Lipopolysaccharide-induced stimulation of CD11b/CD18 expression on neutrophils. Evidence of specific receptor-based response and inhibition by lipid A-based antagonists.
Gram-negative bacterial septicemia is a common clinical syndrome resulting, in part, from the activation of phagocytic leukocytes by LPS. By using flow cytometry, we have characterized LPS-induced expression of the beta 2 integrin CD11b/CD18. After exposure to Salmonella minnesota R595 LPS, expression of neutrophil CD11b/CD18 is rapidly upregulated, beginning within 5 min and achieving a peak fluorescence (typically two- to threefold over base line) by 30 min. The increase in CD11b/CD18 expression was similar in kinetics and magnitude to that produced by FMLP, PMA, and human rTNF-alpha. Concentrations of LPS necessary to stimulate a response were as low as 1 ng/ml of R595 LPS; a maximal response was observed between 30 and 100 ng/ml. The upregulation of CD11b/CD18 due to LPS was not interrupted by protein synthesis inhibitors. A group of glucosamine disaccharide lipid A-like molecules: Rhodobacter sphaeroides lipid A, lipid IVA, KDO2IVA, and deacylated LPS were able to block the stimulatory effect of LPS. This inhibition was specific for the actions of LPS as stimulation of polymorphonuclear leukocytes (PMN) by FMLP, human rTNF alpha, PMA, and rewarming were not altered by the disaccharide inhibitors. PMN which were exposed to the specific disaccharide LPS antagonists and then washed, were refractory to stimulation by LPS. The monosaccharide lipid A precursor lipid X also blocked stimulation of neutrophils by LPS, although with a 100-fold reduction in potency. Unlike the disaccharide inhibitors, PMN exposed to lipid X were still responsive to LPS stimulation after washing. The PMN response to LPS was less sensitive in the absence of serum, although upregulation of CD11b/CD18 could still be seen using higher concentrations of LPS. Monoclonal antibody directed against CD14 (clone 3C10), also specifically inhibited LPS induced PMN CD11b/CD18 expression both in the presence and absence of serum. These findings support the hypothesis that LPS stimulates neutrophils by interacting with specific cellular receptors.
Lynn
W A
WA
Department of Medicine, Maxwell Finland Laboratory for Infectious Diseases, Boston City Hospital, Boston University School of Medicine, MA 02118.
Raetz
C R
CR
Qureshi
N
N
Golenbock
D T
DT
D015703:Antigens, CD / Q000378:metabolism* / Q000502:physiology*; D015214:Antigens, Differentiation, Myelomonocytic / Q000502:physiology*; D018821:CD18 Antigens; D016895:Culture Media, Serum-Free; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D004187:Disaccharides / Q000494:pharmacology; D006017:Glycolipids / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008050:Lipid A / Q000031:analogs & derivatives / Q000737:chemistry / Q000494:pharmacology*; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000737:chemistry*; D016177:Macrophage-1 Antigen / Q000378:metabolism*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D015854:Up-Regulation
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D018821:CD18 Antigens; D016895:Culture Media, Serum-Free; D004187:Disaccharides; D006017:Glycolipids; D008050:Lipid A; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D016177:Macrophage-1 Antigen; D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D003907:Dexamethasone; C044283:lipid X; D003513:Cycloheximide