Gene expression profiling for the identification of G-protein coupled receptors in human platelets

Introduction and materials and methodsG-protein coupled receptors (GPCRs) play an important role in platelet aggregation. To identify new platelet GPCRs, a platelet gene expression profile was generated and validated using quantitative real-time PCR.ResultsIn total, mRNA of 28 GPCR genes was detected in human platelets. The 12 most abundant platelet GPCR transcripts were: thrombin receptor PAR1 (1865 ± 178%), ADP receptor P2Y12 (459 ± 88%), succinate receptor 1 (257 ± 48%), ADP receptor P2Y1 (100%), orphan P2RY10 (68.2 ± 3.3%), lysophosphatidic acid receptors GPR23 (48.2 ± 11%) and GPR92 (26.1 ± 3.3%), adrenergic receptor α2A (18.4 ± 4.4%), orphan EBI2 (6.31 ±0.42), adenosine receptors A2A (2.94 ± 0.24%) and A2B (2.88 ± 0.16%) and lysophosphatidic acid receptor LPA1 (2.59 ± 0.39%) (% relative to the chosen calibrator P2Y1).A surprising G-protein coupled receptor redundancy was found: two ADP receptors (P2Y1 and P2Y12), three adenosine receptors (A2A, A2B, and A1), four lysophosphatidic acid receptors (LPA1, LPA3, GPR23 and GPR92), two l-glutamate receptors (mGlu3 and mGlu4) and two serotonin receptors (5-HT1F and 5-HT4).The adenosine receptor A2B gene expression was validated with protein expression and functional studies. Western blot confirmed A2B receptor protein expression and platelet flow cytometry demonstrated inhibition of the effect of NECA by the adenosine A2B-antagonist MRS1754.ConclusionsWe have detected several GPCRs not previously known to be expressed in platelets, including a functional adenosine A2B receptor. The findings could improve our understanding of platelet aggregation and provide new targets for drug development.