Osmotic regulation of UT-B urea transporters in the RT4 human urothelial cell line.

Alan Farrell,Gavin Stewart

doi:10.14814/phy2.14314

Abstract

Facilitative UT‐B urea transporters play important physiological roles in numerous tissues, including the urino‐genital tract. Previous studies have shown that urothelial UT‐B transporters are crucial to bladder function in a variety of mammalian species. Using the RT4 bladder urothelial cell line, this study investigated the potential osmotic regulation of human UT‐B transporters. Initial end‐point PCR experiments confirmed expression of both UT‐B1 and UT‐B2 transcripts in RT4 cells. Western blotting analysis revealed glycosylated UT‐B protein to be highly abundant and immunolocalization experiments showed it was predominantly located on the plasma membrane. Further PCR experiments suggested that a 48 hr, NaCl‐induced raise in external osmolality increased expression of UT‐B transcripts. Importantly, these NaCl‐induced changes also significantly increased UT‐B protein abundance (p < .01, n = 7, ANOVA), whereas mannitol‐induced changes in external osmolality had no effect (NS, n = 4, ANOVA). Finally, similar increases in both UT‐B RNA expression and protein abundance were observed with urea‐induced changes to external osmolality (p < .05, n = 4, ANOVA). In conclusion, these findings strongly suggest that increases in external osmolality, via either NaCl or urea, can regulate human urothelial UT‐B transporters.

Highlights

Facilitative urea transporters allow rapid movement of urea across plasma cell membranes and, in mammals, are encoded by two genes—Slc14a1 (UT-B) (Olives et al, 1994) and Slc14a2 (UT-A) (You et al, 1993)
It was shown that urea levels were significantly higher in UT-B null urothelium compared to wild type, indicating that UT-B helps remove toxic intracellular urea from bladder urothelial cells (Dong et al, 2013)
Using F1/R5 UT-B primers, it was revealed that UT-B1 was the predominant transcript, with UT-B2 expressed at a lower level (Figure 1b)

Summary

Introduction

Facilitative urea transporters allow rapid movement of urea across plasma cell membranes and, in mammals, are encoded by two genes—Slc14a1 (UT-B) (Olives et al, 1994) and Slc14a2 (UT-A) (You et al, 1993). These transporters play a vital role in a number of physiological processes, including the urinary concentrating mechanism (Fenton, Chou, Stewart, Smith, & Knepper, 2004; Yang, Bankir, Gillespie, Epstein, & Verkman, 2002). UT-B transporter protein has been detected in the bladder urothelial cell layers of various mammalian. It was shown that urea levels were significantly higher in UT-B null urothelium compared to wild type, indicating that UT-B helps remove toxic intracellular urea from bladder urothelial cells (Dong et al, 2013)

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