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  • Title: Direct regulation of Na(+)-dependent myo-inositol transport by sugars in retinal pigment epithelium: role of phorbol ester and staurosporin.
    Author: Khatami M, Cernadas M, Geroff AJ, Chandra P, Cohen MF.
    Journal: Membr Biochem; 1990; 9(4):263-77. PubMed ID: 2152143.
    Abstract:
    An Na(+)-dependent active process for myo-inositol (MI) uptake, sharing a common carrier system with glucose and sensitive to phlorizin, was previously established in primary cultures of bovine retinal pigment epithelial (RPE) cells (26, 32). The present report further examines the nature of glucose-induced inhibition of MI transport in primary cultures of RPE cells. RPE cells were grown in supplemented Dulbecco's modification of Eagle's medium (DMEM) containing 5 mM D-glucose (basic growth media) or 40 mM D-glucose or its nonmetabolizable analogue, alpha-methyl-D-glucoside (alpha MG); 1-5 mM nonradioactive MI, pyruvate, or lactate; or 0.2-20 microM phorbol 12-myristate 13-acetate (TPA) or straurosporin (modified growth media), for up to 4 weeks. The capacity of RPE cells to accumulate 3H-MI (ratios of intracellular transported radioactive MI, [MI]i, to external free MI concentration, [MI]i/[MI]o) decreased by up to 41% or 34% when cells were grown for 10 days or longer with 40 mM D-glucose or 40 mM alpha MG, respectively, compared to cells grown in basic growth media. The rate of uptake of 3H-MI also was reduced to 63 +/- 15% or 48 +/- 8% of the control values when cells were fed 1 or 5 mM nonradioactive MI, respectively. In addition, cellular capacity to bind to [3H]phlorizin was reduced to 52 +/- 7%, 61 +/- 5%, or 38 +/- 6% of the controls when RPE cells were fed 40 mM D-glucose, 40 mM alpha MG, or 5 mM nonradioactive MI, respectively. Growth media containing either pyruvate or lactate, the glucose metabolites, did not suppress the ability of RPE cells to accumulate MI. An 18 +/- 8% reduction in [3H]thymidine incorporation into DNA occurred when cells were grown in 40 mM glucose for 12-14 days, compared to cells grown with 5 mM glucose. Chronic treatment (12-14 days) of the cells with phorbol ester, an activator of protein kinase C, caused up to twofold increase in MI uptake, [3H]phlorizin binding, cell number, and DNA synthesis. However, when the rates of MI uptake into cells grown in basic growth media or TPA-treated media were normalized to cell number, no significant difference in MI uptake was found between the treated and untreated cells. Addition of staurosporin, a protein kinase C inhibitor, together with TPA, in the growth media reversed the phorbol-induced increase of MI uptake.(ABSTRACT TRUNCATED AT 400 WORDS)
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