Negative modulation of human NK cell activity by purinoceptors. 1. Effect of exogenous adenosine triphosphate.
A variety of receptors, including purinergic receptors (for adenosine and adenosine nucleotides), are present on lymphoid cells. We have investigated the negative modulatory action of ATP, a high-energy purinergic compound, on the human peripheral blood natural killer (NK) cells. Micromolar doses of exogenous ATP produced a strong, direct, reversible, and dose-dependent (2.5 x 10(-5) to 8 x 10(-4) M) inhibition of NK cell-mediated cytotoxic activity in 98% of the 131 healthy donors tested. The inhibitory effect of ATP could be demonstrated in unseparated total lymphocytes and enriched NK cells (Percoll density gradient or negative selection by complement lysis) in 2- or 16-hr NK assays, at various effector:target ratios, at suboptimal and saturating concentrations of the K562 targets. The inhibitory constants ID20 and ID50 max for ATP (2 hr), which were 5.6 x 10(-5) and 8 x 10(-5) M, respectively, appear to be a relatively stable feature of an individual. The mechanism of action of ATP was also explored. It is not due to a simple chelation by ATP of the divalent cations Mg2+ and Ca2+ which play a role in the binding/lytic reactions during NK cell-mediated oncolysis. ATP treatment of lymphocytes neither nonspecifically decreased the number of viable cells nor specifically eliminated the NK cells, including their subsets, CD57+16-, CD57+16+, and CD57-16+ as revealed by immunofluorescence staining. Use of an enzyme-like kinetic approach to define the nature of ATP-induced inhibition suggests that it may be an "uncompetitive" type (Lineweaver-Burke reciprocal plot), because both Vmax (maximal velocity of oncolysis) and Km (Michaelis constant) were reduced simultaneously from 1.53 x 10(4) and 2.64 x 10(4) to 0.44 x 10(4) and 0.73 x 10(4), respectively, in presence of ATP. Other methods (Eadie-Hofstee's and Hane's equations) confirmed that ATP does elicit a qualitatively and quantitatively similar shift in these kinetic parameters. Finally, an 18-hr preactivation of effectors by interleukin 2 (IL2) resulted in partial protection from the negative modulation of NK activity by ATP, while the presence of ATP during preincubation did not interfere with the desensitizing effect of IL2. The biological relevance of these findings are discussed and it is concluded that by interacting with appropriate (non-FcRIII?) cell-surface binding sites such as P2 purinergic receptors, ATP is capable of negatively modulating the antitumor cytotoxic activity of human NK cells. This down-regulation can be partially overcome by IL2 that up-regulates NK cells.(ABSTRACT TRUNCATED AT 400 WORDS)
Krishnaraj
R
R
Geriatric Medicine (M/C 787), College of Medicine, University of Illinois, Chicago 60612.
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000328:Adult; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000187:drug effects*; D007700:Kinetics; D008875:Middle Aged; D011941:Receptors, Adrenergic / Q000187:drug effects*; D014407:Tumor Cells, Cultured
D007376:Interleukin-2; D011941:Receptors, Adrenergic; D000255:Adenosine Triphosphate