Lipopolysaccharide and lipid A‐induced human blood lymphocyte activation as detected by a protein A plaque assay

Abstract

Various purified cell wall lipopolysaccharides (LPS) from gram-negative bacteria and derivatives of these LPS were tested for their stimulatory capacity for human peripheral blood cells. Immunoglobulin (Ig) production was tested by an indirect plaque-forming cell assay using Staphylococcus aureus protein A-coupled erythrocytes and specific anti-Ig as developing serum. This method allows the detection of the majority of cells secreting Ig of a single class, and the number of plaque-forming cells detected are approximately 100-1000 times the amount obtained using normal sheep red cells as targets. LPS containing the O antigen-specific chain, as well as mutant products only containing lipid A and ketodeoxyoctonate trisaccharide, could induce cell division and antibody synthesis. The polypeptide antibiotic polymyxin B was found to inhibit LPS-induced activation. Furthermore, purified lipid A, complexed with bovine serum albumin, was also found to activate human peripheral blood B cells. These findings demonstrate that human peripheral blood lymphocytes can be activated by LPS and also indicate that lipid A is the active part of these molecules.

Flow Cytometric Analysis of Peripheral Blood Mononuclear Cells Induced by Experimental Endotoxemia in Horse

“…The percentage of B-lymphocytes (IgM + cells) was significantly increased among the PBMCs population, suggesting that a humoral response might develop. This result corresponded with past reports, in which LPS activated human peripheral blood B cells [3,29]. Moreover, the percentage of MHC class II + cells appeared to be altered specifically in equine endotoxemia, indicating important implications for immunologic and host defense mechanism.…”

Characterization of Human Lysozyme (Muramidase)-Releasing Cells

“…Thus, unless various purifica tion methods are used it must be born in mind that cultures normally thought to contain peripheral blood lymphocytes also contain other cell types. The tissue culture system employed in these experiments was optimal for Ig secretion [3,16,17,31]. Thus, it is probable that the use of a culture procedure deve loped for the maintenance of macrophages and mo nocytes [35] would have resulted in higher PFC values using antimuramidase Ig as a developing agent.…”

“…A similar kinetics for Ig secretion was recorded in control cultures containing purified protein deriva tive of tuberculin. The peak for Ig secretion using this and other mitogens in this culture system has been re ported to occur after approximately I week of culture [3,16,17,31]. AB serum and fetal calf serum were used as supplements and to some cultures the polyan ions dextran sulfate or sodium polyanethole sulfonate were added.…”

“…The effect of various lymphocyte-stimulatory agents such as phytohemagglutinin, pokeweed mito gen, purified protein derivative and Epstein-Barr vi rus, which have all been shown to activate human PBC to Ig secretion [3,16,17,31] was also tested. Since Epstein-Barr virus and C3 receptors have been shown to be associated [19,20], and since macro phages also carry C3 receptors [2], we found it rele vant to study this agent.…”

“…The tissue culture system employed was the one found optimal for immunoglobulin secretion detected as PFC in the protein A plaque assay [3,16,17,31]. AB serum of fe tal calf serum in a final concentration of 10% was used as supplement.…”