Deficiency of leukocyte adhesion molecules

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Study of patients congenitally deficient in expression of the CD11/CD18 dimer has clarified the fundamental role of these transmembrane glycoproteins of the integrin family in multiple functions of polymorphonuclear leukocytes (PMNL). Deficient expression is associated with severe deficiency of PMNL aggregation, adhesion to protein-coated surfaces and endothelial monolayers, locomotion, chemotaxis, phagocytosis and bacterial cell killing. Severely affected patients develop leukocytosis and die in infancy of overwhelming infection. A patient we studied at autopsy died from septicemia and pneumonia. There was striking lack of any acute inflammatory response near sites of necrosis and infection. In the lungs, in contrast, a distinct but deficient acute focal inflammatory response was observed.We used colloidal gold immunolabeling to study the subcellular localization of the CD11/CD18 adhesion molecules in human PMNL and their expression and movement on cell surfaces after stimulation with the chemotactic peptide formyl-met-leu-phe (fMLP). Human PMNL in pyrogen-free PBS were fixed for lh in 1% glutaraldehyde, dehydrated to 70% ethanol and embedded in LR White resin at 55°C. A polyclonal rabbit antiserum recognizing CD18 and absorbed with CD18-deficient PMNL was used to label sites of storage in thin sections, using 5-nm goat-anti-rabbit-IgG gold. We compared sites labeling for CD18, lactoferrin and gelatinase in adjacent thin sections. For surface labeling, cells were fixed in 1% glutaraldehyde for lh, exposed to OKM1 monoclonal antibody, washed and labeled using 36-nm rabbit-anti-mouse-IgG-gold. Controls for both studies included substitution of other or nonspecific primary and secondary antibody. Cells were attached to polylysine-coated coverslips, critical-point dried in CO2, carbon coated and examined by scanning EM using backscattered electron imaging. Other cells were exposed to f(ab’)2 fragments of OKMl and to f (ab’)2-rabbit-anti-mouse prior to fixation, followed by labeling with 36-nm goat-anti-rabbit-gold, or exposed to f(ab’)2-OKMl conjugated to 36-nm gold particles prior to fixation.