Extracellular nucleotides stimulate leukocyte adherence to cultured pulmonary artery endothelial cells

Adenosine, ATP, and various nucleotides were examined for their effects on the adherence of leukocytes to bovine pulmonary artery endothelial cells. Extracellular ATP enhanced adherence of HL-60 cells and human neutrophils to endothelial cells in a dose-dependent fashion. Maximal adherence occurred after 15 min coincubation of ATP and HL-60 cells or neutrophils with endothelial cells. ATP stimulation was mediated by direct effects on both HL-60 cells and endothelial cells. The potency profile of various nucleotides was ATP = 2-MeSATP > beta,gamma-CH2ATP, indicative of a P2y receptor. Interestingly, UTP was as potent as ATP in stimulating HL-60 cell adherence, suggesting the presence of a pyrimidine nucleotide receptor. Photoaffinity labeling of endothelial cells with 8-Az-[alpha-32P]ATP showed the presence of two ATP binding proteins of 48 and 87 kDa. ATP and 2-MeSATP inhibited binding by both proteins. Labeling of the 87-kDa protein was inhibited by beta,gamma-CH2ATP, whereas UTP blocked binding by the 48-kDa protein. Thus photoaffinity labeling experiments support the proposal that endothelial cells possess two ATP receptors, one of which is a P2u nucleotide receptor. These findings show that extracellular nucleotides enhance leukocyte adherence to endothelial cells. Nucleotide release into the extracellular space may be one mechanism of exacerbating vascular cell injury relevant to conditions such as adult respiratory distress syndrome and septic shock.