Cystine Accumulation Impairs Glucose Transport in LLC-PK&lt;sub&gt;1&lt;/sub&gt; Cells

Bennun, Alfred; Bashan, N; Potashnik, R.; Moran, Arie

doi:10.1159/000154617

Cited by 4 publications

(9 citation statements)

References 13 publications

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“…Inhibition of glucose transport was also shown in LLC-PK 1 cells (15). Corresponding to the decreased glucose uptake, cystine loading resulted in a reduced number of glucose transporters on the apical membrane of LLC-PK 1 cells (15) and in brush border membrane vesicles from rats after intraperitoneal CDME injections (16). Salmon et al (13) used the CDME model to show that reduction in proximal tubular transport was due to an inhibition of active transport but not to an increase in proximal tubular permeability, as it was previously supposed for the Fanconi syndrome caused by maleic acid (17).…”

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confidence: 80%

“…So far with this model uptake measurements with radiolabeled solutes were performed in rat (10,18,31,34) and rabbit (11,13,14,32,33) proximal tubules and with cultured porcine LLC-PK 1 cells (15,19,30). These studies showed an inhibition of volume absorption (13,14), reduction of the transepithelial potential difference (13), and inhibition of transport of different solutes (7,10,(13)(14)(15). Most of proximal tubular transport is Na ϩ -coupled; depending therefore on the Na ϩ -gradient from the extrato the intracellular space, the role of energy metabolism, Na ϩ / K ϩ -ATPase activity, intracellular phosphate, and ATP levels in this experimental model have been investigated (14,18,31,33,34).…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, inhibition of glucose, different amino acids, bicarbonate, and phosphate reabsorption in cystine-loaded proximal tubules was demonstrated (7,13). Inhibition of glucose transport was also shown in LLC-PK 1 cells (15). Corresponding to the decreased glucose uptake, cystine loading resulted in a reduced number of glucose transporters on the apical membrane of LLC-PK 1 cells (15) and in brush border membrane vesicles from rats after intraperitoneal CDME injections (16).…”

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confidence: 92%

“…Sakarcan et al (11) showed that cystine in this model is mainly accumulated intralysosomally. So far, uptake measurements of radiolabeled substances by rat and rabbit proximal tubules and by porcine proximal tubular cells (LLC-PK 1 ) have been used to obtain insights into the pathophysiologic mechanisms of the Fanconi syndrome of cystinosis (7,10,(13)(14)(15). Inhibition of volume reabsorption and a decrease in the lumen-negative transepithelial potential difference were shown under these conditions (13,14).…”

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confidence: 99%

“…Most of proximal tubular transport of solutes across the luminal membrane is secondarily coupled with active Na ϩ -transport; studies were therefore performed to investigate the relationship between intracellular cystine accumulation and energy metabolism. Na ϩ /K ϩ -ATPase was not directly inhibited by CDME loading (14,18), but a decrease in intracellular phosphate and ATP-levels of cystine-loaded tubules with subsequent inhibition of Na ϩ /K ϩ -ATPase activity could be shown (14,15,18). According to these findings, there was an increase in intracellular Na ϩ -concentration parallel to a decrease in intracellular K ϩ -concentration in LLC-PK 1 cells (19) and in rat proximal tubules (18).…”

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confidence: 99%

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Inhibition of Na+-Dependent Transporters in Cystine-Loaded Human Renal Cells

Çetinkaya¹,

Schlatter²,

Hirsch³

et al. 2002

Journal of the American Society of Nephrology

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Abstract. Cystinosis is the most common cause of the renal Fanconi syndrome in children, leading to severe electrolyte disturbances and growth failure. A defective lysosomal transporter, cystinosin, results in intralysosomal accumulation of cystine. Loading cells with cystine dimethyl ester (CDME) is the only available model for this disease. This model was used to present electrophysiologic studies on immortalized human kidney epithelial (IHKE-1) cells that had been derived from the proximal tubule with the slow whole-cell patch clamp technique. Basal membrane voltages (V m ) of IHKE-1 cells were Ϫ30.7 Ϯ 0.4 mV (n ϭ 151). CDME concentration-dependently altered V m with an initial depolarization (2.7 Ϯ 0.2 mV; n ϭ 76; 1 mM CDME) followed by a more pronounced hyperpolarization (Ϫ9.9 Ϯ 1.0 mV; n ϭ 49). Three Na ϩ -dependent transporters were examined. Alanine (1 mM) depolarized IHKE-1 cells by 17.6 Ϯ 0.7 mV (n ϭ 59), and phosphate (1.8 mM) depolarized by 9.7 Ϯ 1.1 mV (n ϭ 18).Acidification of IHKE-1 cells with propionate (20 mM) resulted in a depolarization of V m by 7.1 Ϯ 0.3 mV (n ϭ 21) followed by a repolarization by 2.9 Ϯ 0.3 mV/min (n ϭ 17), reflecting Na ϩ /H ϩ -exchanger activity. Acute addition of 1 mM CDME did not alter the alanine-and propionate-induced changes in V m , but it reduced the phosphate-induced depolarization by 37 Ϯ 9% (n ϭ 10). Incubation with 1 mM CDME reduced the activity of all three transporters. Depolarizations by alanine and phosphate and the repolarization after propionate were inhibited by 57 Ϯ 4% (n ϭ 30), 45 Ϯ 9% (n ϭ 9), and 78 Ϯ 15% (n ϭ 8), respectively. In conclusion, this study demonstrates that CDME acutely alters V m of IHKE-1 cells and that at least three Na ϩ -dependent transporters are inhibited, the Na ϩ -phosphate cotransporter most sensitively. This might suggest that phosphate depletion and dissipation of the Na ϩ -gradient are involved in the development of the Fanconi syndrome of cystinosis.

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Section: Discussionmentioning

confidence: 99%

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confidence: 92%

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confidence: 99%

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confidence: 99%

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Inhibition of Na+-Dependent Transporters in Cystine-Loaded Human Renal Cells

Çetinkaya¹,

Schlatter²,

Hirsch³

et al. 2002

Journal of the American Society of Nephrology

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The Fanconi syndrome of cystinosis: insights into the pathophysiology

Baum

1998

Pediatric Nephrology

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Cystinosis is a lysosomal storage disease which is the most-common inherited cause of the Fanconi syndrome. Insights into the pathophysiology of the proximal tubular defect have come from in vitro studies of the cystine-loaded tubule. Proximal tubules loaded with cystine have a generalized proximal tubule transport defect characteristic of the Fanconi syndrome. The decrease in proximal tubular transport with cystine loading is not due to an increase in paracellular permeability with backflux of solute transport from the blood to the tubular lumen, but due to a decrease in active transport. The Na-K-ATPase activity is intact under Vmax conditions in cystine-loaded tubules; however, the production of ATP is severely compromised. The cystine-loaded tubule has a lower intracellular phosphate concentration than that of control tubules. This low intracellular phosphate concentration in cystine-loaded tubules likely plays a critical role in the decrease in intracellular ATP. Preservation of intracellular phosphate at control levels prevents the decrease in intracellular ATP and the proximal tubule respiratory dysfunction with cystine loading. The clinical significance and future directions for investigation are discussed.

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Cystine dimethyl ester reduces the forces driving sodium-dependent transport in LLC-PK1 cells

Bennun

Bashan

Potashnik

et al. 1992

American Journal of Physiology-Cell Physiology

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Cystinosis is an inherited metabolic disease characterized by accumulation of lysosomal cystine and renal impairment. In an attempt to better understand the link between cystine accumulation and renal functions, we studied the effects of cystine loading on the Na(+)-H+ antiporter and the sodium pump in renal epithelial cells (LLC-PK1) in culture. Incubation of LLC-PK1 with 1 mM cystine dimethyl ester (CDME) for 48 h caused lysosomal cystine loading and reduced by 22 +/- 2% the maximal velocity of sodium-hydrogen antiport with no significant change in the affinity of sodium for the transporter. Rubidium influx decreased to 46 +/- 5% of control. Ouabain binding experiments revealed a 10% reduction in the number of Na(+)-K(+)-ATPase units in the intact cells. Na(+)-K(+)-ATPase activity in the particulate fraction of the cells homogenate declined to 50 +/- 7.5% of controls. No significant change was observed in the activity of ouabain-insensitive phosphatases. The intracellular concentration of sodium increased from 20.6 +/- 3.7 to 64.8 +/- 10 mM, and potassium concentration decreased from 103 +/- 6 to 80 +/- 13 mM. In addition to the observed reduction in the sodium gradient and in agreement with the reduction in the intracellular potassium concentration, the membrane potential changed from -80.8 +/- 7.5 to -69.9 +/- 7.0 mV. The results suggest that intracellular accumulation of cystine is associated with reduction in the number and the activity of membrane transporters. The consequence of the changes in the activity of Na(+)-K(+)-ATPase is a reduction in the electrochemical forces that drive transport in the renal cells tested.(ABSTRACT TRUNCATED AT 250 WORDS)

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Cystine Accumulation Impairs Glucose Transport in LLC-PK<sub>1</sub> Cells

Cited by 4 publications

References 13 publications

Inhibition of Na+-Dependent Transporters in Cystine-Loaded Human Renal Cells

Inhibition of Na+-Dependent Transporters in Cystine-Loaded Human Renal Cells

The Fanconi syndrome of cystinosis: insights into the pathophysiology

Cystine dimethyl ester reduces the forces driving sodium-dependent transport in LLC-PK1 cells

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