Corticotropin-releasing hormone and epidermal growth factor: mitogens for anterior pituitary corticotropes

Anterior pituitary corticotropes increase in number after stimulation by adrenalectomy or corticotropin-releasing hormone (CRH). However, is this brought about by mitoses? Furthermore, as epidermal growth factor (EGF) is a potent secretagogue for corticotropes, is it also a mitogen? To address these questions, populations of corticotropes enriched to 88-97% by counterflow centrifugation were studied after growth in 0-10 nM CRH with and without 0.1-10 ng/ml EGF. Three types of assays were used to detect changes in mitotic cells or cell number. An enzyme immunoassay for bromodeoxyuridine uptake during DNA synthesis [bromodeoxyuridine (BrDU) uptake] detected a 3-fold increase in optical density readings in the presence of 0.5 nM CRH or EGF. Together the peptides increased the optical density to 4.8-fold basal levels. No further increases in BrDU uptake were seen with higher concentrations of CRH or EGF. Cytochemical detection of BrDU uptake by immunolabeled corticotropes showed BrDU in 18 +/- 2% of 3- to 5-day ACTH cells. In the presence of 0.5 nM CRH or 0.5 ng/ml EGF, this value increased to 37 +/- 3% or 34 +/- 2% of ACTH cells, respectively. Together CRH and EGF stimulated increases in mitotic activity so that 47 +/- 4% of the ACTH cells were labeled for BrDU after a 1-h exposure. Cell growth/cell death assays in 3-(4,5-dimethyltiazol-2-yl)2,5- diphenyl tetrazolium bromide were also used to detect changes in overall cell number or cell survival in the same groups of enriched corticotrope cultures. Both 0.5 nM CRH and 0.5 ng/ml EGF caused increases to 1.5- to 1.7-fold basal readings. However, higher concentrations did not stimulate increases in number, and their combined effects were not additive. These studies show that CRH and EGF can be mitogens for ACTH-bearing corticotropes, in a limited dose range. In a higher dose range, their differentiating effects may eliminate dividing cells and retard further growth of the population.