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. 1991 Aug 15;278(Pt 1):119–128. doi: 10.1042/bj2780119

Effects of macrophage colony-stimulating factor and phorbol myristate acetate on 2-D-deoxyglucose transport and superoxide production in rat peritoneal macrophages.

R J Rist 1, G E Jones 1, R J Naftalin 1
PMCID: PMC1151457  PMID: 1652936

Abstract

2-D-Deoxyglucose (2-dGlc) uptake and accumulation into rat peritoneal macrophages was increased by colony-stimulating factor (mCSF) by stimulating the coupling between endofacial hexokinase activity and the sugar transporter. The evidence for this is as follows: (1) mCSF significantly decreased the Km for zero-trans uptake (P less than 0.05), without altering Vmax.; (2) the accumulation of free 2-dGlc was increased by mCSF (P less than 0.05); (3) mCSF retarded the rate of exit of accumulated free 2-dGlc. The mCSF-dependent increase in 2-dGlc uptake by macrophages was enhanced by preincubation of the cells in mCSF-free solution. The activity of the hexose monophosphate shunt (HMPS) measured by the differential uptake of 2-d[1-3H]Glc and 2-d[2,6-3H]Glc was not stimulated by mCSF. Also, in quiescent cells, superoxide production, as determined by cytochrome c reduction, was unaffected by mCSF. Phorbol myristate acetate (PMA; 40 nM) stimulated both the HMPS activity and superoxide production. Both these effects were dependent on the uptake of external sugar (2-dGlc). Incubation of the macrophages with mCSF enhanced the sugar transport and PMA-dependent stimulation of HMPS activity and superoxide production, indicating a role for mCSF in the 'priming' of macrophage functions. Both HMPS activity and superoxide production are entirely dependent on uptake of exogenous sugar, since the potent sugar-transport inhibitor cytochalasin B competitively inhibited 2-dGlc uptake, HMPS activity and superoxide generation in PMA-activated cells (Ki approximately 0.3 microM for all three processes). Over a wide range of 2-dGlc concentrations, 4 mol of superoxide were generated/mol of 2-dGlc metabolized in the HMPS pathway, indicating coupling between these processes. The Km of 2-d[2,6-3H]Glc uptake in PMA-treated cells was 0.45 +/- 0.07 mM, and Vmax. was 1.32 +/- 0.05 mumol.min-1.ml of cell water-1. It is evident that there is a large degree of slippage between HMPS activity and membrane-associated hexokinase activity, since the Km for HMPS activity was 0.06 +/- 0.02 mM and the Vmax. was 0.10 +/- 0.03 mumol.min-1.ml of cell water-1.

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Selected References

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