Non-Aquaporin Water Channels.

Water transport through membrane is so intricate that there are still some debates. AQPs are entirely accepted to allow water transmembrane movement depending on osmotic gradient. Cotransporters and uniporters, however, are also concerned in water homeostasis. UT-B has a single-channel water permeability that is similar to AQP1. CFTR was initially thought as a water channel but now not believed to transport water directly. By cotransporters, such as KCC4, NKCC1, SGLT1, GAT1, EAAT1, and MCT1, water is transported by water osmosis coupling with substrates, which explains how water is transported across the isolated small intestine. This chapter provides information about water transport mediated by other membrane proteins except AQPs.

Localization of aquaporin-3 proteins in the bovine rumen.

Urea nitrogen salvaging is a crucial mechanism that ruminants have evolved to conserve nitrogen. Facilitative urea transporter-B proteins are known to be involved in urea transport across the rumen epithelium and thus efficiently facilitate the urea nitrogen salvaging process. Recently, functional studies have suggested that aquaglyceroporin transporters might also play a significant role in ruminal urea transport and aquaporin-3 (AQP3) protein has previously been detected in rumen tissue. In this current study, we investigated the specific localization of AQP3 transporters in the bovine rumen. First, end-point reverse-transcription PCR experiments confirmed strong AQP3 expression in both bovine rumen and kidney. Immunoblotting analysis using 2 separate anti-AQP3 antibodies detected AQP3 protein signals at 25, 32, and 42-45 kDa. Further immunolocalization studies showed AQP3 protein located in all the layers of rumen epithelium, especially in the stratum basale, and in the basolateral membranes of kidney collecting duct cells. These data confirm that AQP3 transporters are highly abundant within the bovine rumen and appear to be located throughout the ruminal epithelial layers. The physiological significance of the multiple AQP3 proteins detected and their location is not yet clear, hence further investigation is required to determine their exact contribution to ruminal urea transport.

Water Transport Mediated by Other Membrane Proteins.

Water transport through membrane is so intricate that there are still some debates. (Aquaporins) AQPs are entirely accepted to allow water transmembrane movement depending on osmotic gradient. Cotransporters and uniporters , however, are also concerned in water homeotatsis. Urea transporter B (UT-B) has a single-channel water permeability that is similar to AQP1. Cystic fibrosis transmembrane conductance regulator (CFTR ) was initially thought as a water channel but now not believed to transport water directly. By cotranporters, water is transported by water osmosis coupling with substrates, which explains how water is transported across the isolated small intestine. This chapter provides information about water transport mediated by other membrane proteins except AQPs .

Modulation of sheep ruminal urea transport by ammonia and pH.

Ruminal fermentation products such as short-chain fatty acids (SCFA) and CO2 acutely stimulate urea transport across the ruminal epithelium in vivo, whereas ammonia has inhibitory effects. Uptake and signaling pathways remain obscure. The ruminal expression of SLC14a1 (UT-B) was studied using polymerase chain reaction (PCR). The functional short-term effects of ammonia on cytosolic pH (pHi) and ruminal urea transport across native epithelia were investigated using pH-sensitive microelectrodes and via flux measurements in Ussing chambers. Two variants (UT-B1 and UT-B2) could be fully sequenced from ovine ruminal cDNA. Functionally, transport was passive and modulated by luminal pH in the presence of SCFA and CO2, rising in response to luminal acidification to a peak value at pH 5.8 and dropping with further acidification, resulting in a bell-shaped curve. Presence of ammonia reduced the amplitude, but not the shape of the relationship between urea flux and pH, so that urea flux remained maximal at pH 5.8. Effects of ammonia were concentration dependent, with saturation at 5 mmol/l. Clamping the transepithelial potential altered the inhibitory potential of ammonia on urea flux. Ammonia depolarized the apical membrane and acidified pHi, suggesting that, at physiological pH (< 7), uptake of NH4 (+) into the cytosol may be a key signaling event regulating ruminal urea transport. We conclude that transport of urea across the ruminal epithelium involves proteins subject to rapid modulation by manipulations that alter pHi and the cytosolic concentration of NH4 (+). Implications for epithelial and ruminal homeostasis are discussed.

Metabolic Modulation of Intracellular Ammonia via Intravesical Instillation of Nanoporter-Encased Hydrogel Eradicates Bladder Carcinoma.

Tumor protein 53 (TP53) mutation in bladder carcinoma (BC), upregulates the transcription of carbamoyl phosphate synthetase 1 (CPS1), to reduce intracellular ammonia toxicity. To leverage ammonia combating BC, here, an intravesically perfusable nanoporter-encased hydrogel system is reported. A biomimetic fusogenic liposomalized nanoporter (FLNP) that is decorated with urea transporter-B (UT-B) is first synthesized with protonated chitosan oligosaccharide for bladder tumor-targeted co-delivery of urease and small interfering RNA targeting CPS1 (siCPS1). Mussel-inspired hydrogel featured with dual functions of bio-adhesion and injectability is then fabricated as the reservoir for intravesical immobilization of FLNP. It is found that FLNP-mediated UT-B immobilization dramatically induces urea transportation into tumor cells, and co-delivery of urease and siCPS1 significantly boosts ammonia accumulation in tumor inducing cell apoptosis. Treatment with hybrid system exhibits superior anti-tumor effect in orthotopic bladder tumor mouse model and patient-derived xenograft model, respectively. Combined with high-protein diet, the production of urinary urea increases, leading to an augmented intracellular deposition of ammonia in BC cells, and ultimately an enhanced tumor inhibition. Together, the work establishes that cascade modulation of ammonia in tumor cells could induce tumor apoptosis and may be a practical strategy for eradication of TP53-mutated bladder cancer.

Expression of Urea Transporter B in Normal and Injured Brain.

Urea transporter B (UT-B) is a membrane channel protein widely distributed in mammals, and plays a significant physiological role by regulating urea and water transportation in different tissues. More and more studies have found that UT-B is related to neurological diseases, including myelinopathy and depression. When urea accumulates in the brains of UT-B knockout mice, the synaptic plasticity of neurons is reduced, and the morphology and function of glial cells are also changed. However, the distribution and expression change of UT-B remain unclear. The purpose of this study is to determine the expression characteristics of UT-B in the brain. Through single-cell RNA sequencing, UT-B was found to express universally and substantially throughout the various cells in the central nervous system except for endothelial and smooth muscle cells. UT-B was detected in the third cerebral ventricular wall, granule cell layer of the dentate gyrus, and other parts of the hippocampal, cerebral cortex, substantia nigra, habenular, and lateral hypothalamic nucleus by immunohistochemistry. Compared with the membrane expression of UT-B in glial cells, the subcellular localization of UT-B is in the Golgi apparatus of neurons. Further, the expression of UT-B was regulated by osmotic pressure in vitro. In the experimental traumatic brain injury model (TBI), the number of UT-B positive neurons near the ipsilateral cerebral cortex increased first and then decreased over time, peaking at the 24 h. We inferred that change in UT-B expression after the TBI was an adaptation to changed urea levels. The experimental data suggest that the UT-B may be a potential target for the treatment of TBI and white matter edema.

Effects of Dietary-SCFA on Microbial Protein Synthesis and Urinal Urea-N Excretion Are Related to Microbiota Diversity in Rumen.

Two experiments were performed in this study. In Experiment 1, twenty goats were fed with an isonitrogenous diet, containing 28% Non-Fiber Carbohydrate (MNFC group, n = 10) or 14% NFC (LNFC group, n = 10). In the MNFC group, the ruminal concentration of Short Chain Fatty Acids (SCFA) increased, and pH declined. Compared with those in the LNFC group, the microbial protein synthesis in rumen and mRNA abundance of urea transporter B (UT-B) in rumen epithelium increased in the MNFC group, although serum urea-N (SUN) did not differ significantly between groups. Simultaneously, urinal urea-N excretion was reduced in the MNFC group. Significant correlations were found between rumen SCFA and UT-B and between UT-B and urinal urea-N excretion. Furthermore, the abundances of SCFA receptor of GPR41 and GPR43 increased in the rumen epithelium of the MNFC group. These results suggest that increases of SUN transported into the rumen and incorporated into microbial protein and decreases of urinal urea-N excretion are related to ruminal SCFA. This is supported by data from our previous study in which added SCFA on the mucosal side caused increases of urea transport rate (flux Jsm urea) from the blood to the ruminal lumen side. In Experiment 2, we used 16S rRNA Amplicon Sequencing to analyze the structure of the ruminal microbiota community in relation to SCFA. An additional eight goats were assigned into the MNFC (n = 4) and LNFC (n = 4) groups. The dietary ingredients, chemical composition, and feeding regimes were the same as those in Experiment 1. Constrained correspondence analysis (CCA analysis) revealed NFC promoted the expansion of microbiota diversity, particularly of SCFA-producing microbes. The function prediction of 19 upregulated Kyoto Encyclopedia of Genes and Genomes (KEGG) ortholog groups showed an NFC-induced increase of the types and abundances of genes coding for enzymes catalyzing N and fatty acid metabolism. Based on our present and previous investigations, our results indicate that, in goats consuming a MNFC diet, the facilitated urea transport in the rumen and improved urea N salvage are triggered by an expansion of ruminal microbiota diversity and are signaled by ruminal SCFA. This study thus provides new insights into the microbiota involved in the dietary modulation of urea-N salvage in ruminant animals.

Clinical significance of the reduction of UT-B expression in urothelial carcinoma of the bladder.

Urea transporter B (UT-B) is a membrane protein and plays an important role in regulating urea concentration in bladder urothelial cells. It has been reported that UT-B gene mutations were related to bladder carcinogenesis, and UT-B deletion could induce DNA damage and apoptosis in bladder urothelium. However, the functions and clinical significance of UT-B in human bladder cancer remain unknown. The most common type of bladder cancer is urothelial carcinoma (UC). We hypothesized that UT-B expression was related to bladder UC progress. In this study, UT-B was detected using immunohistochemistry in 52 paraffin-embedded specimens of bladder UC and 10 normal urothelium specimens. The results showed that UT-B protein expression in UC tumor cells was significantly lower as compared with normal urothelial cells (P = 0.021). UT-B protein expression was significantly reduced with increasing histological grade (P = 0.010). UT-B protein expression in muscle-invasive stage was significantly lower than in non-muscle-invasive stage (P = 0.014). Taken together, our data suggest that the reduction or loss of UT-B expression may be related to the incidence, progression and invasiveness of bladder UC. UT-B may be a novel diagnostic or prognostic biomarker, as well as a potential therapeutic target in UC of the bladder.