Time-dependent loss of Mac-1 from infiltrating neutrophils in the reperfused myocardium

Keith A. Youker, Joshua Beirne, Justin Lee, Lloyd H. Michael, C. Wayne Smith, Mark L. Entman

Research output: Contribution to journalArticle

17 Scopus citations

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 α and β 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.

Original languageEnglish (US)
Pages (from-to)2752-2758
Number of pages7
JournalJournal of Immunology
Volume164
Issue number5
DOIs
StatePublished - Mar 1 2000

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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