Effects of Dietary Selenium Yeast Supplementation on Oxidative Biomarkers of the Brain and Blood in Goats.

The present study evaluated the effects of dietary selenium yeast (SY) on the brain, CSF, and blood of 30 crossbreed goats (5-6 months of age) of both sexes. After the acclimatization of 2 weeks, they were randomly separated into two groups (n = 15) named C and SY groups. The C group received only a basal diet, while SY received a basal diet along with 0.3 mg/kg/diet of SY (Sel-Plex®) in total 0.035 mg/kg/diet of SY for 10 weeks. Se concentration (µg /g dry weight) in 15 different parts of the goat's brain was accessed, and results showed that the highest concentration was found in the occipital cerebrum (322.0 ± 6.146), whereas the lowest concentration was found in the midbrain (10.33 ± 0.232). Besides, the oxidative biomarkers including GSH (12.13 ± 0.191), GSH-Px (206.7 ± 2.362), GST (23.80 ± 0.279), CAT (14.80 ± 0.279), and SOD (152.5 ± 9.540) were increased in SY as compared to GSH (8.200 ± 0.144), GSH-Px (112.9 ± 1.183), GST (18.93 ± 0.284), CAT (12.53 ± 0.215), and SOD (109.0 ± 1.966) of C. The level of cholesterol was also significantly decreased in the serum of the SY group (84.87 ± 0.960) as compared to C (110.5 ± 0.592). In addition, the cholesterol level in CSF decreased significantly in SY (0.3567 ± 0.016) as compared to C (0.509 ± 0.009). The current research suggests that SY supplementation has improved the brain's antioxidant status, blood biochemistry, and cholesterol levels in both serum and CSF of goats.

Molecular Genomic Study of Inhibin Molecule Production through Granulosa Cell Gene Expression in Inhibin-Deficient Mice.

Inhibin is a molecule that belongs to peptide hormones and is excreted through pituitary gonadotropins stimulation action on the granulosa cells of the ovaries. However, the differential regulation of inhibin and follicle-stimulating hormone (FSH) on granulosa cell tumor growth in mice inhibin-deficient females is not yet well understood. The objective of this study was to evaluate the role of inhibin and FSH on the granulosa cells of ovarian follicles at the premature antral stage. This study stimulated immature wild-type (WT) and Inhibin-α knockout (Inha-/-) female mice with human chorionic gonadotropin (hCG) and examined hCG-induced gene expression changes in granulosa cells. Also, screening of differentially expressed genes (DEGs) was performed in the two groups under study. In addition, related modules to external traits and key gene drivers were determined through Weighted Gene Co-Expression Network Analysis (WGCNA) algorithm. The results identified a number of 1074 and 931 DEGs and 343 overlapping DEGs (ODEGs) were shared in the two groups. Some 341 ODEGs had high relevance and consistent expression direction, with a significant correlation coefficient (r2 = 0.9145). Additionally, the gene co-expression network of selected 153 genes showed 122 nodes enriched to 21 GO biological processes (BP) and reproduction and 3 genes related to genomic pathways. By using principal component analysis (PCA), the 14 genes in the regulatory network were fixed and the cumulative proportion of fitted top three principal components was 94.64%. In conclusion, this study revealed the novelty of using ODEGs for investigating the inhibin and FSH hormone pathways that might open the way toward gene therapy for granulosa cell tumors. Also, these genes could be used as biomarkers for tracking the changes in inhibin and FSH hormone from the changes in the nutrition pattern.

Transcriptomic in silico analysis of bovine Escherichia coli mastitis highlights its immune-related expressed genes as an effective biomarker.

BACKGROUND: Mastitis is one of the major diseases causing economic loss to the dairy industry by reducing the quantity and quality of milk. Thus, the objective of this scientific study was to find new biomarkers based on genes for the early prediction before its severity. METHODS: In the present study, advanced bioinformatics including hierarchical clustering, enrichment analysis, active site prediction, epigenetic analysis, functional domain identification, and protein docking were used to analyze the important genes that could be utilized as biomarkers and therapeutic targets for mastitis. RESULTS: Four differentially expressed genes (DEGs) were identified in different regions of the mammary gland (teat cistern, gland cistern, lobuloalveolar, and Furstenberg's rosette) that resulted in 453, 597, 577, and 636 DEG, respectively. Also, 101 overlapped genes were found by comparing 27 different expressed genes. These genes were associated with eight immune response pathways including NOD-like receptor signaling pathway (IL8, IL18, IL1B, PYDC1) and chemokine signaling pathway (PTK2, IL8, NCF1, CCR1, HCK). Meanwhile, 241 protein-protein interaction networks were developed among overlapped genes. Fifty-seven regulatory events were found between miRNAs, expressed genes, and the transcription factors (TFs) through micro-RNA and transcription factors (miRNA-DEG-TF) regulatory network. The 3D structure docking model of the expressed genes proteins identified their active sites and the binding ligands that could help in choosing the appropriate feed or treatment for affected animals. CONCLUSIONS: The novelty of the distinguished DEG and their pathways in this study is that they can precisely improve the detection biomarkers and treatments techniques of cows' Escherichia coli mastitis disease due to their high affinity with the target site of the mammary gland before appearing the symptoms.

The variation in promoter sequences of the Akt3 gene between cow and buffalo revealed different responses against mastitis.

BACKGROUND: Serine/threonine kinase 3 (AKT3) is a protein-coding gene that is associated with several cattle immune diseases including different tumors and cancers. The objective of this study was to investigate the differences in structures and functions of AKT3 of cow and buffalo cattle. METHODS: The sequence differences of gene-coding sequence (CDS) and core promoter region of AKT3 in cow and buffalo were analyzed by using bioinformatics tools and PCR sequencing. Also, the functional analysis of promoter regulating gene expression by RT-PCR was performed using 500 Holstein cows and buffalos. And, evaluation of AKT3 inflammatory response to the lipopolysaccharide (LPS)-induced mastitis was performed between both species. RESULTS: The results revealed the variation in 6 exons out of 13 exons of the two species of CDS. Also, 4 different regions in 3-kb promoters of the AKT3 gene were significantly different between cow and buffalo species, in which cow's AKT3 promoter sequence region was started from - 371 to - 1247, while in buffalo, the sequence was started from - 371 to - 969 of the promoter crucial region. Thus, the promoter was overexpressed in cows compared to buffaloes. As a result, significant differences (P < 0.05) between the two species in the AKT3 gene expression level related to the LPS stimulation in their mammary epithelial cell line. CONCLUSIONS: This study emphasized the great importance of the structural differences of AKT3 between the animal species on their different responses against immune diseases like mastitis.

Adaptive Molecular Evolution of AKT3 Gene for Positive Diversifying Selection in Mammals.

The V-Akt Murine Thymoma Viral Oncogene Homolog 3 (AKT3) gene is of the serine/threonine-protein kinase family and influences the production of milk fats and cholesterol by acting on the sterol administrative area restricting protein (SREBP). The AKT3 gene is highly preserved in animals, and during lactation in cattle, its expression increases. The AKT3 gene is expressed in the digestive system, mammary gland, and immune cells. A phylogenetic investigation was performed to clarify the evolutionary role of AKT3, by maximum probability. The AKT3 gene sequence data of various mammalian species was evident even with animals undergoing breeding selection. From 39 mammalian species studied, there was a signal of positive diversifying selection with Hominidae at 13Q, 16G, 23R, 24P, 121P, 294K, 327V, 376L, 397K, 445T, and 471F among other codon sites of the AKT3 gene. These sites were codes for amino acids such as arginine, proline, lysine, and leucine indicating major roles for the function of immunological proteins, and in particular, the study highlighted the importance of changes in gene expression of AKT3 on immunity.

Transcriptome profiling of anti-müllerian hormone treated preantral/small antral mouse ovary follicles.

The predisposition for the initiation of folliculogenesis in mammals including humans is programmed to start at fetal life and continues until reproductive capacity. The follicles grow from a pool of primordial follicles which retain the major functions in the entire reproductive life of a female. Anti-müllerian hormone (AMH), a glycoprotein belonging to the transforming growth factor-beta family, has an inhibitory effect on ovarian follicle development. The key regulatory target genes in primordial follicle development are of paramount importance in reproductive biology of female. A systems biology method was used to find regulatory genes performing critical role in primordial follicle development. A complete in-depth bioinformatics analysis was performed to investigate the changes in transcriptome of preantral to small antral mouse follicles treated for 12 h and 24 h with two different concentrations; 50 and 200 ng/ml of AMH, and thereby identify candidate genes in time and concentration manner. Firstly, we found differentially expressed genes that were time and concentration dependent in response to AMH. The network analysis of these differentially expressed genes provided new candidate genes and pathways associated with inhibitory action of AMH on the primordial follicle development. To further emphasize the function of AMH, the key identified genes' protein-protein docking was analyzed and found the intracellular and extracellular protein-protein interaction. This study elucidates one of the novel mechanisms of AMH involvement in inhibition of ovarian follicle development which may lead to prolong productive life in female.

Comparative Analysis of V-Akt Murine Thymoma Viral Oncogene Homolog 3 (AKT3) Gene between Cow and Buffalo Reveals Substantial Differences for Mastitis.

AKT3 gene is a constituent of the serine/threonine protein kinase family and plays a crucial role in synthesis of milk fats and cholesterol by regulating activity of the sterol regulatory element binding protein (SREBP). AKT3 is highly conserved in mammals and its expression levels during the lactation periods of cattle are markedly increased. AKT3 is highly expressed in the intestine followed by mammary gland and it is also expressed in immune cells. It is involved in the TLR pathways as effectively as proinflammatory cytokines. The aims of this study were to investigate the sequences differences between buffalo and cow. Our results showed that there were substantial differences between buffalo and cow in some exons and noteworthy differences of the gene size in different regions. We also identified the important consensus sequence motifs, variation in 2000 upstream of ATG, substantial difference in the "3'UTR" region, and miRNA association in the buffalo sequences compared with the cow. In addition, genetic analyses, such as gene structure, phylogenetic tree, position of different motifs, and functional domains, were performed to establish their correlation with other species. This may indicate that a buffalo breed has potential resistance to disease, environment changes, and airborne microorganisms and some good production and reproductive traits.

Knockdown of melatonin receptor 1 and induction of follicle-stimulating hormone on the regulation of mouse granulosa cell function.

Melatonin receptor 1 (MT1) performs a critical role in the regulation of the animal reproductive system, particularly in follicular growth, and has a considerable effect on reproductive performance. However, the role that MT1 plays in regulating hormones associated with reproduction remains unclear. This study was designed to examine the physiological role of constitutive MT1 silencing and follicle stimulating hormone (FSH) treatment in reproduction, making use of mouse granulosa cells (mGCs) as a model. To understand the constitutive role of MT1 in ovarian physiology, the RNAi-Ready pSIREN-RETROQ-ZsGreen Vector mediated recombinant pshRNA was used to silence MT1 gene expression. Furthermore, we observed that the expression of MT1 was successfully inhibited both at the protein and mRNA levels (P<0.001). We demonstrated that RNAi-B-mediated MT1 down-regulation significantly promoted apoptosis (P<0.001), inhibited proliferation, and regulated the cell cycle at the S-phase; conversely, FSH treatment partially aided the apoptotic effect and improved proliferation but showed a significant effect at the S-phase of the cell cycle. Transitory knockdown of MT1 proved essential in the function of mGCs, as it significantly decreased cyclic adenosine monophospahte (cAMP) level and increased cell apoptosis. Following knockdown of MT1, the expression of Bax was significantly up-regulated (P<0.001), but Bcl-2 was slightly down-regulated, both at the transcriptional and at translational levels. Moreover, the silencing of MT1 and its constitutive effect on FSH significantly promoted an increase in estradiol (P<0.001) and slightly decreased the concentration of progesterone. Together, our data indicates that MT1 suppression leads to interference in the normal physiological function of the ovary by enhancing follicular apoptosis, inhibiting proliferation, and influencing hormonal signaling, whereas constitutive FSH treatment counteracted the negative down-regulatory effects of MT1 on mGCs.

Long Non-Coding RNAs: the New Horizon of Gene Regulation in Ovarian Cancer.

Long non-coding RNAs (lncRNAs), a class of non-coding transcripts, have recently been emerging with heterogeneous molecular actions, adding a new layer of complexity to gene-regulation networks in tumorigenesis. LncRNAs are considered important factors in several ovarian cancer histotypes, although few have been identified and characterized. Owing to their complexity and the lack of adapted molecular technology, the roles of most lncRNAs remain mysterious. Some lncRNAs have been reported to play functional roles in ovarian cancer and can be used as classifiers for personalized medicine. The intrinsic features of lncRNAs govern their various molecular mechanisms and provide a wide range of platforms to design different therapeutic strategies for treating cancer at a particular stage. Although we are only beginning to understand the functions of lncRNAs and their interactions with microRNAs (miRNAs) and proteins, the expanding literature indicates that lncRNA-miRNA interactions could be useful biomarkers and therapeutic targets for ovarian cancer. In this review, we discuss the genetic variants of lncRNAs, heterogeneous mechanisms of actions of lncRNAs in ovarian cancer tumorigenesis, and drug resistance. We also highlight the recent developments in using lncRNAs as potential prognostic and diagnostic biomarkers. Lastly, we discuss potential approaches for linking lncRNAs to future gene therapies, and highlight future directions in the field of ovarian cancer research.

Toxicity of Nanoparticles on the Reproductive System in Animal Models: A Review.

In the last two decades, nanotechnologies demonstrated various applications in different fields, including detection, sensing, catalysis, electronics, and biomedical sciences. However, public concerns regarding the well-being of human may hinder the wide utilization of this promising innovation. Although, humans are exposed to airborne nanosized particles from an early age, exposure to such particles has risen dramatically within the last century due to anthropogenic sources of nanoparticles. The wide application of nanomaterials in industry, consumer products, and medicine has raised concerns regarding the potential toxicity of nanoparticles in humans. In this review, the effects of nanomaterials on the reproductive system in animal models are discussed. Females are particularly more vulnerable to nanoparticle toxicity, and toxicity in this population may affect reproductivity and fetal development. Moreover, various types of nanoparticles have negative impacts on male germ cells, fetal development, and the female reproductive system. These impacts are associated with nanoparticle modification, composition, concentration, route of administration, and the species of the animal. Therefore, understanding the impacts of nanoparticles on animal growth and reproduction is essential. Many studies have examined the effects of nanoparticles on primary and secondary target organs, with a concentration on the in vivo and in vitro effects of nanoparticles on the male and female reproductive systems at the clinical, cellular, and molecular levels. This review provides important information regarding organism safety and the potential hazards of nanoparticle use and supports the application of nanotechnologies by minimizing the adverse effects of nanoparticles in vulnerable populations.

Role and mechanism of AMH in the regulation of Sertoli cells in mice.

Sertoli cells produce anti-Müllerian hormone (AMH), a glycoprotein belonging to the transforming growth factor-beta family. AMH mediates the regression of Müllerian ducts in the developing male fetus. However, the role of AMH in the regulation of primary Sertoli cells remains unclear. The present study was designed to investigate the effect of AMH on the viability and proliferation of Sertoli cells, with an additional focus on stem cell factor (SCF). Treatment of Sertoli cells with increasing concentrations of rh-AMH (0, 10, 50, 100, and 800ng/ml) for two days revealed that AMH, at high concentrations, increased apoptosis. These results were confirmed by a significant increase in Caspase-3 and Bax and a decrease in Bcl-2 protein and mRNA expression (P<0.01). Paradoxically, treatment with a low concentration of rh-AMH (10ng/ml), but not higher concentrations (50-800ng/ml), promoted Sertoli cell proliferation, which was verified by an increase in PCNA mRNA (P<0.05). Furthermore, only low concentrations of rh-AMH activated the non-canonical ERK signaling pathway. Similarly, low concentrations of rh-AMH (10-50ng/ml) significantly increased (P<0.05) SCF mRNA and SCF protein levels. These findings indicate that AMH differentially regulates the fate of Sertoli cells in vitro by promoting proliferation at low concentrations and apoptosis at high concentrations. In addition, AMH increased the expression of SCF, an important regulator of Sertoli cell development. Therefore, AMH may play a role in Sertoli cell development.

Expression, Localization of SUMO-1, and Analyses of Potential SUMOylated Proteins in Bubalus bubalis Spermatozoa.

Mature spermatozoa have highly condensed DNA that is essentially silent both transcriptionally and translationally. Therefore, post translational modifications are very important for regulating sperm motility, morphology, and for male fertility in general. Protein sumoylation was recently demonstrated in human and rodent spermatozoa, with potential consequences for sperm motility and DNA integrity. We examined the expression and localization of small ubiquitin-related modifier-1 (SUMO-1) in the sperm of water buffalo (Bubalus bubalis) using immunofluorescence analysis. We confirmed the expression of SUMO-1 in the acrosome. We further found that SUMO-1 was lost if the acrosome reaction was induced by calcium ionophore A23187. Proteins modified or conjugated by SUMO-1 in water buffalo sperm were pulled down and analyzed by mass spectrometry. Sixty proteins were identified, including proteins important for sperm morphology and motility, such as relaxin receptors and cytoskeletal proteins, including tubulin chains, actins, and dyneins. Forty-six proteins were predicted as potential sumoylation targets. The expression of SUMO-1 in the acrosome region of water buffalo sperm and the identification of potentially SUMOylated proteins important for sperm function implicates sumoylation as a crucial PTM related to sperm function.

Association of MAP4K4 gene single nucleotide polymorphism with mastitis and milk traits in Chinese Holstein cattle.

The objective of the studies presented in this Research Communication was to investigate the association of single nucleotide polymorphisms present in the MAP4K4 gene with different milk traits in dairy cows. Based on previous QTL fine mapping results on bovine chromosome 11, the MAP4K4 gene was selected as a candidate gene to evaluate its effect on somatic cell count and milk traits in ChineseHolstein cows. Milk production traits including milk yield, fat percentage, and protein percentage of each cow were collected using 305 d lactation records. Association between MAP4K4 genotype and different traits and Somatic Cell Score (SCS) was performed using General Linear Regression Model of R. Two SNPs at exon 18 (c.2061T > G and c.2196T > C) with genotype TT in both SNPs were found significantly higher for somatic SCS. We found the significant effect of exon 18 (c.2061T > G) on protein percentage, milk yield and SCS. We identified SNPs at different location of MAP4K4 gene of the cattle and several of them were significantly associated with the somatic cell score and other different milk traits. Thus, MAP4K4 gene could be a useful candidate gene for selection of dairy cattle against mastitis and the identified polymorphisms might potentially be strong genetic markers.

MicroRNAs: New Insight in Modulating Follicular Atresia: A Review.

Our understanding of the post-transcriptional mechanisms involved in follicular atresia is limited; however, an important development has been made in understanding the biological regulatory networks responsible for mediating follicular atresia. MicroRNAs have come to be seen as a key regulatory actor in determining cell fate in a wide range of tissues in normal and pathological processes. Profiling studies of miRNAs during follicular atresia and development have identified several putative miRNAs enriched in apoptosis signaling pathways. Subsequent in vitro and/or in vivo studies of granulosa cells have elucidated the functional role of some miRNAs along with their molecular pathways. In particular, the regulatory roles of some miRNAs have been consistently observed during studies of follicular cellular apoptosis. Continued work should gradually lead to better understanding of the role of miRNAs in this field. Ultimately, we expect this understanding will have substantial benefits for fertility management at both the in vivo or/and in vitro levels. The stable nature of miRNA holds remarkable promise in clinical use as a diagnostic tool and in reproductive medicine to solve the ever-increasing fertility problem. In this review, we summarize current knowledge of the involvement of miRNAs in follicular atresia, discuss the challenges for further work and pinpoint areas for future research.