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Abstract

Glucocorticoid induces apoptosis in rat Leydig cells 
Gao,Hui-Bao; Tong,Ming-Han; Hu,Yan-Qin; Guo,Qiang-Su; Ge,Renshan; Hardy,Matthew P.
Endocrinology 143(1): 130-138
Publication date: 2002


The aim of the present study was to investigate whether glucocorticoidinduces apoptosis in rat Leydig cells. To determine whetherthere are developmental differences in glucocorticoid sensitivity,Leydig cells were isolated at distinct stages of their differentiation[mesenchymal-like progenitors (PLC), immature Leydig cells (ILC),and adult Leydig cells (ALC)] from 21-, 35-, and 90-d-old SpragueDawley rats, respectively. Glucocorticoid induction of apoptosiswas evaluated after both in vitro and in vivo exposures. Inthe first set of experiments, PLC, ILC, and ALC were treatedwith 100 nM corticosterone (CORT) for either 4 or 24 h in vitroand then assessed for labeling with the apoptotic marker annexinV. PLC exposed to CORT had levels of annexin V-fluorescein isothiocyanatelabeling that were unchanged relative to control values at bothtime points (P > 0.05). In contrast, CORT-treated ILC andALC had increased frequencies of apoptosis: in ALC, a 22.1 ±1.7% incidence after 4 h and 30.5 ± 2.3% after 24 h comparedwith 7.4 ± 0.8% in untreated controls (P < 0.05).Similar trends were observed for ILC. Ultrastructural analysisconfirmed that the increase in annexin V labeling was associatedwith characteristic signs of apoptosis, including nuclear fragmentationand formation of apoptotic bodies. A second line of experimentsexamined whether apoptosis was evident in purified Leydig cellsafter administration of CORT in vivo. Male rats were subjectedto bilateral adrenalectomy and were treated with CORT by ipinjection twice daily at doses ranging from 2.5-7.5 mg/100g BW starting 3 d after surgery. The frequency of Leydig cellapoptosis was measured at 12, 24, 48, and 72 h after the firstinjection. Administration of the 2.5-mg dose raised circulatingCORT 5-10 times above normal basal concentrations, andLH levels sampled at these times were not altered in the treated animals.Increased Leydig cell apoptosis was measurable after 24 h oftreatment, with an incidence of 21.1 ± 1.8% in ALC compared with5.7 ± 0.8% in untreated controls (P < 0.05). Sharpreductions in immunocytochemical staining intensity were observedin the treated animals for a Leydig cell marker, 11ß-hydroxysteroiddehydrogenase, which occurred concurrently with decreased serumT levels. This was consistent with the hypothesis that CORT-mediatedinduction of apoptosis leads to declines in Leydig cell numbers,thereby affecting T production. These results suggest that excessiveexposure to CORT initiates apoptosis in rat Leydig cells, potentiallycontributing to suppression of circulating T levels during stressand other conditions in which glucocorticoid concentrationsare elevated.