Primary culture of prepubertal human testicular cells isolated from testes collected at necropsy.
The aim of the present work was to develop a method for maintaining prepubertal human testicular cells in culture. Seven pairs of testes of boys who died of causes unrelated to endocrine or metabolic diseases were obtained at necropsy. Histology of the testes was normal. Testes were digested with collagenase and dispersed cells were seeded in multi-well dishes in the presence of 5% bovine fetal serum. After the first day, cells were cultured for five days in serum-free medium in the presence or absence of 918 pmol/l insulin. At the end of culture, microscopic examination showed healthy looking cells with characteristics compatible with pre-Sertoli cells; peritubular cells were identified by immunocytochemistry. In the presence of insulin, cells were able to secrete either testosterone or estradiol into the medium, as well as to reveal aromatase activity. In order to study the effect of the time elapsed between death and beginning of cultures, steroidogenic activity was related to this post mortem time. It was found that, in the presence of insulin, cells obtained from testes with less than 24 h of post mortem time secreted testosterone (64 +/- 7.2 pmol/10(6) cells.24 h, mean +/- SD) while cells obtained from testes with more than 24 h of post mortem time did not secrete testosterone. With long post mortem times, aromatase activity under insulin increased from non-detectable to 35 pmol/10(6) cells.24 h. Time course studies showed that cells with capacity to secrete testosterone increase this secretion gradually up to day 10 of culture, while those with detectable aromatase activity showed increments in this activity during the first week of culture.(ABSTRACT TRUNCATED AT 250 WORDS)
Berensztein
E B
EB
Laboratorio de Investigación, Hospital de Pediatría Garrahan, Buenos Aires, Argentina.
Belgorosky
A
A
Rivarola
M A
MA
D001141:Aromatase / Q000378:metabolism; D001344:Autopsy; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D002675:Child, Preschool; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004958:Estradiol / Q000378:metabolism; D005326:Fetal Proteins / Q000494:pharmacology; D006801:Humans; D007150:Immunohistochemistry; D007223:Infant; D007328:Insulin / Q000494:pharmacology; D008297:Male; D013737:Testis / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D013739:Testosterone / Q000378:metabolism; D013997:Time Factors
D016895:Culture Media, Serum-Free; D005326:Fetal Proteins; D007328:Insulin; D013739:Testosterone; D004958:Estradiol; D001141:Aromatase