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The brocket deer (Genus Mazama) is a highly diverse cervid group distributed from Mexico to Argentina, with a downward population trend. However, literature on the basic reproductive biology of the genus is scarce. This work aimed to study biometric, histological and stereological aspects of the testes of Dwarf Red Brocket (Mazama rufina). Testes from free-ranging adult brockets (n = 3) were retrieved from necropsies. Testes were histologically processed. From histological images, several stereological parameters were estimated, and seminiferous epithelium cycle morphology was described. Testes volumes were between 8.2 and 18.4 ml and weights from 8.3 to 19.4 g. Gonadosomatic index (% paired-testes weight to body weight) went from 0.17 to 0.64. The tubular cross-sectional diameter was 179.8 ± 2.8 µm. Estimated volume densities for parenchyma and interstitium were 78.8% and 21.2% respectively. There were (in millions/ml) 96.0 ± 13.1 germ cells and 37.7 ± 6.0 somatic cells. Specific cell densities were (all expressed in millions/ml) as follows: spermatogonia 13.1 ± 4.2; primary spermatocytes 43.1 ± 5.0; round spermatids 36.8 ± 8.0 (lower density near the caudal pole, p

Effective tools for male contraception are important in the control of reproduction in animal populations. The aim of the present study was to evaluate the effects of active immunization against gonadotropin-releasing hormone (GnRH) on male reproductive function assessing testicular morphological changes and serum-gonadotropin levels in pre-pubertal rabbits, guinea pigs and ram lambs. An anti-GnRH vaccine was developed by linking a GnRH-homologous molecule to a tetanus clostridial toxoid (Al(OH)3 coadjuvant). After vaccination protocols testicular morphometry, histopathological alterations and endocrine responses (FSH, LH, testosterone and cortisol serum levels) were evaluated. Testicular volume was significantly reduced in vaccinated animals with respect to the control group in rabbits, guinea pigs and ram lambs (P

The objective of this review is to outline existing artificial mitochondria transfer techniques and to describe the future steps necessary to develop new therapeutic applications in medicine. Inspired by the symbiotic origin of mitochondria and by the cell’s capacity to transfer these organelles to damaged neighbors, many researchers have developed procedures to artificially transfer mitochondria from one cell to another. The techniques currently in use today range from simple coincubations of isolated mitochondria and recipient cells to the use of physical approaches to induce integration. These methods mimic natural mitochondria transfer. In order to use mitochondrial transfer in medicine, we must answer key questions about how to replicate aspects of natural transport processes to improve current artificial transfer methods. Another priority is to determine the optimum quantity and cell/tissue source of the mitochondria in order to induce cell reprogramming or tissue repair, in both in vitro and in vivo applications. Additionally, it is important that the field explores how artificial mitochondria transfer techniques can be used to treat different diseases and how to navigate the ethical issues in such procedures. Without a doubt, mitochondria are more than mere cell power plants, as we continue to discover their potential to be used in medicine.

The understanding of mammalian spermatogenesis niche factors active during sexual development may be leveraged to impact reproduction in farm animals. The aim of this study was to evaluate the effects of r-met-hu/G-CSF (filgrastim) on prepubertal sexual development of guinea pigs (Cavia porcellus) and ram lambs (Ovis aries). Individuals of both species were administered r-met-hu/G-CSF daily for 4 days. During and after administration protocols, testicular function and development were assessed through hematological responses, hormonal profiles (gonadotropins, testosterone and cortisol) testicular morphometry and germ cell kinetics. As expected, r-met-hu/G-CSF acutely mobilized white-lineage blood cells in both species. LH was increased by r-met-hu/G-CSF in guinea pigs (P0.05). In guinea pigs there were no differences in testicular weights and volumes 2-mo after r-met-hu/G-CSF application (P>0.05). However, ram lambs showed a dose-response effect regarding testis weight (P

Spermatogonial stem cells (SSC) are a small self-renewing subpopulation of type A spermatogonia, which for the rest are composed of differentiating cells with a very similar morphology. We studied the development of primary co-cultures of prepubertal bovine Sertoli cells and A spermatogonia and the effect of glial cell line-derived neurotropic factor (GDNF) on the numbers and types of spermatogonia, the formation of spermatogonial colonies and the capacity of the cultured SSC to colonize a recipient mouse testis. During the first week of culture many, probably differentiating, A spermatogonia entered apoptosis while others formed pairs and chains of A spermatogonia. After 1 week colonies started to appear that increased in size with time. Numbers of single (A(s)) and paired (A(pr)) spermatogonia were significantly higher in GDNF treated cultures at Days 15 and 25 (P

Aspermatogenesis is a severe impairment of spermatogenesis in which germ cells are completely lacking or present in an immature form, which results in sterility in approximately 25% of patients. Because assisted reproduction techniques require mature germ cells, biotechnology is a valuable tool for rescuing fertility while maintaining biological fatherhood. However, this process involves, for instance, the differentiation of preexisting immature germ cells or the production/derivation of sperm from somatic cells. This review critically addresses four potential techniques: sperm derivation in vitro, germ stem cell transplantation, xenologous systems, and haploidization. Sperm derivation in vitro is already feasible in fish and mammals through organ culture or 3D systems, and it is very useful in conditions of germ cell arrest or in type II Sertoli-cell-only syndrome. Patients afflicted by type I Sertoli-cell-only syndrome could also benefit from gamete derivation from induced pluripotent stem cells of somatic origin, and human haploid-like cells have already been obtained by using this novel methodology. In the absence of alternative strategies to generate sperm in vitro, in germ cells transplantation fertility is restored by placing donor cells in the recipient germ-cell-free seminiferous epithelium, which has proven effective in conditions of spermatogonial arrest. Grafting also provides an approach for ex-vivo generation of mature sperm, particularly using prepubertal testis tissue. Although less feasible, haploidization is an option for creating gametes based on biological cloning technology. In conclusion, the aforementioned promising techniques remain largely experimental and still require extensive research, which should address, among other concerns, ethical and biosafety issues, such as gamete epigenetic status, ploidy, and chromatin integrity.

Morphometry is a classical quantitative method often used in biology to provide a data basis for functional interpretations/interactions of a particular organ or system. Herein we took advantage of this valuable approach to evaluate the spermatogonial stem cell niche using the horse testis and immunocytochemical localization of GFRA1 [glial cell line-derived neurotrophic factor receptor produced by Sertoli cells)] as an example. Using the NIH ImageJ free software, we describe in detail all the necessary steps to investigate this specific and crucial microenvironment. Based on several recently published papers from our research group, this approach has proved to be fast, simple, and adaptable to a wide range of species and has the potential to be easily reproducible in different laboratories.

The access to sufficient numbers of spermatogonial stem cells (SSCs) is a prerequisite for the study of their regulation and further biomanipulation. A specialized medium and several growth factors were tested to study thein vitrobehavior of bovine type A spermatogonia, a cell population that includes the SSCs and can be specifically stained for the lectin Dolichos biflorus agglutinin. During short-term culture (2 weeks), colonies appeared, the morphology of which varied with the specific growth factor(s) added. Whenever the stem cell medium was used, round structures reminiscent of sectioned seminiferous tubules appeared in the core of the colonies. Remarkably, these round structures always contained type A spermatogonia. When leukemia inhibitory factor (LIF), epidermal growth factor (EGF), or fibroblast growth factor 2 (FGF2) were added, specific effects on the numbers and arrangement of somatic cells were observed. However, the number of type A spermatogonia was significantly higher in cultures to which glial cell line-derived neurotrophic factor (GDNF) was added and highest when GDNF, LIF, EGF, and FGF2 were all present. The latter suggests that a proper stimulation of the somatic cells is necessary for optimal stimulation of the germ cells in culture. Somatic cells present in the colonies included Sertoli cells, peritubular myoid cells, and a few Leydig cells. A transplantation experiment, using nude mice, showed the presence of SSCs among the cultured cells and in addition strongly suggested a more than 10 000-fold increase in the number of SSCs after 30 days of culture. These results demonstrate that bovine SSC self-renew in our specialized bovine culture system and that this system can be used for the propagation of these cells.

Brahman breed bulls (Bos indicus) are widely used to introduce environmental resistance traits into meat-producing herds. However, their reproductive development is slower than European breeds (Bos taurus). The objective of this study was to assess the development of the seminiferous epithelium in Brahman bulls. Twenty-three prepubertal bulls were castrated and testicular samples taken for histological processing. Light microscopic images were digitized and cells of the seminiferous epithelium were assessed. Immature Sertoli cells gradually decreased in numbers and were no longer detected after approximately 14 months of age; concurrently, the numbers of mature Sertoli cells increased from 10 to 14 months. Spermatogenesis started during the ninth month; prior to that, only gonocytes and immature Sertoli cells were observed. Type A spermatogonia, spermatocytes, round spermatids, elongated spermatids and spermatozoa were first detected at 9.5, 11, 11, 13 and 16 months of age, respectively. The delay in the onset of puberty in Brahman bulls with respect to B. taurus was attributed to a longer duration of the prepubertal period (interval from start of spermatogenesis to puberty) and a later start of spermatogenesis.