Growth and development of ovarian follicles require a series of coordinated events that induce morphological and functional changes within the follicle, leading to cell differentiation and oocyte development. The preantral early antral follicle transition is the stage of follicular development during which gonadotropin dependence is obtained and the progression into growing or atresia of the follicle is made. Follicular growth during this period is tightly regulated by oocyte-granulosatheca cell interactions. A cluster of early expressed genes is required for normal folliculogenesis. Granulosa cell factors stimulate the recruitment of theca cells from cortical stromal cells. Thecal factors promote granulosa cell proliferation and suppress granulosa cell apoptosis. Cell-cell and cell-extracellular matrix interactions influence the production of growth factors in the different follicular compartments (oocyte, granulosa, and theca cells). Several autocrine and paracrine factors are involved in follicular growth and differentiation; their activity is present even at the time of ovulation, decreasing the gap junction communication, and stimulating the theca cell proliferation. In addition, the identification of the factors that promote follicular growth from the preantral stage to the small antral stage may provide important information for the identification for assisted reproduction techniques.
The aim of this study was to elucidate the role of llama seminal plasma in the formation of oviductal sperm reservoirs. Female llamas with follicles in the mature phase were mated with a bulbourethral glands-removed male. Females mated with nonbulbourethral glands-removed males were used as control. Oviducts were obtained by surgery 24 h after mating. The uterotubal junction and isthmus were examined by scanning electron microscopy, and mucopolysaccharides were identified by Alcian blue staining. To know the proteins probably involved in sperm reservoir formation, SDS-PAGE of seminal plasma (8% and 18% resolving gel) was made. Spermatozoa only adhered to the oviductal mucosa surface of uterotubal junction of females mated with nonbulbourethral glands-removed males confirming that seminal plasma and, in particular, bulbourethral secretions are related with the oviductal sperm reservoir formation. Histological sections showed sperm in the lumen, immersed in substance, positive for acid mucopolysaccharides. Alcian blue staining of seminal plasma proteins SDS-PAGE showed a band of high molecular weight containing mucopolysaccharides, only present in nonbulbourethral glands-removed males. Bulbourethral glands would secrete at least eight different proteins that most likely participate in the process of sperm storage in the oviduct.
β‐Nerve growth factor (β‐NGF) is a seminal plasma element, responsible for inducing ovulation in camelids. The main organ of β‐NGF production remains nondescript. The aims of this study were to (a) characterize gene expression and protein localization of β‐NGF and its main receptor tyrosine kinase receptor A (TrKA) in the llama male reproductive tract, and (b) determine whether the seminal β‐NGF interacts with ejaculated sperm by localizing β‐NGF and TrKA in epididymal, ejaculated, and acrosome‐reacted (AR) sperms and, additionally, by identifying β‐NGF presence in sperm‐adsorbed proteins (SAP). Both β‐NGF and TrkA transcripts are widely expressed along the male reproductive tract, with a higher expression level of β‐NGF at prostate (p < 0.05). β‐NGF immunolabeling was only positive for prostate, whereas TrKA label was present in epithelial and muscular cells of testis, prostate, bulbourethral glands, and epididymis. Using an immunofluorescent technique, β‐NGF was colocalized with TrKA in the middle piece of ejaculated and AR sperm. However, only TrKA was observed in epididymal sperm indicating that β‐NGF could have a seminal origin. This was also confirmed by the identification of four β‐NGF isoforms in SAP. This study extends the knowledge about the participation of β‐NGF/TrkA in llama reproduction, providing evidence that may have roles in the regulation of sperm physiology.
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