GM-CSF triggers a rapid, glucose dependent extracellular acidification by TF-1 cells: evidence for sodium/proton antiporter and PKC mediated activation of acid production

• Published in: Journal of cellular physiology Vol. 154; no. 1; p. 129
• Main Authors: Wada, H G, Indelicato, S R, Meyer, L, Kitamura, T, Miyajima, A, Kirk, G, Muir, V C, Parce, J W
• Format: Journal Article
• Language: English
• Published: United States 01.01.1993
• Subjects: Benzoquinones; Bone Marrow - metabolism; Bone Marrow Cells; Carrier Proteins - metabolism; Cell Division; Cell Line; Ethers, Cyclic - pharmacology; Genistein; Glucose - metabolism; Granulocyte-Macrophage Colony-Stimulating Factor - physiology; Humans; Hydrogen-Ion Concentration; Ionomycin - pharmacology; Isoflavones - pharmacology; Kinetics; Lactams, Macrocyclic; Naphthalenes; Okadaic Acid; Phosphoprotein Phosphatases - antagonists & inhibitors; Polycyclic Compounds - pharmacology; Protein Kinase C - antagonists & inhibitors; Protein Kinase C - metabolism; Protein-Tyrosine Kinases - antagonists & inhibitors; Quinones - pharmacology; Rifabutin - analogs & derivatives; Signal Transduction; Sodium-Hydrogen Exchangers; Tetradecanoylphorbol Acetate - pharmacology
• ISSN: 0021-9541
• DOI: 10.1002/jcp.1041540116

The extracellular acidification rate of the human bone marrow cell line, TF-1, increases rapidly in response to a bolus of recombinant granulocyte-macrophage colony stimulating factor (GM-CSF). Extracellular acidification rates were measured using a silicon microphysiometer. This instrument contains micro-flow chambers equipped with potentiometric sensors to monitor pH. The cells are immobilized in a fibrin clot sandwiched between two porous polycarbonate membranes. The membranes are part of a disposable plastic "cell capsule" that fits into the microphysiometer flow chamber. The GM-CSF activated acidification burst is dose dependent and can be neutralized by pretreating the cytokine with anti-GM-CSF antibody. The acidification burst can be resolved kinetically into at least two components. A rapid component of the burst is due to activation of the sodium/proton antiporter as evidenced by its elimination in sodium-free medium and in the presence of amiloride. A slower component of the GM-CSF response is a consequence of increased glycolytic metabolism as demonstrated by its dependence on D-glucose as a medium nutrient. Okadaic acid (a phospho-serine/threonine phosphatase inhibitor), phorbol 12-myristate 13-acetate (PMA, a protein kinase C (PKC) activator), and ionomycin (a calcium ionophore) all produce metabolic bursts in TF-1 cells similar to the GM-CSF response. Pretreatment of TF-1 cells with PMA for 18 h resulted in loss of the GM-CSF acidification response. Although this treatment is reported to destroy protein kinase activity, we demonstrate here that it also down-regulates expression of high-affinity GM-CSF receptors on the surface of TF-1 cells. In addition, GM-CSF driven TF-1 cell proliferation was decreased after the 18 h PMA treatment. Short-term treatment with PMA (1-2 h) again resulted in loss of the GM-CSF acidification response, but without a decrease in expression of high-affinity GM-CSF receptors. Evidence for involvement of PKC in GM-CSF signal transduction was obtained using calphostin C, a specific inhibitor of PKC, which inhibited the GM-CSF metabolic burst at a subtoxic concentration. Genistein and herbimycin A, tyrosine kinase inhibitors, both inhibited the GM-CSF response of TF-1 cells, but only at levels high enough to also inhibit stimulation by PMA. These results indicate that GM-CSF activated extracellular acidification of TF-1 cells is caused by increases in sodium/proton antiporter activity and glycolysis, through protein kinase signalling pathways which can be both activated and down-regulated by PMA.