Glu496 to Ala polymorphism in the P2X7 receptor impairs ATP-induced IL-1 beta release from human monocytes.
Priming of monocytes with LPS produces large quantities of intracellular, biologically inactive IL-1beta that can be processed and released by subsequent activation of the P2X7 receptor by extracellular ATP. We examined whether a loss-of-function polymorphism of the human P2X7 receptor (Glu496Ala) impairs this process. Both ATP-induced ethidium+ uptake and ATP-induced shedding of L-selectin (CD62L) were nearly absent in monocytes from four subjects homozygous for Glu496Ala confirming that this polymorphism impairs P2X7 function. The level of ATP-induced IL-1beta released in 2 h from LPS-activated whole blood from homozygous subjects was 50% of that from wild-type samples. A more marked defect in IL-1beta release was observed from LPS-activated monocytes of homozygous subjects which was only 22% of that released from wild-type monocytes after a 30-min incubation with ATP. However, after a 60-min incubation with ATP, the amount of IL-1beta released from homozygous monocytes was 70% of that released from wild-type monocytes. Incubation of monocytes of either genotype with nigericin resulted in a similar release of IL-1beta. Western blotting demonstrated that ATP induced the release of mature 17-kDa IL-1beta from monocytes, and confirmed that this process was impaired in homozygous monocytes. Finally, ATP-induced 86Rb+ efflux was 9-fold lower from homozygous monocytes than from wild-type monocytes. The results indicate that ATP-induced release of IL-1beta is slower in monocytes from subjects homozygous for the Glu496Ala polymorphism in the P2X7 receptor and that this reduced rate of IL-1beta release is associated with a lower ATP-induced K+ efflux.
Sluyter
Ronald
R
Department of Medicine, University of Sydney at Nepean Hospital, Penrith, New South Wales, Australia. rons@med.usyd.edu.au
Shemon
Anne N
AN
Wiley
James S
JS
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000409:Alanine / Q000235:genetics*; D002478:Cells, Cultured; D004996:Ethidium / Q000037:antagonists & inhibitors / Q000378:metabolism; D018698:Glutamic Acid / Q000235:genetics*; D006720:Homozygote; D006801:Humans; D007375:Interleukin-1 / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D019041:L-Selectin / Q000096:biosynthesis / Q000378:metabolism; D009000:Monocytes / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D011110:Polymorphism, Genetic / Q000276:immunology; D011499:Protein Processing, Post-Translational / Q000276:immunology; D018048:Receptors, Purinergic P2 / Q000235:genetics* / Q000502:physiology; D058486:Receptors, Purinergic P2X7; D012414:Rubidium Radioisotopes / Q000378:metabolism
D007375:Interleukin-1; C546468:P2RX7 protein, human; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D012414:Rubidium Radioisotopes; D019041:L-Selectin; D018698:Glutamic Acid; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D004996:Ethidium; D000409:Alanine