Preparation of an enriched fraction of luteinizing hormone-secreting cells from a mixed population of rat anterior pituitary cells.

Abstract

To determine whether female rat gonadotropes differentially secrete only LH, only FSH, or both LH and FSH, a new method of preparing an enriched fraction of LH-secreting cells from a mixed population of cells was developed. Dispersed female rat pituitary cells were incubated for 4 h in glass tubes with magnetic beads coated with anti-LH antibody. The tubes were then placed in a magnetic particle concentrator to separate cells bound to antibody-coated beads (518b7-bound) from the rest of the cell population (518b7-unbound). Several controls were used in this experiment: 1) mixed cells immediately plated without any preincubation (control); 2) mixed cells incubated without magnetic beads (incubated-only); 3) mixed cells incubated with magnetic beads but not separated (nonseparated); and 4) mixed cells first incubated with magnetic beads coated with normal mouse serum (NMS) and then separated into NMS-bound and NMS-unbound populations. After removal of magnetic beads from cell populations, the cells were plated at 200,000 cells/well. The 518b7-bound cells secreted 1.5-, 2-, 2-, and 5-fold more LH than control, incubated-only, nonseparated, or 518b7-unbound cells, respectively (p < 0.05); 518b7-bound cells secreted 2.5-, 3.8-, 3.8-, and 11-fold more FSH than control, incubated-only, nonseparated, and 518b7-unbound cells. The ratio of LH:FSH secreted did not vary between the different treatment groups, but the ratio of LH:FSH contained in 518b7-selected cells was greater than that for unbound cells. Our data indicate that we have prepared an enriched fraction of LH-secreting pituitary cells that also secrete FSH but have a greater capacity for producing LH than FSH in culture. These results are supportive of the view that LH and FSH are secreted by the same cells.