Mechanosensation induced by urinary bladder distensions in response to bladder filling is critical for proper urinary tract function during micturition cycles. Mechanoreceptors in the bladder urothelium play a crucial role. Recent studies have shown that urothelial cells express numerous mechanoreceptors, including ion channels PIEZO1 and PIEZO2, but their distribution in urothelium has not been confirmed. We investigated PIEZO1 and PIEZO2 expression with western blotting and their localization in various urothelial layers and regions of mouse urinary bladder with immunofluorescence. Selection of antibodies, which recognize different epitopes of both PIEZO proteins, was performed after amino acid sequence analysis. We confirmed PIEZO1 expression with one out of three antibodies while neither of the two PIEZO2 antibodies showed appropriate bands by western blottings. Two PIEZO1 antibodies resulted in positive labeling of basal, intermediate and superficial umbrella cells on cryosections. In umbrella cells reaction was confined to their apical plasma membrane. Similar distribution was observed by one PIEZO2 antibody, except that the apical plasma membrane was not distinctly labeled. No differences were detected between various regions of the urinal bladder urothelium in cryosections and in en face preparations for confocal microscope.
Using a variety of antibodies and immunohistochemical methods we have shown PIEZO mechanoreceptors in basal, intermediate and superficial layer of the urothelium in all regions of the mouse urinal bladder. We have established a foundation for further studies of mechanosensory function of the PIEZO proteins in urinary tract diseases that affect normal micturition.
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