Objective: Uromodulin (UMOD) is the most abundantly secreted protein found within the urine, primarily produced by medullary thick ascending limb (mTAL) epithelial cells of the kidneys. There is accruing genetic evidence implicating UMOD in blood pressure regulation and consequently hypertension. The aim of this study was to investigate the potential role of extracellular calcium and the calcium-sensing receptor (CaSR) on UMOD production and secretion in TAL cells with the hope of defining novel clinical targets for the treatment of hypertension. Design and method: mTAL tubules from Wistar-Kyoto (WKY) and stroke-prone spontaneously hypertensive (SHRSP) female rats were incubated with calcium, nifedipine (calcium channel blocker), CaSR agonist (spermine) and antagonist (NPS2143). UMOD protein levels were detected by Western blot. Gene expression of Umod was determined by qRT-PCR. Results: Calcium increased mTAL UMOD secretion in WKY and SHRSP. Nifedipine slightly decreased UMOD secretion in WKY without calcium. In both strains, NPS2143 increased calcium-induced UMOD secretion, with an enhanced effect in SHRSP. Stimulation of CaSR with spermine decreased UMOD secretion in WKY. Analysis of intracellular UMOD levels in these conditions demonstrated increased accumulation when extracellular secretion was low, and vice versa. Incubation of primary mTAL cells with calcium confirmed increased localisation of UMOD at the membrane compared to cytosol. The Umod mRNA level changes were not statistically significant among conditions. Conclusions: Trafficking of UMOD is influenced by the CaSR ligand and the biased nature of G-protein coupled CaSR signalling. Unravelling the signalling events post-calcium will be necessary for identification of key regulators of UMOD secretion.
Objective: Our traditionally nephrocentric view of Na+ homeostasis has recently been challenged by the concept of Na+ accumulation in tissues, predominantly skin, observed in hypertension. Its proposed water-independent (hypertonic) nature, carrying pathogenic potential, lacks firm demonstration. This study investigates the nature and clinical correlates of skin Na+ accumulation in hypertensive patients. Design and method: Responses to a validated salt intake questionnaire, measures of transepidermal water loss (TEWL), blood samples and a skin punch biopsy were collected from adult, non-pregnant and consenting hypertensive patients at the Glasgow Blood Pressure Clinic. Histochemical analysis was conducted by splitting the skin into epidermis/superficial dermis (ESD) and deep dermis (DD) layers. Water was measured by gravimetric approach (wet weight – dry weight), and Na+ and K+ by flame photometry. Results: Seventy-six patients consented to skin biopsy (age: 58 ± 15, range 22–86 years; females = 47.4%; BMI: 30.3, IQR 27.5–36.3). Na+ concentration ([Na+]) and K+ concentration ([K+]) was lower and higher in ESD (richer in cellularity) compared to DD, respectively (112.6 ± 8.57 and 30.17 ± 4.88 vs 119.0 ± 11.81 and 17.28 ± 3.62 mmol/l), respectively (p < 0.001 for both). Virtually none of the patients had [Na++K+] exceeding normal physiological levels in either layer, thus excluding hypertonic extracellular Na+ accumulation. Water content was positively and negatively correlated with Na+ and K+, respectively. Female DD, which is richer in fat content, contained less water, Na+ and K+ than male dermis (p < 0.01 for all), with no difference in concentrations or in ESD. Age was associated with an increase in ESD water and [Na+], and a decrease in [K+] in both layers, suggestive of oedema accumulation and independent of sex, BMI and estimated Na intake (padj < 0.005). The latter independently predicted epidermal water content (padj = 0.017). Patients with uncontrolled BP had higher ESD [Na+] (padj = 0.02). NT-proBNP levels mirrored all measures of skin oedema accumulation, while no significant interaction with medications or TEWL was observed. Conclusions: Skin Na+ accumulation in hypertensive patients is isotonic, prevalent and reflects subclinical oedema. Tissue architecture considerably impacts on Na+ content and concentration, whereas TEWL has minimal effect. These results have mechanistic, diagnostic and therapeutic implications.
Background and Aims Uromodulin (UMOD) is the most abundant renal protein secreted into urine by the thick ascending epithelial (TAL) cells of the loop of Henle. Genetic studies have demonstrated an association between UMOD risk variants and hypertension. Studies on UMOD overexpressing transgenic mice have shown that UMOD increases the tubular salt reabsorption via enhanced NKCC2 activity. We aimed to dissect the effect of salt-loading and blood pressure on the excretion of UMOD. Method Wistar-Kyoto (WKY) and stroke-prone spontaneously hypertensive (SHRSP) rats (n=8/sex/strain) were maintained on 1% NaCl for three weeks. Salt-loaded SHRSP were treated with nifedipine. Tubule isolation and ex vivo incubation with nifedipine were used to assess its direct effect on TAL. Results Urinary UMOD excretion was significantly reduced after salt loading in both strains (figure). In salt-loaded SHRSP, nifedipine treatment reduced blood pressure and urinary UMOD excretion. The reductions in urinary UMOD excretion were dissociated from unchanged kidney UMOD protein and mRNA levels, however, were associated with UMOD endoplasmic reticulum accumulation, thus suggesting secretion as a key regulatory step. Ex vivo experiments with TAL tubules showed that nifedipine did not have a direct effect on UMOD secretion. Conclusion Our data suggest a direct effect of salt on UMOD secretion independent of blood pressure and a potential role of endoplasmic reticulum stress on the control of UMOD secretion. The role of UMOD as a cardiovascular risk marker deserves mechanistic reappraisal and further investigations based on our findings.
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