A simple efficient method for separating murine uterine epithelial and mesenchymal cells

Abstract

A simple, and very efficient, method for isolating pure uterine epithelium from neonatal, immature, or adult mice and pure uterine mesenchyme from neonatal mice is described. The technique uses mild tryptic digestion of the tissues to loosen the adherence of the epithelium to its underlying mesenchyme followed by gentle mechanical manipulation to effect removal of the luminal epithelium as an intact tube of cells. The epithelial fraction collected by this method was free of stromal cell contamination as judged by microscopic examination of the freshly isolated cells and of cell cultures made from that fraction. The mesenchymal fraction was consistently devoid of epithelium when collected from neonatal mice (5 days old or younger), but mesenchyme from uteri of mice greater than or equal to 10 days old was usually contaminated with epithelial cells due to retention of glandular epithelial crypts in the mesenchyme following removal of the luminal epithelium. Both epithelial and mesenchymal cells obtained by this method are viable, as judged by their ability to attach, spread, and synthesize DNA in vitro. Epithelial cells isolated from 20-day-old or adult animals have a full complement of estrogen receptors, as assessed by whole cell uptake of [3H]estradiol. The technique described here has clear advantages over previously described methods for obtaining pure uterine epithelium and in addition allows mesenchymal tissue free of epithelial contamination to be obtained when applied to uteri of animals less than or equal to 5 days old.