Intracellular pH regulation and proton transport by rabbit renal medullary collecting duct cells

Abstract Proton secretion in the renal medullary collecting duct is thought to occur via a luminal proton-ATPase. In order to determine what mechanism(s) participate in proton transport across medullary collecting duct (MCD) cells membranes, intracellular pH (pHi) regulation and proton extrusion rates were measured in freshly prepared suspensions of rabbit outer MCD cells. Cells were separated by protease digestion and purified by Ficoll gradient centrifugation. pH, was estimated fluorometrically using the entrapped intracytoplasmic pH indicator, 6-carboxyfluorescein. Proton extrusion rates were measured using a pH stat. The resting pH, of MCD cells was 7.19±0.05 (SE) in a nonbicarbonate medium of pH 7.30. When cells were acidified by exposure to acetate salts or by abrupt withdrawal of ammonium chloride, they exhibited pH, recovery to the resting pH, over a 5-min timecourse. Depletion of >95% of cellular ATP content by poisoning with KCN in the absence of glucose inhibited pH, recovery. ATP depletion inhibited proton extrusion from MCD cells. Treatment with N-ethylmaleimide also inhibited pH, recovery. In addition, cellular ATP content was dependent on transmembrane pH gradients, suggesting that proton extrusion stimulated ATP hydrolysis. Neither removal of extracellular sodium nor addition of amiloride inhibited pH, recovery. These results provide direct evidence that a plasma membrane proton-ATPase, but not a Na+/ H' exchanger, plays a role in proton transport and pH, regulation in rabbit MCD

Hypokalemia, Its Contributing Factors and Renal Outcomes in Patients with Chronic Kidney Disease

Background: In the chronic kidney disease (CKD) population, the impact of serum potassium (sK) on renal outcomes has been controversial. Moreover, the reasons for the potential prognostic value of hypokalemia have not been elucidated. Design, Participants & Measurements 2500 participants with CKD stage 1–4 in the Integrated CKD care program Kaohsiung for delaying Dialysis (ICKD) prospective observational study were analyzed and followed up for 2.7 years. Generalized additive model was fitted to determine the cutpoints and the U-shape association between sK and end-stage renal disease (ESRD). sK was classified into five groups with the cutpoints of 3.5, 4, 4.5 and 5 mEq/L. Cox proportional hazard regression models predicting the outcomes were used. Results: The mean age was 62.4 years, mean sK level was 4.2±0.5 mEq/L and average eGFR was 40.6 ml/min per 1.73 m2. Female vs male, diuretic use vs. non-use, hypertension, higher eGFR, bicarbonate, CRP and hemoglobin levels significantly correlated with hypokalemia. In patients with lower sK, nephrotic range proteinuria, and hypoalbuminemia were more prevalent but the use of RAS (renin-angiotensin system) inhibitors was less frequent. Hypokalemia was significantly associated with ESRD with hazard ratios (HRs) of 1.82 (95% CI, 1.03–3.22) in sK 5 mEq/L conferred 1.6-fold (95% CI,1.09–2.34) increased risk of ESRD compared with sK = 4.5–5 mEq/L. Hypokalemia was also associated with rapid decline of renal function defined as eGFR slope below 20% of the distribution range. Conclusion: In conclusion, both hypokalemia and hyperkalemia are associated with increased risk of ESRD in CKD population. Hypokalemia is related to increased use of diuretics, decreased use of RAS blockade and malnutrition, all of which may impose additive deleterious effects on renal outcomes

Nutritional Management of Kidney Stones (Nephrolithiasis)

The incidence of kidney stones is common in the United States and treatments for them are very costly. This review article provides information about epidemiology, mechanism, diagnosis, and pathophysiology of kidney stone formation, and methods for the evaluation of stone risks for new and follow-up patients. Adequate evaluation and management can prevent recurrence of stones. Kidney stone prevention should be individualized in both its medical and dietary management, keeping in mind the specific risks involved for each type of stones. Recognition of these risk factors and development of long-term management strategies for dealing with them are the most effective ways to prevent recurrence of kidney stones

Relation between the anion exchange protein in kidney medullary collecting duct cells and red cell band 3

A membrane protein that is immunochemically similar to the red cell anion exchange protein, band 3, has been identified on the basolateral face of the outer medullary collecting duct (MCD) cells in rabbit kidney. In freshly prepared separated rabbit MCD cells, M.L. Zeidel, P. Silva and J.L. Seifter (J. Clin. Invest.77:1682–1688, 1986) found that C−/HCO3- exchange was inhibited by the stilbene anion exchange inhibitor, DIDS (4,4′-diisothiocyano-2,2′-disulfonic stilbene), with a K1 similar to that for the red cell. We have measured the binding affinities of a fluorescent stilbene inhibitor, DBDS (4,4′-dibenzamido-2,2′-disulfonic stilbene), to MCD cells in 28.5 mM citrate and have characterized both a high-affinity site (K1s=93±24 mM) and a lower affinity site (K2s=430±260 nM), which are closely similar to values for the red cell of 110±51 nM for the high-affinity site and 980±200 nM for the lower affinity site (A.S. Verkman, J.A. Dix & A.K. Solomon,J. Gen. Physiol.81:421–449, 1983). Whn Cl− replaces citrate in the buffer, the two sites collapse into a single one withK1s=1500±400 nM, similar to the singleK1s=1200±200 nM in the red cell (J.A. Dix, A.S. Verkman & A.K. Solomon,J. Membrane Biol.89:211–223, 1986). The kinetics of DBDS binding to MCD cells at 0.25 μM are characterized by a fast process, τ=0.14±0.03 sec, similar to τ=0.12±0.03 sec in the red cell. These similarities show that the physical chemical characteristics of stilbene inhibitor binding to MCD cell ‘band 3’ closely resemble those for red cell band 3, which suggests that the molecular structure is highly conserved

Restricted Cubic Spline Regression Plot of the U-shape Association between sK and the Risk for End Stage Renal Disease.

<p>Covariates included in the model were the same as the Cox regression in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067140#pone-0067140-t004" target="_blank">Table 4</a>. Serum potassium (sK), body mass index, mean blood pressure and C-reactive protein were treated as restricted cubic spline functions. The solid line represents the log transformed multivariable-adjusted hazard ratio of ESRD. The dashed lines indicate the 95% confidence intervals. sK below 4.03 and above 5.11 mEq/L were associated with higher hazard (log hazard ratio >0). Tick marks on the x-axis indicate individual observations at corresponding levels of sK.</p

Baseline Demographic and Clinical Characteristics.

<p>Data expressed as mean ± standard deviation, median (interquartile range) or percentage.</p><p>BMI, body mass index; MBP, mean blood pressure; CKD, chronic kidney disease; eGFR, estimated glomerular filtration rate; CRP, C-reactive protein; HbA1c, glycated hemoglobin; UPCR, Urine protein-to-creatinine ratio; ESRD, End Stage Renal Disease.</p><p>Comparisons are made by ANOVA or the chi-square test.</p>*<p>eGFR slope less than −6.88 mL/min/1.73 m²/yr (%).</p>#<p>Urine protein-to-creatinine (mg/g).</p