Formation Of Spermatids Research Articles

Mettl1-dependent m7G tRNA modification is essential for maintaining spermatogenesis and fertility in Drosophila melanogaster

Modification of guanosine to N7-methylguanosine (m7G) in the variable loop region of tRNA is catalyzed by the METTL1/WDR4 heterodimer and stabilizes target tRNA. Here, we reveal essential functions of Mettl1 in Drosophila fertility. Knockout of Mettl1 (Mettl1-KO) causes no major effect on the development of non-gonadal tissues, but abolishes the production of elongated spermatids and mature sperm, which is fully rescued by expression of a Mettl1-transgene, but not a catalytic-dead Mettl1 transgene. This demonstrates that Mettl1-dependent m7G is required for spermatogenesis. Mettl1-KO results in a loss of m7G modification on a subset of tRNAs and decreased tRNA abundance. Ribosome profiling shows that Mettl1-KO led to ribosomes stalling at codons decoded by tRNAs that were reduced in abundance. Mettl1-KO also significantly reduces the translation efficiency of genes involved in elongated spermatid formation and sperm stability. Germ cell-specific expression of Mettl1 rescues disrupted m7G tRNA modification and tRNA abundance in Mettl1-KO testes but not in non-gonadal tissues. Ribosome stalling is much less detectable in non-gonadal tissues than in Mettl1-KO testes. These findings reveal a developmental role for m7G tRNA modification and indicate that m7G modification-dependent tRNA abundance differs among tissues.

Deletion of Mgat2 in spermatogonia blocks spermatogenesis.

Identifying factors required for spermatogenesis is important for understanding mechanisms of male fertility. Inactivation of either the Mgat1 or Man2a2 gene leads to a block in spermatogenesis causing infertility in male mice. MGAT1 GlcNAc-transferase initiates complex N-glycan synthesis and MAN2A2 mannosidase generates the substrate for MGAT2 GlcNAc-transferase to form a biantennary complex N-glycan. In this paper, we show that conditional deletion of Mgat2 in spermatogonia via Stra8-iCre caused a novel block in spermatogenesis, largely prior to the formation of round spermatids. Mgat2[-/-] germ cells did not bind the lectins Phaseolus vulgaris leucoagglutinin (L-PHA) or Griffonia simplicifolia II (GSA-II), similar to germ cells lacking MGAT1 and complex N-glycans. However, overall spermatogenic defects were distinct in germ cells with deleted Mgat2 versus Mgat1. In addition, RNA-seq analysis at 15 days after birth revealed a unique transcriptomic landscape in Mgat2[-/-] germ cells with genes required for sperm formation and functions being most downregulated. Bioinformatic analyses using the ingenuity pathway analysis (IPA) algorithm identified ERK and AKT as central activities. Western blot analyses of 15-day germ cell lysates confirmed that both AKT and ERK1/2 signaling were increased by loss of MGAT2 in germ cells. By contrast, Mgat1[-/-] germ cells were previously shown to have reduced ERK signaling and unchanged AKT activity. Therefore, since the loss of all complex N-glycans is common to each mutant model, the different immature N-glycans that accumulate in Mgat2[-/-] versus Mgat1[-/-] germ cells are proposed to be the basis of their unique spermatogenic phenotypes.

Spermatocytes have the capacity to segregate chromosomes despite centriole duplication failure

Centrosomes are the canonical microtubule organizing centers (MTOCs) of most mammalian cells, including spermatocytes. Centrosomes comprise a centriole pair within a structurally ordered and dynamic pericentriolar matrix (PCM). Unlike in mitosis, where centrioles duplicate once per cycle, centrioles undergo two rounds of duplication during spermatogenesis. The first duplication is during early meiotic prophase I, and the second is during interkinesis. Using mouse mutants and chemical inhibition, we have blocked centriole duplication during spermatogenesis and determined that non-centrosomal MTOCs (ncMTOCs) can mediate chromosome segregation. This mechanism is different from the acentriolar MTOCs that form bipolar spindles in oocytes, which require PCM components, including gamma-tubulin and CEP192. From an in-depth analysis, we identified six microtubule-associated proteins, TPX2, KIF11, NuMA, and CAMSAP1-3, that localized to the non-centrosomal MTOC. These factors contribute to a mechanism that ensures bipolar MTOC formation and chromosome segregation during spermatogenesis when centriole duplication fails. However, despite the successful completion of meiosis and round spermatid formation, centriole inheritance and PLK4 function are required for normal spermiogenesis and flagella assembly, which are critical to ensure fertility.

The Role of Tubulin Polymerization-Promoting Protein2 (TPPP2) in Spermatogenesis: A Narrative Review.

Tubulin polymerization-promoting protein2 (TPPP2) is one of the three paralogs of mammalian TPPP proteins. Its possible role in spermatogenesis is described in this narrative review. TPPP2 is expressed specifically in the male reproductive system, mainly in testes and sperm, and also in the epididymis. In testes, TPPP2 is exclusively expressed in elongating spermatids; in the epididymis, it is located in the middle piece of the sperm tail. TPPP2 is involved in spermiogenesis, in steps which are determinative for the formation and morphology of spermatids. The inhibition of TPPP2 decreases sperm motility (the curvilinear velocity of sperms), probably due to influencing mitochondrial energy production since TPPP2 knockout mice possess an impaired mitochondrial structure. There are data on the role of TPPP2 in various mammalian species: human, mouse, swine, and various ruminants; there is a significant homology among TPPP2s from different species. Experiments with Tppp2-/--mice show that the absence of TPPP2 results in decreased sperm count and serious dysfunction of sperm, including decreased motility; however, the in vitro capacitation and acrosome reaction are not influenced. The symptoms show that Tppp2-/--mice may be considered as a model for oligoasthenozoospermia.

Core binding factor subunit β plays diverse and essential roles in the male germline.

Much of the foundation for lifelong spermatogenesis is established prior to puberty, and disruptions during this developmental window negatively impact fertility long into adulthood. However, the factors that coordinate prepubertal germline development are incompletely understood. Here, we report that core-binding factor subunit-β (CBFβ) plays critical roles in prepubertal development and the onset of spermatogenesis. Using a mouse conditional knockout (cKO) approach, inactivation of Cbfb in the male germline resulted in rapid degeneration of the germline during the onset of spermatogenesis, impaired overall sperm production, and adult infertility. Utilizing a different Cre driver to generate another Cbfb cKO model, we determined that the function of CBFβ in the male germline is likely limited to undifferentiated spermatogonia despite expression in other germ cell types. Within undifferentiated spermatogonia, CBFβ regulates proliferation, survival, and overall maintenance of the undifferentiated spermatogonia population. Paradoxically, we discovered that CBFβ also distally regulates meiotic progression and spermatid formation but only with Cbfb cKO within undifferentiated spermatogonia. Spatial transcriptomics revealed that CBFβ modulates cell cycle checkpoint control genes associated with both proliferation and meiosis. Taken together, our findings demonstrate that core programs established within the prepubertal undifferentiated spermatogonia population are necessary for both germline maintenance and sperm production.

Histomorphometric study of seminiferous tubules and epididymis in adult ram

This research set out to examine the epididymis and seminiferous tubules of adult male Rams, use both histological and histomorphometric methods. For this purpose, ten paired of testis of healthy adult local rams were taken from local abattoir in Fallujah. Tissue was removed and cut into tiny pieces from different parts of the testicles before being put in Bouin's solution. The tissue samples were processed by common paraffin technique then stained with routine H & E stain as well as Masson's trichrome stain. According to the results of the present histological analysis, the ram's testis is enclosed by a capsule of thick irregular connective tissue, and its lobules are separated by trabecular and interlobular septa, with seminiferous tubules and interstitial tissue making up each lobule. Spermatogenic cells and Sertoli cells border the seminiferous tubules. Spermatozoa develop from spermatogenic cells. Spermatogonia are the earliest stages of spermatogenic cells; they are tiny, spherical cells with black, spherical nuclei that sit on the basement membrane. Primary spermatocytes, which are bigger cells with often-distinct chromatin, are generated during mitosis in the spermotogonia. Because of their rapid second meiotic division and subsequent formation of haploid spermatids, secondary spermatocytes are seldom seen. Clustered near the seminiferous tubule's lumen, the spermatids are spherical cells with pale nuclei. There are less Sertoli cells (sustentacular cells) than spermatogenic cells. The nucleus is oval or triangular in shape and pale in colour, with a large nucleolus. Leydig cells are found in the connective tissue between neighbouring tubules. Epithelium that is pseudostratified lines the epididymis. The stereocilia are long cytoplasmic structures that extend into the lumen. Circular smooth muscle is present in varying amounts, and the epithelium is supported by a connective tissue lamina propria.In conclusion, the current study concluded that the thickness of the epithelium of epididymis was decrease toward the tail while the thickness of the smooth muscle layer was increase toward the tail.

Regulation of Miwi-mediated mRNA stabilization by Ck137956/Tssa is essential for male fertility

BackgroundSperm is formed through spermiogenesis, a highly complex process involving chromatin condensation that results in cessation of transcription. mRNAs required for spermiogenesis are transcribed at earlier stages and translated in a delayed fashion during spermatid formation. However, it remains unknown that how these repressed mRNAs are stabilized.ResultsHere we report a Miwi-interacting testis-specific and spermiogenic arrest protein, Ck137956, which we rename Tssa. Deletion of Tssa led to male sterility and absence of sperm formation. The spermiogenesis arrested at the round spermatid stage and numerous spermiogenic mRNAs were down-regulated in Tssa−/− mice. Deletion of Tssa disrupted the localization of Miwi to chromatoid body, a specialized assembly of cytoplasmic messenger ribonucleoproteins (mRNPs) foci present in germ cells. We found that Tssa interacted with Miwi in repressed mRNPs and stabilized Miwi-interacting spermiogenesis-essential mRNAs.ConclusionsOur findings indicate that Tssa is indispensable in male fertility and has critical roles in post-transcriptional regulations by interacting with Miwi during spermiogenesis.

Oogenesis and spermatogenesis in round sardinella (Sardinella aurita Valenciennes, 1847) from the eastern part of the Adriatic Sea

Background and purpose: Small pelagic fish species round sardinella (Sardinella aurita) has expanded its habitat during the last few decades towards colder parts of the Mediterranean Sea. In the Adriatic Sea, this investigation is the first attempt to study its gonad development histologically.Materials and methods: S. aurita specimens were sampled monthly from commercial landings of purse seine catches from the eastern part of the Adriatic Sea from November 2007 to January 2009. After biometric parameters were determined, gonads were removed from the fish and weighed. Sex and gonad development were determined macroscopically by the shape and the structure of gonads and afterwards gonads were analysed histologically.Results: The phases of gonadal development of both sexes of S. aurita were: immature, developing, spawning capable, regressing, and regenerating. An annual reproduction cycle based on gamete development revealed five stages of oocyte development: chromatin nucleolar stage, perinucleolar stage, yolk vesicle (cortical alveoli) formation, vitellogenic (yolk) stage, and ripe (mature) stage. Spermatogenesis was described through three stages: spermatocytogenesis (formation of primary spermatocytes from spermatogonia), meiosis (formation of secondary spermatocytes and spermatids) and spermiogenesis.Conclusions: Gonadal histology analyses of both sex of S. aurita indicated summer spawning manner of this species, with a peak in June-August period.

Aspartam ve asesülfam K kullanımının testis yapısına etkilerinin ince yapı düzeyinde incelenmesi

Amaç: Bu çalışma, ülkemizde ve dünyada yaygın olarak kullanılan aspartam ve asesülfam K’nın, sıçanlarda testis dokuları üzerindeki etkilerinin ışık ve elektron mikroskobik düzeyde araştırılması amacıyla yapılmıştır.Gereç ve Yöntem: Çalışmada 60 adet erkek sıçan kullanıldı ve bu denekler rastgele olarak 6 eşit gruba ayrıldı. 1. gruptaki hayvanlar kontrol grubu olarak değerlendirildi; 2.gruptaki hayvanlara 200 mg/kg/gün aspartam; 3.gruptaki hayvanlara 300 mg/kg/gün aspartam; 4. gruptaki hayvanlara 300 mg/kg/gün asesülfam K; 5. gruptaki hayvanlara 600 mg/kg/gün asesülfam K ve son gruba da 300 mg/kg/gün aspartam+300 mg/kg/gün asesülfam K birlikte 8 hafta süreyle verildi. 8. haftanın sonunda deneklerden testis dokuları alındı ve dokular ışık ve elektron mikroskobik olarak değerlendirildi.Bulgular: Aspartam ve asesülfam K kullanımının testis dokusunda dejenerasyona yol açtığı ve spermatogenez sürecinde sağlıklı spermatid oluşumunu engellediği izlendi.Sonuç: Aspartam ve asesülfam K tüketiminin erkek sıçanlarda testis dejenerasyonuna neden olduğu görüldü. Bununla birlikte, erkek üreme sistemindeki aspartam ve asesülfam K toksisitesinin doğrulanması için daha kapsamlı çalışmaların yapılması gerektiği kanaatine varıldı.

N, N-Dimethylacetamide, an FDA approved excipient, acts post-meiotically to impair spermatogenesis and cause infertility in rats

N, N-Dimethylacetamide is an FDA approved solvent widely used in pharmaceutical industry to facilitate the solubility of lipophilic, high molecular weight drugs with poor water solubility. However, the cytotoxic effects of DMA raises the concern about its use in clinical applications. In the present study, we address the effect of DMA on spermatogenesis. Male Sprague Dawley rats were injected intra-peritoneally for 8 weeks, once a week at a dose of 862 mg/kg. Analysis of reproductive parameters revealed that DMA treated animals exhibit spermatid formation defects within the testis describing the characteristics of oligozoospermia. A subsequent decrease in epididymal sperm concentration along with distortion of sperm morphology was observed. The mitochondrial and microtubule organization in the sperm is considerably modified by DMA. This disrupts the sperm kinetics thus decreasing the total and progressive sperm motility. Finally, DMA treatment resulted in loss of fertility. Our results indicate that exposure to DMA has a negative impact on spermatogenesis and leads to infertility in male rats by inhibiting the post meiotic stages of sperm development. Therefore, the use of DMA in humans must be closely monitored.

Kinesin-7 CENP-E regulates chromosome alignment and genome stability of spermatogenic cells

Kinesin-7 CENP-E is an essential kinetochore motor required for chromosome alignment and congression. However, the specific functions of CENP-E in the spermatogenic cells during spermatogenesis remain unknown. In this study, we find that CENP-E proteins are expressed in the spermatogonia, spermatocytes, and the elongating spermatids. CENP-E inhibition by specific inhibitor GSK923295 results in the disruption of spermatogenesis and cell cycle arrest of spermatogenic cells. Both spermatogonia and spermatocytes are arrested in metaphase and several chromosomes are not aligned at the equatorial plate. We find that CENP-E inhibition leads to chromosome misalignment, the spindle disorganization, and the formation of the aneuploidy cells. Furthermore, the inhibition of CENP-E results in the defects in the formation of spermatids, including the sperm head condensation and the sperm tail formation. We have revealed that kinesin-7 CENP-E is essential for chromosome alignment and genome stability of the spermatogenic cells.

Meiotic arrest occurs most frequently at metaphase and is often incomplete in azoospermic men

To establish which meiotic checkpoints are activated in males with severe spermatogenic impairment to improve phenotypic characterization of meiotic defects. Retrospective observational study. University medical center research laboratory and andrology clinic. Forty-eight patients with confirmed spermatogenic impairment (Johnsen scores 3-6) and 15 controls (Johnsen score 10). None. Quantitative assessment of immunofluorescent analyses of specific markers to determine meiotic entry, chromosome pairing, progression of DNA double-strand break repair, crossover formation, formation of meiotic metaphases, metaphase arrest, and spermatid formation, resulting in a novel classification of human meiotic arrest types. Complete metaphase arrest was observed most frequently (27%), and the patients with the highest frequency of apoptotic metaphases also displayed a reduction in crossover number. Incomplete metaphase arrest was observed in 17% of the patients. Only four patients (8%) displayed a failure to complete meiotic chromosome pairing leading to pachytene arrest. Two new types of meiotic arrest were defined: premetaphase and postmetaphase arrest (15% and 13%, respectively). Meiotic arrest in men occurs most frequently at meiotic metaphase. This arrest can be incomplete, resulting in low numbers of spermatids, and often occurs in association with reduced crossover frequency. The phenotyping approach described here provides mechanistic insights to help identify candidate infertility genes and to assess genotype-phenotype correlations in individual cases.

Sub-active Toxicity Effects of Iron Oxide Nanoparticles on Selected Organs in Male Mice

Background: Decreased particle size and increased surface area to volume ratio are beneficial properties of nanoparticles. However, there are contrasting reports on their potential organotoxic effects. The dose de-pendent toxicity effects of iron III oxide nanoparticles (Fe2O3NPs) on the biochemical indices and histology of select-ed organs of adult male mice were investigated. Methods: Fifty six male mice weighing between 25-32g were randomly assigned into 4 groups (n=14), the control/group 1 was given the vehicle/distilled water, while animals in groups (2-4) received 5 mg/kg, 25 mg/kg and 50mg/kg body weights of Fe2O3NPs (INP) respectively by intraperitoneal route of administration for 14 days after which blood samples were drawn for biochemical analysis. Histopathology studies on the effect of graded doses of INP on the architecture of the liver, kidney and testes of mice were carried out. Results: There were significant increases in plasma sodium, creatinine, urea, chloride, albumin, alanine and aspartate aminotransferases (ALT and AST) in the experimental animals after the 7th and 14th days of ad-ministration of graded doses of INP compared with the control group(p<0.05). These changes were dose-dependent. Photomicrographs showed morphological alterations including kupffer cells hyperplasia, hepatocyte necrosis, interstitial nephritis and eosinophilia of the renal tubular cells but only mild reduction in spermatid formation in testes at 25 and 50mg/kg dosage respectively. Conclusion: The Administration of INP brought about dose-dependent alterations in plasma markers of renal functions as well as hepatic enzymes activity. Deleterious morphological changes in architecture of the liver and kidney as well as mild changes in testes of mice were associated with INP administration

MODIFICATION OF FORMS OF SPERM IN THE TESTICLES OF MEN WITH INFLAMMATORY DISEASES OF THE GENITOURINARY SYSTEM

This paper presents information on the process of the formation of germ cells in men with inflammatory diseases of the genital organs that have been exposed to electromagnetic fields.Оbservations of spermatozoa showed that there were more living cells than non-living (respectively, 50.19 % and 48.81 %).During the observation disorder of cell motility was detected in the direction of increasing the number of motionless cells to 37.4 % and sedentary to 56,33 %.The influence of the electromagnetic field led to disruption of the spermatogenesis process at the stage of formation of late spermatids and spermatozoa.As a result of the noncoordinated divergence by acrosome of germ cells, cells with double heads, deformed necks and bifurcated flagella were formed.The sperm heads, which accounted for 32.6 %, subjected to greatest deformation,changes in the structures of the cervix are defined 7.21 % and changes in the structures of the tails – 3.72 %.

New data on spermiogenesis and trepaxonematan axoneme in basal tapeworms (Cestoda, Caryophyllidea, Lytocestidae) parasitizing cyprinid fishes

Monozoic caryophyllidean cestodes, intestinal parasites of cyprinid fishes, represent a group of tapeworms with an unclear evolutionary history. As spermatology may provide phylogenetically important data, the spermiogenesis and ultrastructure of the mature spermatozoon have been investigated using an integrative approach combining transmission electron microscopy, cytochemistry and electron tomography in Khawia rossittensis (Szidat, 1937). The process of spermatid formation is accompanied by the presence of ultrastructural characters not described in traditional models of spermiogenesis, e.g., apical electron-dense material, the two striated roots situated unusually opposite each other, branching of typical striated roots, an intercentriolar body comprising five electron-dense and four electron-lucent layers, rotation of both free flagella and flagellar buds to the median cytoplasmic process at 90°, and a complete proximodistal fusion. The synchronous rotation of both flagellar buds and growing free flagella is an evolutionarily linked pattern favouring the hypothesis that the Caryophyllidea are not ancestral but are secondarily derived from polyzoic forms. Electron tomography analysis has revealed a unique feature of two helicoidal tubular structures in the central electron-dense core of the axoneme of mature spermatozoon. These data provide new insights into the architecture of the 9 + ‘1’ axoneme, which is shared by male gametes of all trepaxonematan Platyhelminthes.

In-vitro differentiation of early pig spermatogenic cells to haploid germ cells.

Spermatogonial stem cells (SSCs) self-renew and contribute genetic information to the next generation. Pig is wildly used as a model animal for understanding reproduction mechanisms of human being. Inducing directional differentiation of porcine SSCs may be an important strategy in exploring the mechanisms of spermatogenesis and developing better treatment methods for male infertility. Here, we established an in-vitro culture model for porcine small seminiferous tubule segments, to induce SSCs to differentiate into single-tail haploid spermatozoa. The culture model subsequently enabled spermatozoa to express the sperm-specific protein acrosin and oocytes to develop to blastocyst stage after round spermatid injection. The addition of retinoic acid (RA) to the differentiation media promoted the efficiency of haploid differentiation. RT-PCR analysis indicated that RA stimulated the expression of Stra8 but reduced the expression of NANOS2 in spermatogonia. Genes involved in post-meiotic development, transition protein 1 (Tnp1) and protamine 1 (Prm1) were upregulated in the presence of RA. The addition of an RA receptor (RAR) inhibitor, BMS439, showed that RA enhanced the expression of cAMP responsive-element binding protein through RAR and promoted the formation of round spermatids. We established an efficient culture system for in-vitro differentiation of pig SSCs. Our study represents a model for human testis disease and toxicology screening. Molecular regulators of SSC differentiation revealed in this study might provide a therapeutic strategy for male infertility.

MIB-1 Is Required for Spermatogenesis and Facilitates LIN-12 and GLP-1 Activity in Caenorhabditis elegans.

Covalent attachment of ubiquitin to substrate proteins changes their function or marks them for proteolysis, and the specificity of ubiquitin attachment is mediated by the numerous E3 ligases encoded by animals. Mind Bomb is an essential E3 ligase during Notch pathway signaling in insects and vertebrates. While Caenorhabditis elegans encodes a Mind Bomb homolog (mib-1), it has never been recovered in the extensive Notch suppressor/enhancer screens that have identified numerous pathway components. Here, we show that C. elegans mib-1 null mutants have a spermatogenesis-defective phenotype that results in a heterogeneous mixture of arrested spermatocytes, defective spermatids, and motility-impaired spermatozoa. mib-1 mutants also have chromosome segregation defects during meiosis, molecular null mutants are intrinsically temperature-sensitive, and many mib-1 spermatids contain large amounts of tubulin. These phenotypic features are similar to the endogenous RNA intereference (RNAi) mutants, but mib-1 mutants do not affect RNAi. MIB-1 protein is expressed throughout the germ line with peak expression in spermatocytes followed by segregation into the residual body during spermatid formation. C. elegans mib-1 expression, while upregulated during spermatogenesis, also occurs somatically, including in vulva precursor cells. Here, we show that mib-1 mutants suppress both lin-12 and glp-1 (C. elegans Notch) gain-of-function mutants, restoring anchor cell formation and a functional vulva to the former and partly restoring oocyte production to the latter. However, suppressed hermaphrodites are only observed when grown at 25°, and they are self-sterile. This probably explains why mib-1 was not previously recovered as a Notch pathway component in suppressor/enhancer selection experiments.

B chromosome influence on macro-spermatid amount in the grasshopper Abracris flavolineata (Orthoptera: Acrididae)

BB chromosomes are additional elements that occur in some individuals of determined populations. These chromosomes are regularly known to behave as a parasitic element that could promote harmful effects to your hosts despite their ability of transmission related to distinct drive mechanisms. One of the effects of B chromosome presence is related to the abnormal formation of spermatids that could has interference on viability of gametes. Here, we tested if B chromosome harboring individuals (1B) of Abracris flavolineata present different rate of abnormal spermatids (presenting ploidy higher than haploidy “n”) in comparison to individuals with no B chromosomes (0B). Abnormal spermatids can be easily recognized by size and number of centriolar adjuncts labeled through silver nitrate treatment followed by Giemsa 5% staining. The counting of 400 spermatids of four individuals from Rio Claro/SP for each condition revealed mean percentage of macro spermatids of 0,625% in 0B samples, while for 1B samples a slightly increase was noticed, with 0,812%. For both conditions diploid spermatids were noticed, but for 1B individuals tetraploidy was also common. Our data, with occurrence of 30% more macro-spermatids in 1B individuals is contrasting with the observed in Eyprepocnemis plorans that showed an increase of almost five times the number of macro-spermatids on B carriers. The data suggests a less harmful effect of B chromosomes in reproduction capacity of A. flavolineata. A next step is to investigate if B chromosome would be present in these abnormal spermatids, indicating a defensive mechanism by the host genome, through B chromosome expulsion.

Melatonin up-regulates the expression of the GATA-4 transcription factor and increases testosterone secretion from Leydig cells through RORα signaling in an in vitro goat spermatogonial stem cell differentiation culture system.

Because androgen function is regulated by its receptors, androgen-androgen receptor signaling is crucial for regulating spermatogenesis. Androgen is mainly testosterone secreted by testis. Based on the results of early studies in goats, the administration of melatonin over an extended period of time increases steroid production, but the underlying mechanism remains unclear. Here, we report the expression of the melatonin membrane receptors MT1 and MT2 and the retinoic acid receptor-related orphan receptor-alpha (RORα) in the goat testis. An in vitro differentiation system using spermatogonial stem cells (SSCs) cultured in the presence of testicular somatic cells was able to support the formation of sperm-like cells with a single flagellum. The addition of 10-7 M melatonin to the in vitro culture system increased RORα expression and considerably improved the efficiency of haploid cell differentiation, and the addition of the RORα agonist CGP52608 significantly increased the testosterone concentration and expression of GATA binding factor 4 (GATA-4). Furthermore, inhibitors of melatonin membrane receptors and a RORα antagonist (T0901317) also led to a considerable reduction in the efficiency of haploid spermatid formation, which was coupled with the suppression of GATA-4 expression. Based on these results, RORα may play a crucial role in enhancing melatonin-regulated GATA-4 transcription and steroid hormone synthesis in the goat spermatogonial stem cell differentiation culture system.

Obesity causes weight increases in prepubertal and pubertal male offspring and is related to changes in spermatogenesis and sperm production in rats.

The effect of obesity on testicular activity in prepubertal and pubertal rats was investigated in the present study. Obesity was induced in adult females by feeding a high-calorie diet (HCD). These females were mated with normal males and were fed an HCD during pregnancy and lactation. The male offspring born to obese mothers and fed an HCD after weaning were found to be obese. Seminiferous tubules of offspring from control mothers (OCM) and offspring from HCD-fed mothers (OHCDM) had the same set of germ cells at different age intervals, namely spermatogonia, leptotene spermatocytes, zygotene spermatocytes, pachytene spermatocytes and round and elongated spermatids on postnatal days (PND) 7, 13, 17, 24 and 36, and on the day of preputial separation, respectively. However, there was a significant decrease in round and elongated spermatids and the epididymal sperm count, coupled with a significant decrease in testosterone and an increase in leptin serum concentrations in OHCDM compared with OCM. These results show that obesity in prepubertal rats does not affect the age-dependent appearance of germ cells according to developmental hierarchy, but it does interfere with spermatid formation, resulting in a reduced sperm count, which may be due to a deficiency of testosterone mediated by hyperleptinaemia.