Adenosine Diphosphate Stimulation of Cultured Hematopoietic Cell Lines

• Published in: Blood 1993-05, Vol.81 (10), p.2652-2657
• Main Authors: Kalambakas, Stacey A., Robertson, Fredika M., O’Connell, Sean M., Sinha, Shini, Vishnupad, Kalpana, Karp, George I.
• Format: Article
• Language: English
• Subjects: Adenosine Diphosphate - metabolism; Adenosine Diphosphate - pharmacology; Adenosine Triphosphate - pharmacology; Animals; Biological and medical sciences; Blood coagulation. Blood cells; Calcium - metabolism; Cells, Cultured; Collagen - pharmacology; Epinephrine - pharmacology; Fundamental and applied biological sciences. Psychology; General aspects, investigation methods, hemostasis, fibrinolysis; Humans; Ionomycin - pharmacology; Kinetics; Leukemia, Erythroblastic, Acute; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Lymphoma, Large B-Cell, Diffuse; Megakaryocytes - drug effects; Megakaryocytes - metabolism; Molecular and cellular biology; Rats; Receptors, Purinergic - drug effects; Receptors, Purinergic - metabolism; Thrombin - pharmacology; Tumor Cells, Cultured
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V81.10.2652.2652

Adenosine diphosphate (ADP) plays a critical role in platelet activation both by exogenous stimulation and the release of endogenous intracellular stores. As the platelet ADP receptor is not well defined, we have chosen to identify and characterize several cell lines that possess functional receptors for this nucleotide. Rat promegakaryoblasts (RPM), human erythroleukemia cells (HEL), U937, and K562 leukemia cells responded to ADP, as measured by a rapid increase in intracellular calcium. In the case of RPM cells, ADP was the only naturally occurring platelet agonist capable of eliciting this response. Binding studies with [3H]ADP and fixed cells showed 3.99 ± 1.77 × 105 binding sites/cell for RPM cells (apparent dissociation constant [kd] = 7.75 ± 2.3 × 10–8 mol/L), 8.19 ± 3.25 × 105 sites/cell for HEL cells (kd = 2.15 ± 0.84 × 10–7 mol/L), 1.15 ± 0.23 × 106 sites/cell for U937 cells (kd = 2.20 ± 0.53 × 10–7 mol/L) and 5.39 ± 2.80 × 105 sites/cell for K562 cells (kd = 1.37 ± 0.39 × 10–7 mol/L). Inhibition studies with unlabeled nucleotides and analogues showed that binding was approximately 85% specific and the inhibitory pattern was similar to that seen with mature platelets. The purine base adenosine resulted in little or no inhibition. These studies indicate that both human and rat hematopoietic cell lines possess intact ADP receptors and may be useful tools in future studies of the structure and function of this important platelet-activation system.