Urea Flux Research Articles

Influence of submarine groundwater discharge on the nutrient dynamics of a fringing-reef lagoon

Study regionThis study investigates nutrient distribution and flux dynamics in a coral reef lagoon in Quintana Roo, Mexico, located on a permeable limestone coast of the Mesoamerican Barrier Reef System. Study FocusEmphasis is placed on submarine groundwater discharge (SGD) as a crucial contributor to nutrient pathways, including ammonium (NH4+), nitrate and nitrite (NOx-), hydrogen silicate (HSiO3-), hydrogen phosphate (HPO42-), and urea. Inputs vary with SGD magnitudes and sources and by proximity to active spring discharges. Groundwater multi-tracer analysis and multiple linear regression identify 226Ra as explaining NH4+ variability due to long-term groundwater processes, while 223Ra predicts NOx-, HSiO3-, and urea due to short-term inputs. No significant relationship was found between HPO42- and any radium isotope, indicating complex behavior in coastal karst aquifers. New hydrological insights for the regionThe findings highlight complex nutrient dynamics in coastal karst settings, with SGD-derived fluxes primarily consisting of dissolved inorganic nitrogen (DIN) and HSiO3-. Although lower in concentration, HPO42- and urea fluxes are significant compared to other karst environments. Radium isotopes distinguish between short-term and long-term, as well as new and recycled nutrient inputs. Groundwater inputs transport fresh nutrients to healthier reefs, whereas processed, recycled inputs were detected near degraded reefs. These insights are essential for understanding global nutrient cycles and coral health, particularly in the context of global change and anthropogenic disturbances affecting coral reef ecosystems.

Acute effects of testosterone on whole body protein metabolism in hypogonadal and eugonadal conditions: a randomized, placebo-controlled, crossover study.

Chronic testosterone (T) substitution and short-term T administration positively affect protein metabolism, however, data on acute effects in humans are sparse. This study aimed to investigate T's acute effects on whole body protein metabolism in hypogonadal and eugonadal conditions. We designed a randomized, double-blind, placebo-controlled, crossover study, including 12 healthy young males. Whole body protein metabolism was evaluated during 1) eugonadism, and after medically induced hypogonadism, with application of a gel on each trial day containing either 2) placebo, 3) T 50 mg, or 4) T 150 mg; under basal (5-h basal period) and insulin-stimulated conditions (3-h clamp). The main outcome measure was a change in net protein balance. The net protein loss was 62% larger in the placebo-treated hypogonadal state compared with the eugonadal state during the basal period (-5.5 ± 3.5 µmol/kg/h vs. -3.4 ± 1.2 µmol/kg/h, P = 0.038), but not during the clamp (P = 0.06). Also, hypogonadism resulted in a 25% increase in whole body urea flux (P = 0.006). However, T did not result in any significant changes in protein breakdown, synthesis, or net balance during either the basal period or clamp (all P > 0.05). Protein breakdown was reduced during clamp compared with the basal period regardless of gonadal status or T exposure (all P ≤ 0.001). In conclusion, the application of transdermal T did not counteract the negative effects of hypogonadism with no effects on protein metabolism within 5 h of administration. Insulin (during clamp) mitigated the effects of hypogonadism. This study is the first to investigate acute protein metabolic effects of T in hypogonadal men.NEW & NOTEWORTHY In a model of medically induced hypogonadism in male volunteers, we found increased whole body urea flux and net protein loss as an expected consequence of hypogonadism. Our study demonstrates the novel finding that the application of transdermal testosterone had no acute effects on whole body protein metabolism under eugonadal conditions, nor could it mitigate the hypogonadism-induced changes in protein metabolism. In contrast, insulin (during clamp) mitigated the effects of hypogonadism on protein metabolism.

Whole-body urea kinetics and functional roles of urea transporters and aquaporins in urea secretion into the rumen in sheep fed diets varying in crude protein content and corn grain processing method.

The objectives were to determine the effects of dietary crude protein (CP) content and corn grain processing on whole-body urea kinetics and the functional roles of urea transporter-B (UT-B) and aquaporins (AQP) in serosal-to-mucosal urea flux (Jsm-urea) in ovine ruminal epithelia. Thirty-two Rideau-Arcott ram lambs were blocked by bodyweight into groups of 4 and then randomly allocated within blocks to 1 of 4 diets (n = 8) in a 2 × 2 factorial design. Dietary factors were CP content (11% [LP] vs. 16% [HP]) and corn grain processing (whole-shelled [WSC] vs. steam-flaked [SFC] corn). Whole-body urea kinetics and N balance were determined using 4-d continuous intrajugular infusions of [15N15N]-urea with concurrent collections of urine and feces with four blocks of lambs (n = 4). After 23 d on diets, lambs were killed to collect ruminal epithelia for mounting in Ussing chambers to determine Jsm-urea and the measurement of mRNA abundance of UT-B and AQP. Serosal and mucosal additions of phloretin and NiCl2 were used to inhibit UT-B- and AQP-mediated urea transport, respectively. Lambs fed HP had a greater (P < 0.01) N intake (29.4 vs. 19.1g/d) than those fed LP; however, retained N (g/d or % of N intake) was not different. As a % of N intake, lambs fed SFC tended (P = 0.09) to have a lower N excretion (72.2 vs. 83.5%) and a greater N retention (27.8 vs. 16.6%) compared to those fed WSC. Endogenous urea-N production (UER) was greater in lambs fed HP compared to those fed LP (29.9 vs. 20.6g/d; P = 0.02), whereas urea-N secreted into the gut (GER; g/d) and urea-N used for anabolic purposes (UUA; g/d) were similar. Lambs fed LP tended (P = 0.05) to have greater GER:UER (0.78 vs. 0.66) and UUA:GER (0.23 vs. 0.13) ratios, and a greater Jsm-urea (144.7 vs. 116.1 nmol/[cm2 × h]; P = 0.07) compared to those fed HP. Lambs fed SFC tended to have a lower NiCl2-insensitive Jsm-urea (117.4 vs. 178.4 nmol/[cm2 × h]; P = 0.09) and had a lower phloretin-insensitive Jsm-urea (87.1 vs. 143.1 nmol/[cm2 × h]; P = 0.02) compared to those fed WSC. The mRNA abundance of UT-B (0.89 vs. 1.07; P = 0.08) and AQP-3 (0.90 vs. 1.05; P = 0.07) tended to be lower in lambs fed SFC compared to those fed WSC. Overall, reducing CP content tended to increase the GER:UER ratio with no changes in the expression or function of UT-B and AQP. Although corn grain processing had no effects on GER, feeding SFC increased the portion of urea secretion into the rumen that was mediated via UT-B and AQP.

High flux novel polymeric membrane for renal applications

Biocompatibility and the ability to mediate the appropriate flux of ions, urea, and uremic toxins between blood and dialysate components are key parameters for membranes used in dialysis. Oxone-mediated TEMPO-oxidized cellulose nanomaterials have been demonstrated to be excellent additives in the production and tunability of ultrafiltration and dialysis membranes. In the present study, nanocellulose ionic liquid membranes (NC-ILMs) were tested in vitro and ex vivo. An increase in flux of up to two orders of magnitude was observed with increased rejection (about 99.6%) of key proteins compared to that of polysulfone (PSf) and other commercial membranes. NC-ILMs have a sharper molecular weight cut-off than other phase inversion polymeric membranes, allowing for high throughput of urea and a uremic toxin surrogate and limited passage of proteins in dialysis applications. Superior anti-fouling properties were also observed for the NC-ILMs, including a > 5-h operation time with no systemic anticoagulation in blood samples. Finally, NC-ILMs were found to be biocompatible in rat ultrafiltration and dialysis experiments, indicating their potential clinical utility in dialysis and other blood filtration applications. These superior properties may allow for a new class of membranes for use in a wide variety of industrial applications, including the treatment of patients suffering from renal disease.

Dialysate regeneration via urea photodecomposition with TiO2 nanowires at therapeutic rates.

The standard weekly treatment for end-stage renal disease patients is three 4-h-long hemodialysis sessions with each session c'onsuming over 120 L of clean dialysate, which prevents the development of portable or continuous ambulatory dialysis treatments. The regeneration of a small (~1L) amount of dialysate would enable treatments that give conditions close to continuous hemostasis and improve patient quality of life through mobility. Small-scale studies have shown that nanowires of TiO2 are highly efficient at photodecomposing urea into CO2 and N2 when using an applied bias and an air permeable cathode. To enable the demonstration of a dialysate regeneration system at therapeutically useful rates, a scalable microwave hydrothermal synthesis of single crystal TiO2 nanowires grown directly from conductive substrates was developed. These were incorporated into 1810 cm2 flow channel arrays. The regenerated dialysate samples were treated with activated carbon (2min at 0.2g/mL). The photodecomposition system achieved the therapeutic target of 14.2g urea removal in 24 h. TiO2 electrode had a high urea removal photocurrent efficiency of 91%, with less than 1% of the decomposed urea generating NH4 + (1.04 μg/h/cm2 ), 3% generating NO3 - and 0.5% generating chlorine species. Activated carbon treatment could reduce total chlorine concentration from 0.15 to <0.02 mg/L. The regenerated dialysate showed significant cytotoxicity which could be removed by treatment with activated carbon. Additionally, a forward osmosis membrane with sufficient urea flux can cut off the mass transfer of the by-products back into the dialysate. Urea could be removed from spent dialysate at a therapeutic rate using a TiO2 based photooxidation unit, which can enable portable dialysis systems.

Fermentable Fibers or Oligosaccharides Reduce Urinary Nitrogen Excretion by Increasing Urea Disposal in the Rat Cecum

The availability of fermentable carbohydrates could influence the digestive degradation and disposal of blood urea. The effects of a poorly fermented cellulosic oat fiber, a soluble fermentable fiber (gum arabic) or one of two oligosaccharides (fructooligosaccharide or xylooligosaccharide) on nitrogen excretion were compared with a wheat starch-based control diet in male Wistar rats. The fibers and oligosaccharides were added to the semipurified diets at 7.5 g/100 g in place of wheat starch. The diets contained 13 g casein/100 g. Oat fiber did not cause an enlargement of the cecum. In contrast, gum arabic and the oligosaccharides elicited a 35–60% enlargement of the cecal wall and a 2 to 2.6-fold mean increase in the cecal pool of short chain fatty acids. Compared with rats fed the oat fiber-based diet, urea flux from blood to cecum was nearly 50% greater and more than 120% greater in those fed the gum arabic and oligosaccharide diets, respectively. In those groups, net nitrogen retention in the cecum more than doubled (nitrogen retention was calculated as the difference between net urea nitrogen flux into the cecum and ammonia nitrogen reabsorption). As a percentage of total excreted nitrogen, fecal nitrogen was 20% in the oat fiber group and 27–29% in the gum arabic and oligosaccharide groups, compared with only 10% in fiber-free controls. Results indicate that under these dietary conditions, the addition of oligosaccharides to the diet induced a 20 to 30% decrease in blood urea and renal nitrogen excretion relative to the control, indicating a potential for oligosaccharide diet therapy in chronic renal disease.

PSX-A-2 Effects of Dietary Crude Protein Level and Corn Grain Processing on Whole-Body Urea Kinetics and in-Vitro Urea Secretion Into the Rumen of Sheep

Abstract In ruminants, urea-N salvaging allows secretion of endogenously-produced urea (UER) into the rumen, where it can provide N for microbial growth. Our objective was to determine the effects of dietary crude protein content and corn grain processing on whole-body urea kinetics and in vitro serosal-to-mucosal urea flux (Jsm-urea) across ruminal epithelia. Thirty-two Rideau-Arcott ram lambs were blocked by BW into groups of 4 and then randomly allocated within block to one of 4 dietary treatments (n = 8) in a 2 × 2 factorial design. Dietary factors were crude protein content (10.9% [LP] vs. 15.8% [HP]) and corn grain processing (whole-shelled [WSC] vs. steam-flaked [SFC]). Feeding SFC was expected to provide more ruminally-fermentable energy than WSC. Four blocks of animals (n = 4) were used in 4-d continuous infusions of 15N-15N with concurrent collections of urine and feces to determine whole-body urea kinetics. After 23 d of dietary exposure, lambs were killed to collect ruminal epithelia which were mounted in Ussing chambers to determine Jsm-urea. Lambs fed HP had greater N intake (P = 0.01) and total N excretion (P = 0.002) than those fed LP; however, retained N (g/d or % of N intake) was similar between the 2 groups. When expressed as absolute amounts, urea-N secreted into the gut (GER) and urea-N used for anabolic purposes (UUA) were similar across diets; however, lambs fed the LP diet had a greater GER/UER ratio (P = 0.02) and tended to have a greater UUA/GER ratio (P = 0.05) than lambs fed the HP diet. Diet had no effect on ruminal Jsm-urea. Grain processing had no effects on GER, UUA and Jsm-urea. These results suggest ruminants fed protein-restricted diets have a greater dependence on urea secretion into the gut to provide N for anabolic use.

Modeling ammonia volatilization following urea and controlled-release urea application to paddy fields

• The WHCNS model performed well in simulating NH 3 flux measured by a continuous ventilation method. • Urea split application reduced the peak of NH 3 flux but increased total NH 3 loss. • Controlled-release urea application significantly reduced NH 3 loss compared to urea. • The controlled-release urea rate was recommended to reduce N loss without reducing yield. Ammonia volatilization, a main pathway of nitrogen (N) loss in paddy fields, can easily lead to environmental problems, such as haze and water eutrophication. Field measurement of ammonia volatilization is time-consuming and expensive. Soil–crop system models can simulate the effect of different fertilizer management practices on NH 3 volatilization and compensate for the limitations of field measurement. Therefore, the WHCNS (soil Water Heat Carbon Nitrogen Simulator) model was used to evaluate the feasibility of simulating NH 3 volatilization in paddy fields under different N fertilizer types and application methods. The field experiment, including four treatments of farmer’s practice (FP), basal application of urea in one dose (SBN), split application of urea in three doses (SPN), and basal application of controlled-release urea in one dose (CRU), was conducted from 2013 to 2014. All treatments were conducted with a fertilizer rate of 165 kg N ha −1 , except for FP with a rate of 210 kg N ha −1 . The measured dry matter weight, grain yield, N uptake, and NH 3 flux were used to test the WHCNS model. The model evaluation indices of Nash–Sutcliffe efficiency ( NSE ) and index of agreement ( d ) were close to 1 for the measured variables, except for NSE of yield (0.16). The normalized root mean square error ( NRMSE ) of NH 3 flux was slightly over 30%, but lower than 30% for the other items. Results showed that the model can successfully simulate rice yield and NH 3 flux of urea and controlled-release urea under different application methods. The simulated annual average cumulative NH 3 losses of FP, SBN, SPN, and CRU accounted for 18.9%, 17.8%, 20.6%, and 10.1% of their N inputs, respectively. Compared with the FP treatment, the ratios of NH 3 losses of the SBN and CRU treatments were decreased by 1.2%-8.8%. Compared with the SBN treatment (the same rate of 165 kg N ha −1 ), SPN reduced the peak of NH 3 flux but increased the cumulated NH 3 losses and CRU significantly reduced the NH 3 flux and cumulative NH 3 losses. The application rate of controlled-release urea could be further optimized by refined model calculations to reduce NH 3 volatilization and total N loss while maintaining yield.

Simulation of nitrogen fertigation schedule for drip irrigated paddy

A robust soil-water system is essential for achieving most of the Sustainable Development Goals of United Nations. Systems approach is essential to keep the soil-water system to derive sustainable benefits. Prevention is better than cure; hence accurate prediction of the likely results of our action on soil-water system is needed. Hydrus is one of the handy tools for simulating root zone under drip irrigation to derive best management practices for water and nutrient application. Hydrus is parameter intensive software. Hence, mostly, there is a tendency to use typical values of parameters available in the literature for modelling. In this research work, we generated data for Hydrus by intensive field experimentation to improve the reliability of modelling. The first order rate constants of urea-ammonium-nitrate reactions were obtained by field experimentation. HYDRUS was used numerically to simulate nitrogen transport for paddy crop under drip irrigation. From the simulation, various fluxes of urea, ammonium and nitrate were quantified, to develop a N fertigation schedule for entire crop duration. The fertigation schedule was implemented by raising a paddy crop and evaluation was done by estimating N assimilation of crop by plant sample analysis. The simulated values of nitrogen uptake were in good agreement with the observed values of nitrogen uptake.

PROTON PUMP INHIBITORS (PPI) INDUCES COLONIC TIGHT JUNCTION BARRIER (TJ) DYSFUNCTION VIA AN UPREGULATION OF TJ PORE FORMING CALUDIN-2 PROTEIN

Abstract Background Proton pump inhibitors (PPIs) are highly effective antagonists of gastric acid secretion and are widely used to treat a number of gastroesophageal disorders, including peptic ulcer disease, GERD, and Barrett’s esophagus. PPI-induced elevation in intra-gastric pH and subsequent alterations of gastrointestinal physiology are thought to cause undesired effects on the entire GI tract. Defective intestinal Tight Junction (TJ) barrier is an important pathogenic factor for intestinal inflammation. Claudin-2 is a pore forming TJ protein whose overexpression causes selective increase in TJ permeability to small molecules. Claudin-2 expression is known to be up-regulated in intestinal inflammation and inflammation-associated colon carcinogenesis. The effect of PPI on Intestinal TJ barrier is not known. Aim: The aim of the present study was to study the effect of PPI on Intestinal TJ permeability. Methods A cell culture model of filter grown human intestinal epithelial Caco-2 monolayers, was used to study intestinal epithelial TJ barrier function. The mouse colonic permeability was measured by Ussing chambers studies. Western blot (WB) and Immunofluorescence (IF) was used to study the protein expression of claudin-2 in Caco-2 cells and in mouse colon. Cell surface biotinylation was used to study intracellular trafficking of TJ proteins. Results PPIs, caused a concentration- and time-dependent decrease in transepithelial resistance (TER), and increase in urea flux in filter-grown Caco-2 cells. Further studies on intestinal TJ revealed that PPI caused a significant increase in pore forming TJ protein claudin-2 protein level (western blot), but not mRNA (RT-PCR) in Caco-2 monolayers. Claudin-2 was co-localized to its known intercellular trafficking vesicles, clathrin pits. PPI treatment caused an increase in expression and junctional localization of claudin-2 and a decrease in its cytoplasmic co-localization with clathrin. Similarly, pulse-chase assay showed that the claudin-2 half-life was increased by PPI. In cell surface biotinylation assay, the rate of claudin-2 endocytosis was significantly reduced by PPI. In ex-vivo, Ussing chamber studies, PPI administration for 45 days resulted in a significant decrease in murine colonic TER and increase in urea flux. Immunofluorescence and Western blot analysis showed that PPI also caused an increase in TJ protein claudin-2 expression in mice colonic mucosa. Conclusion Our results suggest that the PPIs induces colonic TJ permeability via increasing pore forming claudin-2 levels. PPIs affect intracellular trafficking to reduce claudin-2 degradation and increased its junctional localization.

Influence of forage level and corn grain processing on whole-body urea kinetics, and serosal-to-mucosal urea flux and expression of urea transporters and aquaporins in the ovine ruminal, duodenal, and cecal epithelia.

The objectives were to determine the effects of forage level and grain processing on whole-body urea kinetics, N balance, serosal-to-mucosal urea flux (Jsm-urea), and messenger ribonucleic acid (mRNA) abundance of urea transporter-B (UT-B; SLC14a1) and aquaporins (AQP) in ovine ruminal, duodenal, and cecal epithelia. Thirty-two wether lambs were blocked by body weight into groups of four and assigned to one of four diets (n = 8) in a 2 × 2 factorial design. Dietary factors were forage level (30% [LF] vs. 70% [HF]) and corn grain processing (whole-shelled [WS] vs. steam-flaked [SF]). Four blocks of lambs (n = 4) were used to determine urea kinetics and N balance using 4-d [15N15N]-urea infusions with concurrent fecal and urine collections. Lambs were killed after 23 d of dietary adaptation. Ruminal, duodenal, and cecal epithelia were collected to determine Jsm-urea and mRNA abundance of UT-B and AQP. Lambs fed LF had greater intakes of dry matter (DMI; 1.20 vs. 0.86 kg/d) and N (NI; 20.1 vs. 15.0 g/d) than those fed HF (P < 0.01). Lambs fed SF had greater DMI (1.20 vs. 0.86 kg/d) and NI (20.6 vs. 14.5 g/d) than those fed WS (P < 0.01). As a percentage of NI, total N excretion was greater in lambs fed HF compared with those fed LF (103% vs. 63.0%; P < 0.01) and was also greater in lambs fed WS compared with those fed SF (93.6% vs. 72.1%; P = 0.02). Retained N (% of NI) was greater in lambs fed LF compared with those fed HF (37.0% vs. -2.55%; P < 0.01). Lambs fed SF had a greater (P = 0.02) retained N (% of NI; 28.0% vs. 6.50%) compared with those fed WS. Endogenous urea production (UER) tended (P = 0.09) to be greater in lambs fed HF compared with those fed LF. As a proportion of UER, lambs fed HF had a greater urinary urea-N loss (0.38 vs. 0.22) and lower urea-N transferred to the gastrointestinal tract (GIT; 0.62 vs. 0.78) or urea-N used for anabolism (as a proportion of urea-N transferred to the GIT; 0.12 vs. 0.26) compared with lambs fed LF (P < 0.01). Ruminal Jsm-urea was unaffected by diet. Duodenal Jsm-urea was greater (P < 0.01) in lambs fed HF compared with LF (77.5 vs. 57.2 nmol/[cm2 × h]). Lambs fed LF had greater (P = 0.03) mRNA expression of AQP3 in ruminal epithelia and tended (P = 0.06) to have greater mRNA expression of AQP3 in duodenal epithelia compared with lambs fed HF. Expression of UT-B mRNA was unaffected by diet. Our results showed that feeding more ruminally available energy improved N utilization, partly through a greater proportion of UER being transferred to the GIT and being used for anabolic purposes.

Spatiotemporal Urea Distribution, Sources, and Indication of DON Bioavailability in Zhanjiang Bay, China

In marine environments, urea is an important component of the biogeochemical cycle of nitrogen. The autochthonous and allochthonous sources (rivers, aquaculture, waste water input, etc.) of urea play a key role in urea cycles in adjacent coastal waters. Because urea is a specific marker to trace the sewage fluxes in coastal waters, we investigated urea associated with terrestrial source input and coastal water in Zhanjiang Bay (ZJB) during the time from November 2018 to July 2019, and the spatiotemporal urea distribution and the bioavailability of dissolved organic nitrogen (DON) based on urea concentration in the ZJB were explored. The results showed that the urea enrichment in coastal water was mainly due to discharge from urban sewage systems, rivers, and coastal aquaculture. The concentration of urea ranged from 1.14 to 5.53 μmol·L−1, and its mean value was 3.13 ± 1.02 μmol·L−1 in the ZJB. The urea concentration showed a significantly different seasonal variation in the ZJB (p &lt; 0.05), and the highest and lowest concentrations were found in November 2018 and April 2019, respectively. Its high value appeared in the north and northeast of the ZJB, which were polluted by coastal aquaculture and agriculture fertilizer utilization. The range of urea concentration of terrestrial source inputs in the ZJB was 1.31–10.29 μmol·L−1, and the average urea concentration reached 3.22 ± 0.82 μmol·L−1. Moreover, the total urea flux surrounding the ZJB was 2905 tons·year−1. The seasonal terrestrial source of urea flux contributions had significant seasonal variation in wet, normal, and dry seasons (p &lt; 0.05). The ZJB was subjected to a large flux of urea by estuaries and sewage outlet discharges. The seasonal urea concentration in all stations (&gt;1 μmol·L−1) indicated that urea in the ZJB may have a bioavailable DON source. As a bioavailable nitrogen source, the ability of terrestrial source-derived urea to increase eutrophication should not be ignored in ZJB.

Fabrication of double crosslinked chitosan/gelatin membranes with Na+ and pH dual-responsive controlled permeability

A double crosslinked membrane based on chitosan/gelatin with controlled release properties was synthesized in aqueous system using potassium pyroantimonate and genipin as the ionic crosslinker and covalent crosslinker, respectively. The membranes were characterized using FT-IR, UV–vis spectra, and SEM. In addition, the thermal stability, contact angle, mechanical and controlled release properties were measured. Double-crosslinking significantly improved the hydrophobicity, thermal stability, and tensile strength of the membrane. The permeabilities of various plant nutrients, including urea and plant growth regulators (PGRs), were evaluated. Under the optimized conditions, the lowest flux (J) of urea was 2.89 mg/ (cm2·h). Urea and PGR release were sustained for up to 3 and 10 days, respectively. The release of agrochemicals in this study followed the Weibull model, demonstrating the Na+-responsive and pH-responsive permeability of the films. This dual-responsive film has potential applications in salinized barrier soil, water treatment, separations, and other membrane applications.

173 Characterization of urea transport mechanisms in the intestinal tract of growing pigs

Abstract Previous studies have indicated that pigs are capable of nitrogen salvage via urea recycling, which involves the movement of urea into the gastrointestinal tract and incorporation of nitrogen into endogenous or microbially produced amino acids. Aquaporins (AQP) and urea transporter B (UT-B) have been shown to contribute to urea transport in ruminants; however, it is unclear whether the same processes contribute to urea movement in the intestinal tract of the pig. The objective of this study was to characterize the presence and relative contribution of known urea transporters to urea flux in the growing pig. A total of 9 barrows of 50.8±0.9 kg BW were euthanized and samples of intestinal tissue were obtained from the duodenum, jejunum, ileum, cecum, and colon. All tissue samples were analyzed for mRNA abundance of UT-B and AQP-3, 7, and 10 via qPCR. Immediately after tissue collection, samples from jejunum and cecum were placed in Ussing chambers for analysis of serosal-to-mucosal urea flux using 14C-urea (49.95 kBq). Serosal-to-mucosal urea flux was measured across intestinal tissue samples with no inhibition or with addition of phloretin (1 mM) to inhibit UT-B-mediated transport, NiCl2 (1 mM) to inhibit AQP-mediated transport, or both inhibitors. UT-B was most highly expressed in the cecum (P &amp;lt; 0.05), while AQP-3, 7, and 10 were most highly expressed in the jejunum (P &amp;lt; 0.05). Serosal-to-mucosal urea flux occurred in both the jejunum and the cecum and was higher in the cecum (42.7 vs. 67.8±5.01 µmol/cm2/h; P &amp;lt; 0.05), confirming the capacity for urea recycling into the gut in pigs; however, neither flux rate was influenced by urea transporter inhibitors (P &amp;gt; 0.05). The results of this study indicate that while known urea transporters are present in the gastrointestinal tract of pigs, they do not play a significant role in urea transport.

Characterization of urea transport mechanisms in the intestinal tract of growing pigs.

Pigs are capable of nitrogen salvage via urea recycling, which involves the movement of urea in the gastrointestinal tract. Aquaporins (AQP) and urea transporter B (UT-B) are involved in urea recycling in ruminants; however, their contribution to urea flux in the intestinal tract of the pig is not known. The objective of this study was to characterize the presence and relative contribution of known urea transporters to urea flux in the growing pig. Intestinal tissue samples (duodenum, jejunum, ileum, cecum, and colon) were obtained from nine barrows (50.8 ± 0.9 kg) and analyzed for mRNA abundance of UT-B and AQP-3, -7, and -10. Immediately after tissue collection, samples from the jejunum and cecum were placed in Ussing chambers for analysis of the serosal-to-mucosal urea flux (Jsm-urea) with no inhibition or when incubated in the presence of phloretin to inhibit UT-B-mediated transport, NiCl2 to inhibit AQP-mediated transport, or both inhibitors. UT-B expression was greatest (P < 0.05) in the cecum, whereas AQP-3, -7, and -10 expression was greatest (P < 0.05) in the jejunum. The Jsm-urea was greater in the cecum than the jejunum (67.8 . 42.7 ± 5.01 µmol·cm-2·h-1; P < 0.05), confirming the capacity for urea recycling in the gut in pigs; however, flux rate was not influenced (P > 0.05) by urea transporter inhibitors. The results of this study suggest that, although known urea transporters are expressed in the gastrointestinal tract of pigs, they may not play a significant functional role in transepithelial urea transport.NEW & NOTEWORTHY We characterized the location and contribution of known urea transporters to urea flux in the pig. Aquaporins are located throughout the intestinal tract, and urea transporter B is expressed only in the cecum. Urea flux occurred in both the jejunum and cecum. Transporter inhibitors had no affect on urea flux, suggesting that their contribution to urea transport in the intestinal tract is limited. Further work is required to determine which factors contribute to urea flux in swine.

Size-segregated water-soluble N-bearing species in the land-sea boundary zone of East China

Size-segregated aerosol samples were collected in the land-sea boundary zone of East China, the offshore area of East China Sea (ECS), to determine the concentrations of water-soluble inorganic nitrogen (WSIN), water-soluble organic nitrogen (WSON), urea, and free amino acids (FAAs), to quantify their sources, and to estimate their dry deposition fluxes. The results indicated that the mean concentrations of WSIN, WSON, urea, and FAAs in total suspended particle (TSP) were 547.14 nmol/m3, 258.64 nmol/m3, 12.69 nmol/m3, and 2589.03 pmol/m3, respectively. The WSIN, WSON, and urea concentrations in TSP showed remarkably spatial variation with the higher one during the shipping line (SL) 1 and SL4, while they exhibited the lower ones during SL3 and SL5. The biomass burning and fertilizer application in the continent provided important precursors for WSIN, WSON, and urea in the offshore areas. Besides, O3-related photochemical process also promoted the secondary formation of these species. In contrast, these anthropogenic sources played the minor roles on the relatively remote marine region (SL5). It was interesting to note that the total FAAs during SL5 did not show the lowest concentration, which might be contributed by the bubble bursting on the sea surface and the release of bacteria. The Positive Matrix Factorization (PMF) method identified fertilizer application and secondary formation were important sources for WSIN (35.54% and 27.53%) and WSON (43.07% and 40.46%), respectively. However, fertilizer application and combustion sources played the crucial important roles on urea (65.93% and 19.13%) and FAAs (51.35% and 16.35%). The mean dry deposition fluxes of WSIN, WSON, urea, and FAAs in the ECS offshore area were 206763.57 nmol m−2 d−1, 103763.07 nmol m−2 d−1, 4746.20 nmol m−2 d−1, and 1047.08 nmol m−2 d−1, respectively. The present study revealed that the ambient N-bearing particles in the land-sea boundary zone suffered from the combined effects of continental transport and the release of marine organisms.

Physiological protective action of dissolved organic carbon on ion regulation and nitrogenous waste excretion of zebrafish (Danio rerio) exposed to low pH in ion-poor water.

Dissolved organic carbon (DOC) represents a heterogeneous group of naturally-occurring molecules in aquatic environments, and recent studies have evidenced that optically dark DOCs can exert some positive effects on ionoregulatory homeostasis of aquatic organisms in acidic waters. We investigated the effects of Luther Marsh DOC, a dark allochthonous DOC, on ion regulation and N-waste excretion of zebrafish acutely exposed to either neutral or low pH in ion-poor water. In the first experiment, simultaneous exposure to pH 4.0 and DOC greatly attenuated the stimulation of Na+ diffusive losses (J outNa ), and prevented the blockade of Na+ uptake (J inNa ) seen in zebrafish exposed to pH 4.0 alone, resulting in much smaller disturbances in Na+ net losses (J netNa ). DOC also attenuated the stimulation of net Cl- losses (J netCl ) and ammonia excretion (J netAmm ) during acidic challenge. In the second experiment, zebrafish acclimated to DOC displayed similar regulation of J inNa and J outNa , and, therefore, reduced J netNa at pH 4.0, effects which persisted even when DOC was no longer present. Protective effects of prior acclimation to DOC on J netCl and J netAmm at pH 4.0 also occurred, but were less marked than those on Na+ balance. Urea fluxes were unaffected by the experimental treatments. Overall, these effects were clearly beneficial to the ionoregulatory homeostasis of zebrafish at low pH, and were quite similar to those seen in a recent parallel study using darker DOC from the upper Rio Negro. This suggests that dark allochthonous DOCs share some chemical properties that render fish tolerant to ionoregulatory disturbances during acidic challenge.

Transepithelial Na and Urea Transport Pathways in Rat Inner Medullary Thin Limbs of Henle's Loop

Identification of key transport pathways for Na and urea in thin limbs of Henle's loops will further explain how solute recycling and countercurrent mechanisms are integrated to produce the corticomedullary osmotic gradient. We studied transepithelial Na and urea fluxes by in vivo microperfusion of papillary descending and ascending thin limbs (DTLs and ATLs, respectively) and by in vitro microperfusion of isolated DTLs and ATLs from the upper and lower 50% of the inner medulla (IM). In vitro studies have shown that transepithelial water permeability (Pf) is high in DTLupper and low in DTLlower, ATLupper and ATLlower. In vivo microperfusion studies confirmed the low Pf in papillary DTLs and ATLs. Mean transepithelial Na and urea permeabilities (PNa, Purea) with in vitro lumen perfusion rates of ~30 nl/min in DTLupper were ~60 E‐5 cm/sec and in DTLlower and ATL were ~350 to 450 E‐5 cm/sec; Na and urea fluxes in DTLlower are directly proportional to lumen perfusion rates between 5 to 30 nl/min. In vivo microperfusion studies confirmed the high PNa and Purea in papillary DTLs and ATLs. With in vitro microperfusion, Purea in DTLupper and DTLlower was not inhibited by peritubular phloretin (0.25 mM) indicating that urea fluxes occur independently of known facilitated urea transporters such as UT‐A1, UT‐A2 or UT‐A3. Furthermore, Purea in DTLlower was not inhibited by ouabain (10 mM) or by a lumen‐to‐bath Na concentration gradient (143 mM/lumen and 0 mM/bath) indicating the absence of Na‐dependent urea transport. On the other hand, Purea in DTLupper and DTLlower were almost completely inhibited by peritubular lanthanum (5 mM), with Purea recovering in both segments following 20 min lanthanum washout. Lanthanum (5 mM) also completely inhibited PNa in DTLlower; however, PNa remained completely inhibited following 20 min washout. Activation energy for transepithelial Na and urea transport determined by in vitro microperfusion during 37°‐ to −16° C temperature transitions was found to be low in DTLupper (~1–2 kJ/mol) compared to DTLlower and ATL (~13 kJ/mol). Urea transport in DTLupper and Na and urea transport in DTLlower and ATL likely occur in large part, by way of a plasma membrane or paracellular channel‐like pathway. The distinctive transport characteristics suggest that there are different mechanisms involved with PNa and Purea in upper and lower DTLs, respectively.Support or Funding InformationNIDDK DK083338 (TP)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Freeze Tolerance and Cryoprotection of Erythrocytes from Warm and Cold Acclimated D. chrysoscelis

Dryophytes chrysoscelis (Cope's gray treefrog) is a freeze tolerant frog that survives the freezing of extracellular fluids throughout winter. Prior to freezing, D. chrysoscelis goes through a cold acclimation period during which the temperature, hours of daylight, and food intake decrease. Cold‐acclimated frogs have elevated plasma levels of glycerol and urea, solutes that function as cryoprotectants by minimizing osmotically induced cell damage during freezing and thawing. To minimize damage, urea and glycerol must be taken up by erythrocytes. Erythrocytes from cold‐acclimated frogs have enhanced abundance and membrane localization of an aquaglyceroporin, HC‐3, through which the intra‐ and extracellular flux of glycerol and urea occur. It is hypothesized that erythrocytes frozen in solution containing glycerol/urea would have greater post‐freeze (PF) viability than cells frozen without glycerol or urea. It is further hypothesized that cells frozen with naturally accumulating, and HC‐3 permeating, solutes (glycerol and urea) would have enhanced PF viability compared to cells frozen with solutes that do not accumulate during cold acclimation (glucose, NaCl, sorbitol). In this study, erythrocytes were obtained from warm‐acclimated (22°C) or cold‐acclimated (4°C) frogs, suspended in phosphate buffered saline (PBS) ± solutes, then incubated at 0 or −8°C for 30 minutes. A separate aliquot of cells was frozen in liquid nitrogen for a 100% hemolysis control. Hemolysis was determined by a colorimetric assay. The percentage of free hemoglobin from the total hemoglobin in the erythrocyte sample was calculated and was normalized to the 100% hemolysis control. Viability of the liquid nitrogen sample was taken as 0%. Viability of cells from warm‐acclimated frogs was reduced by 85% when cells were incubated at −8°C in PBS as oppose to 0°C (p&lt;0.001). Cell viability was reduced approximately 50%, when cells from cold‐acclimated frogs were incubated at −8°C compared to 0°C (p&lt;0.001). Erythrocytes from cold‐acclimated frogs have a 2.6‐fold increase in PF viability compared to cells from warm‐acclimated frogs. Cells from warm‐acclimated frogs had a 2.8‐fold increase in PF viability with the addition of urea in the PBS (p&lt;0.005). Cells from cold‐acclimated frogs had a 0.6‐fold increase in PF viability with the addition of urea or glycerol to the PBS (p&lt;0.001 or p&lt;0.005, respectively). The addition of glucose, NaCl, or sorbitol to PBS did not significantly alter PF viability of cells from warm‐ or cold‐acclimated frogs (p&gt;0.05). Enhanced PF viability of cells from cold‐acclimated frogs, compared to cells from warm‐acclimated frogs suggests cold acclimation aids in the freeze tolerance of D. chrysoscelis by mechanisms other than glycerol/urea accumulation. The results of the cryoprotection assay support the involvement of glycerol and urea in the complex cryoprotectant system of D. chrysoscelis.Support or Funding InformationThis research was supported by NSF IOS‐1121457 (CMK and DLG), Schuellein Chair in the Biological Sciences to CMK, and Schuellein Graduate Research Fellowship to LG.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

GRHL2 Is Required for Collecting Duct Epithelial Barrier Function and Renal Osmoregulation.

Collecting ducts make up the distal-most tubular segments of the kidney, extending from the cortex, where they connect to the nephron proper, into the medulla, where they release urine into the renal pelvis. During water deprivation, body water preservation is ensured by the selective transepithelial reabsorption of water into the hypertonic medullary interstitium mediated by collecting ducts. The collecting duct epithelium forms tight junctions composed of barrier-enforcing claudins and exhibits a higher transepithelial resistance than other segments of the renal tubule exhibit. However, the functional relevance of this strong collecting duct epithelial barrier is unresolved. Here, we report that collecting duct-specific deletion of an epithelial transcription factor, grainyhead-like 2 (GRHL2), in mice led to reduced expression of tight junction-associated barrier components, reduced collecting duct transepithelial resistance, and defective renal medullary accumulation of sodium and other osmolytes. In vitro, Grhl2-deficient collecting duct cells displayed increased paracellular flux of sodium, chloride, and urea. Consistent with these effects, Grhl2-deficient mice had diabetes insipidus, produced dilute urine, and failed to adequately concentrate their urine after water restriction, resulting in susceptibility to prerenal azotemia. These data indicate a direct functional link between collecting duct epithelial barrier characteristics, which appear to prevent leakage of interstitial osmolytes into urine, and body water homeostasis.