Identification of genes specifically expressed in adult and fetal testis is important in furthering our understanding of testis development and function. In this study, a novel human transcript, designated human testis cAMP-responsive element-binding protein (htCREB), was identified by hybridization of adult and fetal human testis cDNA probes with a human cDNA microarray containing 9216 clones. The htCREB transcript (GenBank Accession no. AY347527) was expressed at 2.35-fold higher levels in adult human testes than in fetal testes. Sequence and ntBLAST analyses against the human genome database indicated that htCREB was a novel splice variant of human CREB. RT-PCR-based tissue distribution experiments demonstrated that the htCREB transcript was highly expressed in adult human testis and in healthy sperm, but not in testes from patients with Sertoli cell-only syndrome. Taken together, these results suggest that the htCREB transcript is chiefly expressed in germ cells and is most likely involved in spermatogenesis.
Xiaoyan Huang, Jun Zhang, Li Lu, Lanlan Yin, Min Xu, Youqun Wang, Zuomin Zhou and Jiahao Sha
Jun Yin, Bing Ni, Yi-dong Yang, Zhong-wei Tang, Zhi-qi Gao, Lan Feng, Wei-gong Liao and Yu-qi Gao
Autophagy and apoptosis are interlocked in an extensive crosstalk. Our previous study demonstrated that hypotonic hypoxia-induced marked apoptosis of a spermatocyte-derived cell line (GC-2). However, whether hypoxia-induced apoptosis is mediated by inhibition of autophagy under hypoxic conditions remains unclear. In this study, GC-2 cells were cultured in 1% O2 and harvested at different time points. Autophagy was determined by acridine orange staining, cyto-ID staining, mCherry-GFP-LC3B adenovirus transfection and Western blotting for various autophagy markers. Apoptosis was detected by TUNEL staining, flow cytometry, JC-1 staining and Western blotting of apoptosis-related proteins. We found that hypoxia-induced apoptosis of GC-2 cells through mitochondrial and death receptor pathways and inhibited autophagic flux in GC-2 cells in a time-dependent manner. However, while marked autolysosome formation was observed in GC-2 cells before 24-h culture in hypoxic conditions, apparent apoptosis was observed only after 24-h culture in hypoxic conditions. Caspase-8 siRNA treatment induced cell survival, accompanied by induction of the mature autophagosome, acidic vesicular organelle formation and autophagic flux. Furthermore, Beclin-1 overexpression markedly attenuated the impairment of spermatogenesis in mice by inhibiting apoptosis of spermatocytes. The results of this study demonstrate that hypoxia inhibits autophagy, which further enhances hypoxia-induced apoptosis of mouse spermatocytes by promoting caspase-8 activation in a time-dependent manner, suggesting that combined application of apoptosis inhibition and autophagy activation might be a therapeutic strategy for treating hypoxia-induced male infertility.
Shu-Fang Wang, Xi-Hua Chen, Bin He, De-Dong Yin, Hai-Jun Gao, Hao-Qi Zhao, Nan Nan, Shi-Ge Guo, Jian-Bing Liu, Bin Wu and Xiang-Bo Xu
Stress impacts the reproductive axis at the level of the hypothalamus and the pituitary gland, which exert an effect on the ovary. Menstruation is regulated by the hypothalamic–pituitary–ovary (HPO) axis. However, the role of stress in menstruation remains unclear. The objective of this study was to explore the role of stress in endometrial breakdown and shedding, using the pseudopregnant mouse menstrual-like model. Female mice were mated with vasectomized males and labeled day 0.5, upon observation of a vaginal seminal plug. On day 3.5, decidualization was induced in pseudopregnant mice using arachis oil. On day 5.5, pseudopregnant mice with artificial decidualization were placed in restraint tubes for 3 h. The findings indicated that acute restraint stress resulted in the disintegration of the endometrium. While corticosterone concentration in the serum increased significantly due to restraint stress, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and progesterone (P4) levels in the serum decreased significantly. An endometrial histology examination indicated that progesterone implants may rescue P4 decline caused by acute stress and block endometrium breakdown and shedding. In addition, mice were treated with metyrapone, an inhibitor of corticosterone synthesis, 1 h prior to being subjected to restraint stress. Interestingly, metyrapone not only inhibited stress-induced endometrium breakdown and shedding, but also prevented stress-induced reduction of P4, LH and FSH. Furthermore, real-time PCR and western blot showed that mRNA and protein expression of CYP11A1 (cytochrome P450, family 11, subfamily A, polypeptide 1) and steroidogenic acute regulatory protein (StAR), the two rate-limiting enzymes for progesterone synthesis in the ovary, decreased following acute stress. But metyrapone prevented the reduction of StAR expression induced by restraint stress. Overall, this study revealed that acute stress results in an increase in corticosterone, which may inhibit LH and FSH release in the serum and CYP11A1 and StAR expression in the ovary, which finally leads to the breakdown and shedding of the endometrium. These experimental findings, based on the mouse model, may enable further understanding of the effects of stress on menstruation regulation and determine the potential factors affecting stress-associated menstrual disorders.