Isolation and characterization of populations of mature and immature rat colonocytes

Abstract

A nonenzymatic method is described for the isolation of viable populations of mature and immature rat colonocytes. Histology was used to monitor colocyte dissociation and to systematically characterize the amount of cross-contamination between populations of mature luminal cells and immature crypt cells. The mature colonocytes were 87 +/- 9% pure with respect to contamination from cells from the lower half of the colonic crypt, and the immature populations were 98% pure with respect to contamination with cells from the upper half of the colonic crypt. Neither population contained significant numbers of cells from the lamina propria. Cell viability and synthetic function were maintained for 10-12 h in short-term culture. Alkaline phosphatase activity was 1.59 +/- 0.01-fold higher in the mature cells than in the immature cells, and in vivo [3H]thymidine incorporation was 2.9 +/- 0.4-fold greater in the immature than the mature populations. Immature colonocytes synthesized protein in vitro at a rate of 2.5 +/- 0.4-fold higher than the mature cells, whereas fucoprotein synthetic rates and the secretory products were comparable in the two populations. Cell surface iodination revealed that the major iodinatable cell surface proteins were common to both cell populations. These studies demonstrate that highly enriched populations of mature and immature rat colonocytes that maintain viability and synthetic function in short-term culture can be prepared. The intrinsic rate of protein synthesis is higher in immature colonocytes, and a shift to synthesis of a higher percentage of fucoproteins occurs during colonocyte differentiation. In contrast to results in the small intestine, only modest gradients of differentiation markers and cell surface protein expression were observed between mature and immature colonocytes.