Identification of distinct mesenchymal factors in the developing bladder and urethra
The lower urinary is derived from the endodermal precursor, the cloaca. Despite common embryological origins, the epithelial linings and surrounding mesenchyme of the bladder and urethra show divergence. The bladder forms from the anterior part of the urogenital sinus, a cloacal derivative formed after septation of the cloaca to form the anorectal and urogenital tracts. The urethra elongates posteriorly and shows sex-specific differences as a result of hormonal action. The bladder is a target for regenerative therapies and tissue reconstruction strategies. A significant shortcoming of these strategies has been the inability to recapitulate all layers of bladder epithelial cells, most importantly the apical uroplakin expressing cell layer that forms a tight, impermeable barrier against components in the urine. Epithelial-mesenchymal interactions are critical for organogenesis. Several studies have shown the ability of embryonic bladder mesenchyme to reprogram non-bladder epithelium to resemble bladder tissue at the molecular and cellular level. In this study, we performed bulk RNA-sequencing on the mesenchymal layers of the developing mouse bladder and urethra and identified compartmentalized organ-specific gene expression. Our study identified transcription factor genes and secreted protein genes specific to the bladder or urethra mesenchymal compartments. Additionally, we describe sex-specific differences in the lower urinary tract mesenchyme which are more pronounced between the male and female urethral mesenchyme and less evident in the bladder mesenchyme between sexes. Overall design: The objective of this study was to identify compartment specific gene expression in the developing bladder and urethra. We obtained timed-pregnant CD-1 female mice and collected embryos at embryonic day 16.5. The lower urinary tract, including bladder and pelvic urethra, were dissected from the embryos. The epithelial layer of the bladder and urethra were mechanically separated from the mesenchymal layer after enzymatic digest with trypsin. Male and female samples from a litter were separately pooled. Each litter yielded 4 samples: Female bladder mesenchyme, Female urethral mesenchyme, Male bladder mesenchyme, Male urethral mesenchyme. A total of 4 biological replicates for each sample were obtained from 4 independent litters.