Apoptotic cell death plays an important role in limiting testicular germ cell population during spermatogenesis and its dysregulation has been shown to be associated with male infertility. The growing evidence on the role of the transcription factor nuclear factor (NF)-kappa B in controlling apoptosis prompted us to investigate NF-kappa B activity in the normal human testis and its role in testis tissue undergoing excessive apoptosis in vitro. In electrophoretic mobility shift assays, low-level constitutive NF-kappa B DNA-binding activity was found and, by immunostaining of the RelA and p50 NF-kappa B subunits, was localized to Sertoli cell nuclei. During in vitro-induced testicular apoptosis, the Sertoli cell nuclear NF-kappa B levels and whole seminiferous tubule NF-kappa B DNA-binding activity increased previous detection of germ cells undergoing apoptosis. The anti-inflammatory drug sulfasalazine effectively suppressed stress-induced NF-kappa B DNA binding and NF-kappa B-mediated I kappa B alpha gene expression. Importantly, concomitantly with inhibiting NF-kappa B, sulfasalazine blocked germ cell apoptosis. These results suggest that during testicular stress Sertoli cell NF-kappa B proteins exert proapoptotic effects on germ cells, which raises the possibility that pharmacological inhibition of NF-kappa B could be a therapeutic target in transient stress situations involving excessive germ cell death.

Purpose: To explore whether the increase observed in referrals to child and adolescent gender identity services (GIDS) has been similar in four Nordic countries and in the UK.

Male germ cells are susceptible to radiation-induced injury, and infertility is a common problem after total-body irradiation. Here we investigated, first, the effects of irradiation on germ cells in mouse testis and, second, the role of sphingosine-1-phosphate (S1P) treatment in radiation-induced male germ cell loss. Irradiation of mouse testes mainly damaged the early developmental stages of spermatogonia. The damage was seen by means of DNA flow cytometry 21 days after irradiation as decreasing numbers of spermatocytes and spermatids with increasing amounts of ionizing radiation (0.1-2.0 Gy). Intratesticular injections of S1P given 1-2 h before irradiation (0.5 Gy) did not protect against short-term germ cell loss as measured by in situ end labeling of DNA fragmentation 16 h after irradiation. However, after 21 days, in the S1P-treated testes, the numbers of primary spermatocytes and spermatogonia at G2 (4C peak as measured by flow cytometry) were higher at all stages of spermatogenesis compared with vehicle-treated testes, indicating protection of early spermatogonia by S1P, whereas the spermatid (1C) populations were similar. In conclusion, S1P appears to protect partially (16%-47%) testicular germ cells against radiation-induced cell death. This warrants further studies aimed at development of therapeutic agents capable of blocking sphingomyelin-induced pathways of germ cell loss.

The necessity of estrogens for male fertility was recently discovered in studies on both estrogen receptor ␣ knockout and aromatase (cyp 19 gene) knockout mice. However, direct testicular effects of estrogens in male reproduction have remained unclear. Here we studied the protein expression of ER␣ and the recently described estrogen receptor ␤ in the human seminiferous epithelium and evaluated the role of 17␤-estradiol, the main physiological estrogen, in male germ cell survival. Interestingly, both estrogen receptors ␣ and ␤ were found in early meiotic spermatocytes and elongating spermatids of the human testis. Furthermore, low concentrations of 17␤-estradiol (10 Ϫ9 and 10 Ϫ10 mol/L) effectively inhibited male germ cell apoptosis, which was induced in vitro by incubating segments of human seminiferous tubules without survival factors (i.e. serum and hormones). Dihydrotestosterone, which, in addition to estradiol, is an end metabolite of testosterone, was also capable of inhibiting testicular apoptosis, but at a far higher concentration (10 Ϫ7 mol/L) than estradiol. Thus, estradiol appears to be a potent germ cell survival factor in the human testis. The novel findings of the present study together with the previously reported indirect effects of estrogens on male germ cells indicate the importance of estrogens for the normal function of the testis. (J Clin Endocrinol Metab 85: 2057-2067, 2000 R ECENT CONCERN over the potentially harmful effects of environmental estrogen-like chemicals on male fertility (1, 2) has aroused growing interest in understanding the physiological effects of estrogens in the male. Although estrogens have been regarded as female steroid hormones, they are now known to have profound effects on both female and male reproductive systems. In males, estrogens are synthesized mainly in the testis, where they are formed from testosterone by the enzyme P450 aromatase. In the rat, there is an age-related change in the aromatization site from Sertoli cells in the immature testis to Leydig cells in the adult testis (3, 4). In various other species, including humans, P450 aromatase has been found to be present in the Leydig cells of the adult testis (5-9). In addition, in the mouse (6), rat (7), brown bear (9), and rooster (10), aromatase has been found in meiotic and postmeiotic germ cells of the testis, predominantly in spermatids.Estrogens exert their cellular effects through estrogen receptors (ER) that exist in at least two subtypes, ER␣ (11, 12) and the recently described ER␤ (13, 14), which differ in the C-terminal ligand-binding domain and in the N-terminal trans-activation domain. These two subtypes of ER have similar high affinities for 17␤-estradiol, but some synthetic and naturally occurring ligands have different relative affinities for ER␣ and ER␤ (15). In the male reproductive tract, ER␣ has been shown to be strongly expressed in the epididymis and efferent ductules (16 -20). It has also been found in the Leydig cells of the rat testis (16), whereas the seminiferous epitheliu...

It has been suggested that apoptosis is controlled by two intracellular sphingolipids, ceramide and sphingosine-1-phosphate (S1P), which are widely distributed in mammalian tissues. In the ovary, S1P was found to effectively block apoptosis caused by cancer therapies. Its role in male germ cell death, however, was unknown. In this study, we investigated the effects of ceramide and S1P on human male germ cell apoptosis. Germ cell death was induced by incubation of segments of seminiferous tubules in vitro. During apoptosis, ceramide levels increased rapidly before appearance of caspase 3 activation and DNA laddering, suggesting a role for ceramide in the induction of germ cell death. Ceramide appeared to regulate an early step of apoptosis because n-acetyl-L-cysteine and blockade of mitochondrial respiration inhibited apoptosis but had no effect on ceramide levels. Moreover, fumonisin B1 (ceramide synthetase inhibitor) did not significantly affect testicular apoptosis. Therefore, elevated ceramide levels are likely to result from breakdown of sphingomyelin rather than from de novo synthesis. Finally, we found that S1P at 1 and 10 micromol/liter suppressed germ cell apoptosis by 30% (P < 0.001). Taken together, sphingolipids appear to play a role in male germ cell apoptosis and can partly be inhibited by S1P.