Characterisation of the morphogenetic course and secretion of two different types of mucoid material by granulated metrial gland/lymphokine-activated killer cells

Abstract

Lymphocytes from mesenteric lymph nodes of ordinary and nude mice were grown in microtitre wells on embryonic mesenchymal-fibroblast monolayers. Human recombinant interleukin-2 (80 units ml-1) was added. Clones of lymphokine-activated killer (LAK) cells developed. The incidence of clone-forming cells was 52-136 per 10(5) cells in lymph nodes from nude mice and 4.2-8.3 per 10(5) cells in lymph nodes from ordinary mice. On a limited number of fibroblast monolayers propagated in culture, the maturing LAK cells were induced to synthesise and secrete 2 types of flowing mucoid material. After methanol fixation and Alcian blue/periodic acid-Schiff (PAS) (at pH 1) staining, the first type of material was distinctively stained turquoise, indicating a highly sulphated proteoglycan, chondroitin sulphate; the second type of material, a macromolecular neutral polysaccharide, was not stained and appeared to have been dissolved. Glycogen was stained deep brilliant purple. After treatment with PAS alone, the chondroitin sulphate was not stained and appeared as a bright area, the neutral polysaccharide mass being stained deep red. This polysaccharide material was characteristically secreted as droplets or 'streamlets' emerging from the cell surface and extending through the first type of material to coalesce with the already accumulated main extracellular mucoid layer spreading between the cells. Clones of secretory LAK cells were obtained from gravid and nongravid mouse uteri as well as from tracheal explants. Change of medium or passage with fresh medium to a new inducing batch of monolayer, at the blastoid-large granular lymphocyte stage (on d 3 to 7), was critical for high reproducibility of secretion. The course of differentiation was found ultimately to be dependent on the embryonic mesenchymal monolayer, suggesting induction by a morphogenetic signal. A correlation can be drawn between the secretory activity and the morphological profile at maturation of highly distinctive organised cells.