The expression of adenosine receptors mRNA in human leukocytes. A study on patients with advanced valvular disease undergoing cardiac surgery

Purpose: Adenosine is a retaliatory metabolite acting as a potent anti-inflammatory autacoid; it is defined as a "danger signal" for cells and organs. Extracellular ATP is generally released in conditions of injury or stress and it is dephosphorylated to AMP by the ectonuclease triphosphate diphosphohydrolase (CD39), followed by degradation to adenosine by the ecto-5"-nucleotidase (CD73). Adenosine mediates its cellular functions by four specific transmembrane receptors (A1R, A2aR, A2bR and A3R). In different animal models an up-regulation of specific adenosine receptors (ARs) has been documented in cardiac tissue during heart failure (HF). Recently ARs mRNA expression has been assessed in human whole blood. The aim of this study was to study the possible changes of mRNA expression profile of A1R, A2aR, A2bR and A3R in human leukocytes of patients (pts) with valvular HF (Vlv) as compared to healthy subjects (C). Methods. Total RNA was extracted from leukocytes of C (n=7) and of Vlv pts before cardiac surgery (n= 6, NYHA III-IV) with PAXgene Blood RNA Kit (Qiagen, Milan, Italy). These extracts were retrotranscripted to cDNA, then Real time PCR analysis was performed and optimized for each ARs primer. CD39 and CD73 mRNA expression was also evaluated both in C and in Vlv pts. The experimental results were normalized with the three most stably expressed genes (SRP14, EEF1A, RPL13A). Results. In human whole blood of Vlv pts significantly higher levels of mRNA expression were observed in each receptor with respect to C (A1R: C=1.97±0.72, Vlv pts=10.9±3.27, p=0.0098; A2aR: C=0.45±0.15, Vlv pts=3.19±0.94, p=0.0057; A2bR: C=0.43±0.16, Vlv pts=5.00±2.98, p=0.05; A3R: C=1.39±0.26, Vlv pts=2.9±0.71, p=0.04). An upregulation of CD39 and CD73 was also found in Vlvs pts. Significant correlations (p<0.05) were reported between all AR themselves as well as with CD39 and CD73. Conclusions. An increase of AR mRNA expression in the peripheral circulating cells of Vlv pts undergoing cardiac surgery was documented, suggesting a protective CD39-CD73-dependent adenosine production in Vlv pts in advanced NYHA classes. The approach on human leucocites allows the assessment of ARs transcriptomic profiling in HF; a potential application of this method for monitoring the response to treatment can be foreseen in HF, a pathology still burdened by high mortality and morbidity.