P2X Deficiency Blocks Lesional Inflammasome Activity and Ameliorates Atherosclerosis in Mice.

BACKGROUND

Extracellular adenosine triphosphate (ATP) binds as a danger signal to purinergic receptor P2X and promotes inflammasome assembly and interleukin-1β expression. We hypothesized a functional role of the signal axis ATP-P2X in inflammasome activation and the chronic inflammation driving atherosclerosis.

METHODS

P2X-competent and P2X-deficient macrophages were isolated and stimulated with lipopolysaccharide, ATP, or both. To assess whether P2X may have a role in atherosclerosis, P2X expression was analyzed in aortic arches from low density lipoprotein receptor mice consuming a high-cholesterol or chow diet. P2X and P2X low density lipoprotein receptor mice were fed a high-cholesterol diet to investigate the functional role of P2X knockout in atherosclerosis. Human plaques were derived from carotid endarterectomy and stained against P2X.

RESULTS

Lipopolysaccharide or ATP stimulation alone did not activate caspase 1 in isolated macrophages. However, priming with lipopolysaccharide, followed by stimulation with ATP, led to an activation of caspase 1 and interleukin-1β in P2X-competent macrophages. In contrast, P2X-deficient macrophages showed no activation of caspase 1 after sequential stimulation while still expressing a basal amount of interleukin-1β. P2X receptor was higher expressed in murine atherosclerotic lesions, particularly by lesional macrophages. After 16 weeks of a high-cholesterol diet, P2X-deficient mice showed smaller atherosclerotic lesions than P2X-competent mice (0.162 cm±0.023 [n=9], P2X low density lipoprotein receptor : 0.084 cm±0.01 [n=11], =0.004) with a reduced amount of lesional macrophages. In accord with our in vitro findings, lesional caspase 1 activity was abolished in P2X mice. In addition, intravital microscopy revealed reduced leukocyte rolling and adhesion in P2X-deficient mice. Last, we observe increased P2X expression in human atherosclerotic lesions, suggesting that our findings in mice are relevant for human disease.

CONCLUSIONS

P2X deficiency resolved plaque inflammation by inhibition of lesional inflammasome activation and reduced experimental atherosclerosis. Therefore, P2X represents an interesting potential new target to combat atherosclerosis.