postreperfusion cardiac lymph with R15.7 (anti-CD18) or MY904 (anti-CD11b) was positive in all animals but not for CD11a (R7.1), indicating a specific loss of Mac-1. Immunoprecipitation with either R15.7 or MY904 resulted in identical peptides (a doublet at 190 kDa and a band at 80 kDa), suggesting that both alpha and beta subunits of Mac-1 heterodimer were released. Immunoprecipitation of control PMN lysates revealed bands of 198 kDa and 91 kDa slightly greater than those from the released Mac-1. An in vitro model of homotypic aggregation showed a similar loss of Mac-1 from PMNs; immunoprecipitates of the supernatant demonstrated peptide bands identical with those found in postischemic cardiac lymph. The appearance of soluble Mac-1 in vitro was prevented by anti-CD18 mAb, R15.7, and also by protease inhibition by PMSF. Thus, in vivo and in vitro, activated PMNs lose Mac-1 in a process that may be dependent upon adhesion and subsequent proteolysis.
authors.affiliation: Section of Cardiovascular Sciences, DeBakey Heart Center and Department of Medicine, Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: K A
authors.initials: KA
authors.lastname: Youker
authors.forename: J
authors.initials: J
authors.lastname: Beirne
authors.forename: J
authors.initials: J
authors.lastname: Lee
authors.forename: L H
authors.initials: LH
authors.lastname: Michael
authors.forename: C W
authors.initials: CW
authors.lastname: Smith
authors.forename: M L
authors.initials: ML
authors.lastname: Entman
mesh_terms: D000818:Animals; D002465:Cell Movement / Q000276:immunology*; D002474:Cell-Free System / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D004285:Dogs; D005260:Female; D008196:Lymph / Q000737:chemistry / Q000276:immunology; D016177:Macrophage-1 Antigen / Q000032:analysis / Q000378:metabolism*; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000276:immunology* / Q000378:metabolism* / Q000473:pathology; D009206:Myocardium / Q000737:chemistry / Q000276:immunology; D009504:Neutrophils / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis / Q000276:immunology; D011233:Precipitin Tests; D013194:Staining and Labeling; D013997:Time Factors
chemical_list: D016177:Macrophage-1 Antigen; D010446:Peptide Fragments