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KMID : 0358619960300010063
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Korean Journal of Physiology
1996 Volume.30 No. 1 p.63 ~ p.76
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Effects of Insulin and IGFs on Phosphate Uptake in Primary Cultured Rabbit Renal Proximal Tubule Cells
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Han Ho-Jae
Park Kwon-Moo
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Abstract
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The aim of present study was to characterize phosphate uptake and to investigate the mechanism for the insulin and insulin-like growth factor(IGF) stimulation of phosphate uptake in primary cultured rabbit renal proximal tubule cells. Results were as follows : 1. The primary cultured proximal tubule cells had accumulated nmole phosphate/mg protein in the presence of 140 mM NaCl and nmole phosphate/mg protein in the presence of 140 mM KCl during a 60 minute uptake period. Raising the concentration of extracellular phosphate to 100 mM induced decrease in phosphate uptake compared with that in control cells maintained in 1 mM phosphate. Optimal phosphate uptake was observed at pH 6.5 in the presence of 140 mM NaCl. Phosphate uptake at pH 7.2 and pH 7.9 decreased to of that of pH 6.5, respectively. 2. Phosphate uptake was inhibited by iodoacetic acid(IAA) or valinomycin treatment . When IAA and valinomycin were added together, phosphate uptake was inhibited to of that of control. Phosphate uptake by the primary proximal tubule cells was significantly reduced by ouabain treatment. Inhibition of protein and/or RNA synthesis by either cycloheximide or actinomycin D markedly attenuated phosphate uptake. 3. Extracellular CAMP and phorbol 12-myristate 13 acetate(PMA) decreased phosphate uptake in a dose-dependent manner in all experimental conditions. Treatment of cells with pertussis toxin or cholera toxin inhibited phosphate uptake. cAMP concentration between significantly inhibited phosphate uptake. Phosphate uptake was blocked to about 25% of that of control at 100 ng/ml PMA. 3-Isobutyl-1-methyl-xanthine(IBMX) inhibited phosphate uptake. However, in the presence of IBMX, the inhibitory effect of exogenous cAMP was not significantly potentiated. Forskolin decreased phosphate transport. Acetylsalicylic acid did not inhibit phosphate uptake. The 1,2-dioctanoyl-sn-glycorol(DAG) and 1-oleoyl-2-acetyl-sn- glycerol(OAG) showed a inhibitory effect. However, staurosporine had no effect on phosphate uptake. When PMA and staurosporine were treated together, inhibition of phosphate uptake was not observed. In conclusion, phosphate uptake is stimulated by high sodium and low phosphate and pH 6.5 in the culture medium. Membrane potential and intracellular energy levels are also an important factor fer phosphate transport. Insulin and IGF-I stimulate phosphate uptake through a mechanisms that involve do novo protein and/or RNA synthesis and decrease of intracellular cAMP level. Also protein kinase C(PKC) is may play a regulatory role in transducing the insulin and IGF-I signal for phosphate transport in primary cultured proximal tubule cells.
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KEYWORD
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Kidney, Insulin, IGF, Phosphate
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