Subsequently, these alpha-actin positive cells sequentially acquire the remaining smooth muscle markers. The isolated bladder mesenchyme, bladder mesenchyme recombined with urothelium and intact embryonic bladders were then grown underneath the kidney capsule of syngeneic hosts. The intact 14 day embryonic bladders underwent normal differentiation, defined by the expression of smooth muscle proteins. In contrast, when isolated bladder mesenchyme was grown alone underneath the kidney capsule, the tissue survived, but the mesenchyme did not differentiate into smooth muscle. However, when bladder mesenchyme was recombined with bladder epithelium, smooth muscle differentiation occurred, although this was less organized than if the epithelium was left intact in its natural state.
Epithelium is required for bladder mesenchyme to differentiate into bladder smooth muscle.
The same experiments were repeated in vitro , where intact embryonic bladders prior to smooth muscle differentiation were grown on top of filter paper nourished by standard tissue culture medium. Compared to bladder mesenchyme cultured alone, there was impressive growth of the intact organs. When epithelium was recombined with bladder mesenchyme, again the growth was greatly improved compared to the mesenchyme alone. Labeling of the organ culture with 3H-thymidine showed extensive cellular proliferation in the tissue recombinants composed of bladder mesenchyme plus bladder epithelium and in intact bladders, whereas in the mesenchyme alone, little growth occurred. Histologically, tissue recombinants of bladder mesenchyme plus epithelium structures revealed healthy cellular histology where as the bladder mesenchyme alone showed evidence of cellular degeneration.