Cell production in mouse intestinal epithelium measured by stathmokinetic flow cytometry and coulter particle counting

Abstract

The rate of cell production in the crypt population of the intestinal epithelium has been determined previously. However, the complex geometry of the tissue makes determination of the rate of cell production in the epithelium as a whole, by traditional methods, extremely difficult if not impossible. In this report, stathmokinetic flow cytometry was used for the direct determination of the rate of cell production in the mouse intestinal epithelium. Mice were given an intraperitoneal injection of colcemid (2.5 mg/ kg) and killed 25, 45, 65 and 90 min after injection. The percentage of cells with 4N DNA (G₂ and M phase cells) in the epithelium at each time interval was determined with flow cytometry. The results were fitted by linear regression and the rate of cell production derived from the slope of the regression line. Thus, the rate of cell production was found to be 1.86% ± 0.44 (x ± SE) per hour in the jejunal epithelium and 1.66% ± 0.48 per hour in the colonic epithelium. The turnover time of the epithelium as a whole was determined from the inverse of the rate of cell production, and was found to be 53.8 h ± 12.6 in jejunum and 60.2 h ± 17.3 in colon. Coulter particle counting was used to measure the number of cells in the intestinal epithelium. The number of epithelial cells was found to be 1.90 × 10⁷ ± 3.11 × 10⁵ cells per cm² in the jejunum and 8.98 × 10⁶ ± 8.05 × 10⁵ cells per cm² in the colon. A number of other kinetic parameters, which were not available, were derived from the values determined in this study. For example, it was estimated that 3.53 × 10⁵ ± 8.33 × 10⁴ cells were produced per cm² per hour in the jejunal epithelium, and 1.49 × 10⁵ ± 4.45 × 10⁴ cells per cm² per hour in the colonic epithelium. The rate of cell production at the level of the crypt was found to be 25.1 ± 6.44 cells per crypt per hour in the jejunum, while in the colon 16.4 ± 5.20 cells were produced per crypt per hour. In the jejunum, it was concluded that on the average, about 241.0 ± 65.8 cells were lost each hour from a villus. The kinetic profle of the crypt was also derived. There were on the average about 609.0 ± 50.5 cells per crypt in the jejunum and 985.0 ± 77.98 cells per crypt-surface epithelium unit in the colon. Of these, about 310 cells constitute the cycling population in a jejunal crypt while about 305 cells constitute the cycling population in a colonic crypt-surface epithelium unit. The cycling population in a jejunal crypt was distributed approximately as follows: 91 cells in the G₁ phase, 171 cells in the S phase, 48 cells in the G₂ + M phases. The cycling population in a colonic crypt-surface epithelium unit was distributed approximately as follows: 140 cells in the G₁ phase, 124 cells in the S phase, and 41 cells in the G₂ + M phases.