ISOLATION AND CULTURE OF SERTOLI CELLS FROM THE TESTES OF ADULT SIBERIAN HAMSTERS - ANALYSIS OF PROTEINS SYNTHESIZED AND SECRETED BY SERTOLI CELLS CULTURED FROM HAMSTERS RAISED IN A LONG OR A SHORT PHOTOPERIOD

In the present study, we describe two procedures for isolation and cul ture of Sertoli cells from the testes of adult Siberian hamsters. In p rocedure I, collagenase and pancreatin were used for differential enzy matic digestion of the tissue at 32 degrees C, and effects of several attachment factors were examined. Sertoli cells isolated from adult ha msters were not affected by addition of Na selenite, epidermal growth factor, insulin, and transferrin, with or without 5% fetal bovine seru m, to Dulbecco's modified Eagle's medium +Ham's F-12. Significant incr ease in the yield and plating efficiency of Sertoli cells was achieved by developing a novel procedure (procedure II) for Sertoli cell isola tion. In this procedure, testicular tissue was exposed to only one enz yme (collagenase I) for two consecutive digestions at 37 degrees C, an d the total period of enzymatic exposure was reduced relative to that of procedure I. Another objective of this study was to compare the fun ctions of Sertoli cells isolated from spermatogeneitcally active and i nactive adult testes. Isolated Sertoli cells from 60 +/- 5-day-old ham sters raised in long day conditions (LD, 16L:8D) or in short day condi tions (SD, 6L:18D) were stimulated with FSH and testosterone in the pr esence of S-35-methionine for 24 h on Day 4 of culture. The medium was concentrated, and equal amounts of radioactive proteins from LD and S D hamster Sertoli cell cultures were analyzed by two-dimensional PAGE. Compared to Sertoli cells derived from SD hamsters, Sertoli cells fro m LD animals produced greater amounts of two secretory proteins and a smaller amount of one. The fourth species of secretory protein discemi ble in the gel was present in similar amounts in cultures derived from LD- and SD-raised adults.