Childs G V, Rougeau D, Unabia G
Department of Anatomy and Neurosciences, University of Texas Medical Branch, Galveston 77555-1043.
Endocrinology. 1995 Apr;136(4):1595-602. doi: 10.1210/endo.136.4.7895669.
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.
肾上腺切除或促肾上腺皮质激素释放激素（CRH）刺激后，垂体前叶促肾上腺皮质激素细胞数量增加。然而，这是由有丝分裂引起的吗？此外，由于表皮生长因子（EGF）是促肾上腺皮质激素细胞的有效促分泌剂，它也是一种有丝分裂原吗？为了解决这些问题，我们对通过逆流离心富集至88%-97%的促肾上腺皮质激素细胞群体进行了研究，这些细胞在含有和不含有0.1-10 ng/ml EGF的0-10 nM CRH中生长。使用了三种检测方法来检测有丝分裂细胞或细胞数量的变化。一种用于检测DNA合成过程中溴脱氧尿苷摄取的酶免疫测定法[溴脱氧尿苷（BrDU）摄取]显示，在存在0.5 nM CRH或EGF的情况下，光密度读数增加了3倍。两种肽共同作用使光密度增加到基础水平的4.8倍。更高浓度的CRH或EGF未观察到BrDU摄取的进一步增加。通过免疫标记促肾上腺皮质激素细胞对BrDU摄取进行细胞化学检测显示，在3至5天的促肾上腺皮质激素（ACTH）细胞中，18±2%的细胞中有BrDU。在存在0.5 nM CRH或0.5 ng/ml EGF的情况下，该值分别增加到ACTH细胞的37±3%或34±2%。CRH和EGF共同刺激有丝分裂活性增加，因此在暴露1小时后，47±4%的ACTH细胞被标记为BrDU。还使用3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四氮唑溴盐进行细胞生长/细胞死亡测定，以检测同一组富集的促肾上腺皮质激素细胞培养物中总细胞数量或细胞存活率的变化。0.5 nM CRH和0.5 ng/ml EGF均使读数增加到基础读数的1.5至1.7倍。然而，更高浓度并未刺激细胞数量增加，且它们的联合作用并非相加性。这些研究表明，在有限的剂量范围内，CRH和EGF可以是携带ACTH的促肾上腺皮质激素细胞的有丝分裂原。在更高的剂量范围内，它们的分化作用可能会消除分裂细胞并阻碍细胞群体的进一步生长。
