HORMONAL-CONTROL OF APOPTOTIC CELL-DEATH IN THE TESTIS - GONADOTROPINS AND ANDROGENS AS TESTICULAR CELL-SURVIVAL FACTORS

Although the requirement for pituitary gonadotropins during testicular cell differentiation is well documented, the possible role of FSH and LH in regulating testicular cell survival has not been studied. Using a quantitative autoradiographic method for the detection of internucleosomal DNA fragmentation, a hallmark feature of apoptosis, the hormonal control of apoptotic cell death was studied in testicular cells collected from immature rats after hypophysectomy. After surgery, animals were treated with daily injections of 20 IU long-acting FSH agonist (FSH-CTP) or 50 IU human CG (hCG) for 2 days. Hypophysectomy decreased testis weight by 25%, but treatment with FSH-CTP or hCG prevented the effect of hypophysectomy. Testes of intact animals contained predominantly high-mol wt DNA, whereas hypophysectomy increased DNA cleavage into low-mol wt (<15 kilobases) ladders characteristic of apoptosis. In contrast, treatment with FSH-CTP or hCG inhibited hypophysectomy-induced apoptotic DNA cleavage by 84% and 51%, respectively. Hypophysectomy-induced DNA fragmentation was found in both interstitial cells and seminiferous tubules. Similar to whole testis, treatment with FSH-CTP suppressed hypophysectomy-induced apoptosis by over 90% in seminiferous tubules and interstitial cells. In contrast, hCG treatment was less effective in preventing hypophysectomy-induced DNA cleavage (46% suppression in tubules and 77% suppression in interstitial cells). Furthermore, testosterone replacement also suppressed hypophysectomy-induced DNA fragmentation by 75% in the whole testis tissue, 64% in tubules, and 55% in interstitial cells. To further study the role of gonadotropins, intact animals were treated with a potent GnRH antagonist (Azaline B, 10 mug/day) to decrease serum gonadotropin levels. This treatment increased testicular DNA fragmentation to levels comparable to those induced by hypophysectomy, suggesting that gonadotropins are the primary pituitary factors regulating testicular cell survival. These data indicate that, in addition to their well known role in stimulating testicular cell differentiation and growth, both FSH and LH/hCG are essential for preventing testicular cell death in both seminiferous tubules and interstitial cells. Furthermore, the suppressive effect of LH/hCG on apoptotic cell death in the seminiferous epithelium may be partially mediated by androgens. Future use of the present autoradiographic method to study hormonal regulation of testicular apoptosis should provide new insight on testis physiology and pathophysiology.