Mechanisms Underlying Calcium Nephrolithiasis.

Nephrolithiasis is a worldwide problem with increasing prevalence, enormous costs, and significant morbidity. Calcium-containing kidney stones are by far the most common kidney stones encountered in clinical practice, and thus, hypercalciuria is the greatest risk factor for kidney stone formation. Hypercalciuria can result from enhanced intestinal absorption, increased bone resorption, or altered renal tubular transport. Kidney stone formation is complex and driven by high concentrations of calcium-oxalate or calcium-phosphate in the urine. After discussing the mechanism mediating renal calcium salt precipitation, we review recent discoveries in renal tubular calcium transport from the proximal tubule, thick ascending limb, and distal convolution. Furthermore, we address how calcium is absorbed from the intestine and mobilized from bone. The effect of acidosis on bone calcium resorption and urinary calcium excretion is also considered. Although recent discoveries provide insight into these processes, much remains to be understood in order to provide improved therapies for hypercalciuria and prevent kidney stone formation.

Axial and cellular heterogeneity in electrolyte transport pathways along the thick ascending limb.

The thick ascending limb (TAL) extends from the border of the inner medulla to the renal cortex, thus ascending through regions with wide differences in tissue solute and electrolyte concentrations. Structural and functional differences between TAL cells in the medulla (mTAL) and the cortex (cTAL) would therefore be useful to adapt TAL transport function to a changing external fluid composition. While mechanisms common to all TAL cells play a central role in the reclamation of about 25% of the NaCl filtered by the kidney, morphological features, Na+ / K+ -ATPase activity, NKCC2 splicing and phosphorylation do vary between segments and cells. The TAL contributes to K+ homeostasis and TAL cells with high or low basolateral K+ conductances have been identified which may be involved in K+ reabsorption and secretion respectively. Although transport rates for HCO3- do not differ between mTAL and cTAL, divergent axial and cellular expression of H+ transport proteins in TAL have been documented. The reabsorption of the divalent cations Ca2+ and Mg2+ is highest in cTAL and paralleled by differences in divalent cation permeability and the expression of select claudins. Morphologically, two cell types with different cell surface phenotypes have been described that still need to be linked to specific functional characteristics. The unique external environment and its change along the longitudinal axis require an axial functional heterogeneity for the TAL to optimally participate in conserving electrolyte homeostasis. Despite substantial progress in understanding TAL function, there are still considerable knowledge gaps that are just beginning to become bridged.

A single simple procedure for dewaxing, hydration and heat-induced epitope retrieval (HIER) for immunohistochemistry in formalin fixed paraffin-embedded tissue.

Heat-induced epitope retrieval (HIER) is widely used for immunohistochemistry on formalin fixed paraffin-embedded tissue and includes temperatures well above the melting point of paraffin. We therefore tested whether traditional xylene-based removal of paraffin is required on sections from paraffin-embedded tissue, when HIER is performed by vigorous boiling in 10 mM Tris/0.5 mM EGTA-buffer (pH=9). Immunohistochemical results using HIER with or without prior dewaxing in xylene were evaluated using 7 primary antibodies targeting proteins located in the cytosol, intracellular vesicles and plasma membrane. No effect of omitting prior dewaxing was observed on staining pattern. Semiquantitative analysis did not show HIER to influence the intensity of labelling consistently. Consequently, quantification of immune labelling intensity using fluorescent secondary antibodies was performed at 5 dilutions of primary antibody with and without prior dewaxing in xylene. No effect of omitting prior dewaxing on signal intensity was detectable indicating similar immunoreactivity in dewaxed and non-dewaxed sections. The intensity of staining the nucleus with the DNA-stain ToPro3 was similarly unaffected by omission of dewaxing in xylene. In conclusion, the HIER procedure described and tested can be used as a single procedure enabling dewaxing, hydration and epitope retrieval for immunohistochemistry in formalin fixed paraffin-embedded tissue.

Opposing effects of NaCl restriction and carbohydrate loading on urine volume in diabetic rats.

AIM: To test the effects of dietary NaCl and carbohydrate content on urine volume in diabetic rats. METHODS: Streptozotocin-induced diabetic rats were subjected to NaCl restriction using either a NaCl-deficient carbohydrate-rich synthetic diet (Altromin C1036) supplemented to contain 0.16% NaCl (C1036 + lowNaCl) or a modified normal cereal-based diet (Altromin 1320) containing 0.086% NaCl (lowNaCl-1320). Normal diet contained 0.2683% NaCl. RESULTS: Using the C1036 + lowNaCl diet, earlier reported paradoxical increases in water intake and urine volume of diabetic rats were reproduced. However, water intake and urine volume also increased in diabetic rats offered the synthetic C1036 diet supplemented with NaCl to normal levels. Using the lowNaCl-1320 diet, water intake and urine volume were markedly reduced. Highly significant correlations between urine volume and both osmotic output and urinary glucose excretion were found in diabetic rats on normal diet, but these correlations were absent in diabetic rats on synthetic diet, which showed higher urine volumes than expected from the correlations. In contrast, urine volume was significantly correlated with carbohydrate intake in diabetic rats, irrespective of the diet. CONCLUSIONS: (i) The synthetic diet dramatically increases the urine volume in STZ-DM rats irrespectively of NaCl content. (ii) Rats with STZ-DM on a normal diet show reduced water intake and urine volume in response to dietary NaCl restriction. (iii) A shift to high carbohydrate diet induces polyuria in STZ-DM rats. (iv) Urine volume in all STZ-DM rats only shows correlation with dietary carbohydrate intake. (v) Glucose-driven osmotic diuresis is unlikely to explain the carbohydrate-induced polyuria.