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Phenotype of the accessory cell necessary for mitogen-stimulated T and B cell responses in human peripheral blood: delineation by its sensitivity to the lysosomotropic agent, L-leucine methyl ester.
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1983
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ApoptosisImmunologyBlood CellCell DeathHuman Peripheral BloodImmunotherapyB Cell MitogensAccessory CellsHematologyAutophagyLymphocyte BiologyCell SignalingL-leucine Methyl EsterAutoimmunityCell BiologyPhagocyteAccessory CellCellular Immune ResponseMedicine
The study used the lysosomotropic compound L‑leucine methyl ester to identify the phenotype of accessory cells required for human B and T cell activation in vitro. Leu‑OMe selectively disrupts lysosomes and kills monocytes, depleting peripheral blood mononuclear cells of these accessory cells and abolishing their responsiveness to various T and B cell mitogens, a loss that can be restored by adding monocyte‑enriched adherent cells. The findings demonstrate that L‑leucine methyl ester selectively kills lysosome‑rich monocytes identified by anti‑M phi antibody 63D3, and that these monocytes are the essential accessory cells required for mitogen‑induced human B and T cell activation.
The lysosomotropic compound L-leucine methyl ester (Leu-OMe) was utilized to delineate the phenotype of the accessory cells involved in human B and T cell activation in vitro. Leu-OMe was shown to cause lysosomal disruption and selective death of human monocytes (M phi). After 30-45 minute incubations with this agent, human peripheral blood mononuclear cells (PBM) were nearly completely depleted of M phi. Associated with this M phi depletion, PBM were rendered unresponsive to a variety of T and B cell mitogens including the plant lectins phytohemagglutinin, concanavalin A, and pokeweed mitogen as well as the oxidative mitogens sodium periodate and neuraminidase plus galactose oxidase. Leu-OMe mediated loss of responsiveness was the result of a selective loss of an accessory cell necessary for each of these responses since reconstitution was accomplished by the addition of a M phi-enriched adherent cell population. While intact adherent cells could reconstitute responsiveness, crude M phi supernatants or highly purified human IL 1 alone were ineffective. Further identification of the Leu-OMe sensitive accessory cell indicated that it was entirely contained within the fraction of the adherent population identified by the monoclonal anti-M phi antibody, 63D3. The mechanism by which Leu-OMe Killed M phi was dependent on the lysosomal content of these cells, since agents that altered lysosomal enzyme activity such as chloroquine or NH4Cl protected M phi from Leu-OMe. Thus, the selective killing of M phi by Leu-OMe appeared to relate to the characteristically rich endowment of lysosomes within these cells. These results support the conclusion that a lysosome-rich, leucine methyl ester-sensitive, intact M phi identified by the monoclonal anti-M phi antibody 63D3 is the circulating accessory cell required for mitogen-triggered human B and T cell activation.