Adenosine and TNF-alpha exert similar inotropic effect on heart cultures, suggesting a cardioprotective mechanism against hypoxia

摘要

When cardiomyocytes were subjected to hypoxia, tumor necrosis factor-a (TNF-alpha; 3-50 ng/ml) or adenosine (1-100 mu M), decreased hypoxic damage as was detected by lactate dehydrogenase (LDH) release, MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) absorbance, ROS (reactive oxygen species) measurement or desmin immunostaining. This cardioprotection was not prevented in TNF-alpha-treated cultures by 5-hydroxydecanoic acid (5-HD). Our aim was to elucidate whether adenosine and TNF-a mediate a similar protective mechanism against hypoxia in primary heart cultures and in H9c2 cardiomyocytes. Adenosine and TNF-alpha are known for their negative inotropic effects on the heart. We have suggested that deoxyglucose uptake reflects heart contractility in cell cultures; therefore, we assayed its accumulation under various conditions. Treatment for 20 min with adenosine, R-PIA [(-)-N(6)-phenylisopropyladenosine] (10 mu M), or TNF-a reduced H-3-deoxyglucose uptake in primary heart cultures and also in H9c2 cardiomyocytes by 30-50%. Isoproterenol accelerated 3H-deoxyglucose uptake by 50%. Adenosine, R-PIA, or TNF-a attenuated the stimulatory effect of isoproterenol on 3H-deoxyglucose uptake to control levels. Hypoxia reduced 3H-deoxyglucose uptake by 50%, as in the treatment of the hypoxic cultures with TNF-alpha or adenosine. Glibenclamide (2 PM), 5-HD (300 mu M), or diazoxide (50 mu M) increased 3 H-deoxyglucose uptake by 50-80%. Adenosine (100 mu M) and TNF-a (50 ng/ml) stimulated Rb-86 efflux. Glibenclamide attenuated this effect. We demonstrate that TNF-alpha, like adenosine, accelerated Ca2+ uptake into the sarcoplasmic reticulum (SR) by 50-100% and therefore prevented cardiomyocyte Ca2+ overload. Our findings further suggest that TNF-alpha, as well as adenosine, may mediate an adaptive effect in the heart by preventing Ca2+ overload via activation of SR Ca-ATPase (SERCA(2)a). (c) 2007 Elsevier Inc. All rights reserved.