Whole genome sequencing revealed genetic diversity, population structure, and selective signature of Panou Tibetan sheep.

BACKGROUND: The detection of selective traits in different populations can not only reveal current mechanisms of artificial selection for breeding, but also provide new insights into phenotypic variation in new varieties and the search for genes associated with important traits. Panou sheep is a cultivated breed of Tibetan sheep in China with stable genetic performance, consistent appearance and fast growth and development after decades of artificial selection and cultivation. Due to long-term adaptation to the high altitude, cold and hypoxic environment in the plateau area, they may have formed a unique gene pool that is different from other Tibetan sheep breeds. To explore the genetic resources of Panou sheep, we used next-generation sequencing technology for the first time to investigate the genome-wide population structure, genetic diversity, and candidate signatures of positive selection in Panou sheep. RESULTS: Comparative genomic analysis with the closely related species Oula sheep (a native breed of Tibetan sheep in China) was used to screen the population selection signal of Panou sheep. Principal component analysis and neighbor joining tree showed that Panou sheep and Oula sheep had differences in population differentiation. Furthermore, analyses of population structure, they came from the same ancestor, and when K = 2, the two populations could be distinguished. Panou sheep exhibit genetic diversity comparable to Oula sheep, as shown by observed heterozygosity, expected heterozygosity and runs of homozygosity. Genome-wide scanning using the Fst and π ratio methods revealed a list of potentially selected related genes in Panou sheep compared to Oula sheep, including histone deacetylase 9 (HDAC9), protein tyrosine kinase 2 (PTK2), microphthalmia-related transcription factor (MITF), vesicular amine transporter 1 (VAT1), trichohyalin-like 1 (TCHHL1), amine oxidase, copper containing 3 (AOC3), interferon-inducible protein 35 (IFI35). CONCLUSIONS: The results suggest that traits related to growth and development and plateau adaptation may be selection targets for the domestication and breeding improvement of Tibetan sheep. This study provides the fundamental footprints for Panou sheep breeding and management.

Identification of copy number variation in Tibetan sheep using whole genome resequencing reveals evidence of genomic selection.

BACKGROUND: Copy number variation (CNV) is an important source of structural variation in the mammalian genome. CNV assays present a new method to explore the genomic diversity of environmental adaptations in animals and plants and genes associated with complex traits. In this study, the genome-wide CNV distribution characteristics of 20 Tibetan sheep from two breeds (10 Oula sheep and 10 Panou sheep) were analysed using whole-genome resequencing to investigate the variation in the genomic structure of Tibetan sheep during breeding. RESULTS: CNVs were detected using CNVnator, and the overlapping regions of CNVs between individual sheep were combined. Among them, a total of 60,429 CNV events were detected between the indigenous sheep breed (Oula) and the synthetic sheep breed (Panou). After merging the overlapping CNVs, 4927 CNV regions (CNVRs) were finally obtained. Of these, 4559 CNVRs were shared by two breeds, and there were 368 differential CNVRs. Deletion events have a higher percentage of occurrences than duplication events. Functional enrichment analysis showed that the shared CNVRs were significantly enriched in 163 GO terms and 62 KEGG pathways, which were mainly associated with organ development, neural regulation, immune regulation, digestion and metabolism. In addition, 140 QTLs overlapped with some of the CNVRs at more than 1 kb, such as average daily gain QTL, body weight QTL, and total lambs born QTL. Many of the CNV-overlapping genes such as PPP3CA, SSTR1 and FASN, overlap with the average daily weight gain and carcass weight QTL regions. Moreover, VST analysis showed that XIRP2, ABCB1, CA1, ASPA and EEF2 differed significantly between the synthetic breed and local sheep breed. The duplication of the ABCB1 gene may be closely related to adaptation to the plateau environment in Panou sheep, which deserves further study. Additionally, cluster analysis, based on all individuals, showed that the CNV clustering could be divided into two origins, indicating that some Tibetan sheep CNVs are likely to arise independently in different populations and contribute to population differences. CONCLUSIONS: Collectively, we demonstrated the genome-wide distribution characteristics of CNVs in Panou sheep by whole genome resequencing. The results provides a valuable genetic variation resource and help to understand the genetic characteristics of Tibetan sheep. This study also provides useful information for the improvement and breeding of Tibetan sheep in the future.

Differential tissue expression of sex steroid-synthesizing enzyme CYP11A1 in male Tibetan sheep (Ovis aries).

Steroid metabolism is a fundament to testicular development and function. The cytochrome P450, family 11, subfamily A, polypeptide 1 (CYP11A1) is a key rate-limiting enzyme for catalyzing the conversion of cholesterol to pregnenolone. However, despite its importance, what expression and roles of CYP11A1 possesses and how it regulates the testicular development and spermatogenesis in Tibetan sheep remains largely unknown. Based on this, we evaluated the expression and localization patterns of CYP11A1 in testes and epididymides of Tibetan sheep at three developmental stages (three-month-old, pre-puberty; one-year-old, sexual maturity and three-year-old, adult) by quantitative real-time PCR (qPCR), western blot and immunofluorescence. The results showed that CYP11A1 mRNA and protein were expressed in testes and epididymides throughout the development stages and obviously more intense in one- and three-year-old groups than three-month-old group (except for the caput epididymidis). Immunofluorescence assay showed that the CYP11A1 protein was mainly located in Leydig cells and epididymal epithelial cells. In addition, positive signals of CYP11A1 protein were observed in germ cells, epididymal connective tissue and sperms stored in the epididymal lumen. Collectively, these results suggested that the CYP11A1 gene might be mainly involved in regulating spermatogenesis and androgen synthesis in developmental Tibetan sheep testis and epididymis.

Responses of rhizosphere microbial community structure and metabolic function to heavy metal coinhibition.

Metal mineral mining results in releases of large amounts of heavy metals into the environment, and it is necessary to better understand the response of rhizosphere microbial communities to simultaneous stress from multiple heavy metals (HMs), which directly impacts plant growth and human health. In this study, by adding different concentrations of cadmium (Cd) to a soil with high background concentrations of vanadium (V) and chromium (Cr), the growth of maize during the jointing stage was explored under limiting conditions. High-throughput sequencing was used to explore the response and survival strategies of rhizosphere soil microbial communities to complex HM stress. The results showed that complex HMs inhibited the growth of maize at the jointing stage, and the diversity and abundance of maize rhizosphere soil microorganisms were significantly different at different metal enrichment levels. In addition, according to the different stress levels, the maize rhizosphere attracted many tolerant colonizing bacteria, and cooccurrence network analysis showed that these bacteria interacted very closely. The effects of residual heavy metals on beneficial microorganisms (such as Xanthomonas, Sphingomonas, and lysozyme) were significantly stronger than those of bioavailable metals and soil physical and chemical properties. PICRUSt analysis revealed that the different forms of V and Cd had significantly greater effects on microbial metabolic pathways than all forms of Cr. Cr mainly affected the two major metabolic pathways: microbial cell growth and division and environmental information transmission. In addition, significant differences in rhizosphere microbial metabolism under different concentrations were found, and this can serve as a reference for subsequent metagenomic analysis. This study is helpful for exploring the threshold for the growth of crops in toxic HM soils in mining areas and achieving further biological remediation.

Intelligent Fault Identification for Rolling Bearings Fusing Average Refined Composite Multiscale Dispersion Entropy-Assisted Feature Extraction and SVM with Multi-Strategy Enhanced Swarm Optimization.

Rolling bearings act as key parts in many items of mechanical equipment and any abnormality will affect the normal operation of the entire apparatus. To diagnose the faults of rolling bearings effectively, a novel fault identification method is proposed by merging variational mode decomposition (VMD), average refined composite multiscale dispersion entropy (ARCMDE) and support vector machine (SVM) optimized by multistrategy enhanced swarm optimization in this paper. Firstly, the vibration signals are decomposed into different series of intrinsic mode functions (IMFs) based on VMD with the center frequency observation method. Subsequently, the proposed ARCMDE, fusing the superiorities of DE and average refined composite multiscale procedure, is employed to enhance the ability of the multiscale fault-feature extraction from the IMFs. Afterwards, grey wolf optimization (GWO), enhanced by multistrategy including levy flight, cosine factor and polynomial mutation strategies (LCPGWO), is proposed to optimize the penalty factor C and kernel parameter g of SVM. Then, the optimized SVM model is trained to identify the fault type of samples based on features extracted by ARCMDE. Finally, the application experiment and contrastive analysis verify the effectiveness of the proposed VMD-ARCMDE-LCPGWO-SVM method.

Annual Characteristics, Source Analysis of PM1-Bound Potentially Harmful Elements in the Eastern District of Chengdu, China.

Chengdu, a megacity in southwestern China, experiences severe air pollution; however, knowledge of the seasonal variation in mass concentration, extent of potentially harmful elements (PHEs) contamination, and sources caused by heavy metals remains lacking. This study adopted a weighting method to calculate the daily mass concentration of PM1 and used ICP-MS to determine PHE concentrations. Results indicated that PM1 mass concentration was in the range 5.44-105.91 µg/m3. Seasonal PM1 mass concentration could be arranged in the following order: winter > fall > spring > summer. The concentrations of PHEs in the PM1 sample mostly showed the same seasonal variation characteristics as mass concentration. The average concentration of each PHE decreased as follows: Cu (107.44) > Zn (81.52) > Pb (22.04) > As (8.17) > Sb (1.91) > Ni (1.87) > Cr(VI) (0.84) > Cd (0.40) > Tl (0.33) (ng/m3). Enrichment factor values varied markedly from mild to anomalous enrichment. Principal component analysis revealed mainly derived from the fossil fuel combustion (55.215%).

Multiscale engineered artificial compact bone via bidirectional freeze-driven lamellated organization of mineralized collagen microfibrils.

Bone, renowned for its elegant hierarchical structure and unique mechanical properties, serves as a constant source of inspiration for the development of synthetic materials. However, achieving accurate replication of bone features in artificial materials with remarkable structural and mechanical similarity remains a significant challenge. In this study, we employed a cascade of continuous fabrication processes, including biomimetic mineralization of collagen, bidirectional freeze-casting, and pressure-driven fusion, to successfully fabricate a macroscopic bulk material known as artificial compact bone (ACB). The ACB material closely replicates the composition, hierarchical structures, and mechanical properties of natural bone. It demonstrates a lamellated alignment of mineralized collagen (MC) microfibrils, similar to those found in natural bone. Moreover, the ACB exhibits a similar high mineral content (70.9 %) and density (2.2 g/cm3) as natural cortical bone, leading to exceptional mechanical properties such as high stiffness, hardness, and flexural strength that are comparable to those of natural bone. Importantly, the ACB also demonstrates excellent mechanical properties in wet, outstanding biocompatibility, and osteogenic properties in vivo, rendering it suitable for a broad spectrum of biomedical applications, including orthopedic, stomatological, and craniofacial surgeries.

CBCT-based three-dimensional dual-phase vascular image fusion: a novel technique for interventional real-time TIPS guidance.

BACKGROUND: Due to the invisibility of the portal vein (PV), how to puncture the PV accurately and safely in transjugular intrahepatic portosystemic shunt (TIPS) creation remains a challenge of the procedure. OBJECTIVES: We aimed to provide the first evaluation of the safety, feasibility, and efficiency of cone beam computed tomography (CBCT)-based three-dimensional (3D) dual-phase vascular image fusion for interventional real-time guided PV puncture during TIPS procedures. MATERIALS AND METHODS: From January 2021 to May 2021, 13 patients undergoing TIPS were prospectively enrolled in this study. Images of the hepatic artery (HA) and PV in 3D were acquired and overlaid on interventional fluoroscopy images in a dual-phase display mode for real-time PV puncture guidance. The number of PV puncture attempts, puncture time, overlaid image accuracy, dose area product, fluoroscopy time, and interventional complications were recorded. RESULTS: Portal vein puncture guided by CBCT-based 3D dual-phase vascular image fusion was successfully performed on 92.3% (12/13) patients. The mean number of PV puncture attempts was 1.8 ± 0.7 (1-3). The mean puncture time and fluoroscopy time was 3.5 ± 1.2 (2-6) min and 25.1 ± 9.4 (15-45) min, respectively. The mean dose area product was 39.49 ± 7.88 (28.81-52.87) mGym2. The error between the reference position of the fusion image and the interventional PV angiography image was less than 0.5 cm. No interventional complication was observed. CONCLUSION: Our results show that 3D dual-phase vascular image fusion might be a safe and feasible technique for interventional real-time guided PV puncture during TIPS. This novel technique might help to reduce the number of PV puncture attempts and the puncture time as well as lower the risks of interventional complications.

Characterization of middle ear soft tissue damping and its role in sound transmission.

Damping plays an important role in the middle ear (ME) sound transmission system. However, how to mechanically characterize the damping of ME soft tissues and the role of damping in ME sound transmission have not yet reached a consensus. In this paper, a finite element (FE) model of the partial external and ME of the human ear, considering both Rayleigh damping and viscoelastic damping for different soft tissues, is developed to quantitatively investigate the damping in soft tissues effects on the wide-frequency response of the ME sound transmission system. The model-derived results can capture the high-frequency (above 2 kHz) fluctuations and obtain the 0.9 kHz resonant frequency (RF) of the stapes velocity transfer function (SVTF) response. The results show that the damping of pars tensa (PT), stapedial annular ligament (SAL) and incudostapedial joints (ISJ) can help smooth the broadband response of the umbo and stapes footplate (SFP). It is found that, between 1 and 8 kHz, the damping of the PT increases the magnitude and phase delay of the SVTF above 2 kHz while the damping of the ISJ can avoid excessive phase delay of the SVTF, which is important in maintaining the synchronization in high-frequency vibration but has not been revealed before. Below 1 kHz, the damping of the SAL plays a more important role, and it can decrease the magnitude but increases the phase delay of the SVTF. This study has implications for a better understanding of the mechanism of ME sound transmission.

Origins of brain tissue elasticity under multiple loading modes by analyzing the microstructure-based models.

Constitutive behaviors and material properties of brain tissue play an essential role in accurately modeling its mechanical responses. However, the measured mechanical behaviors of brain tissue exhibit a large variability, and the reported elastic modulus can differ by orders of magnitude. Here we develop the micromechanical models based on the actual microstructure of the longitudinally anisotropic plane of brain tissue to investigate the microstructural origins of the large variability. Specifically, axonal fiber bundles with the specified configurations are distributed in an equivalent matrix. All micromechanical models are subjected to multiple loading modes, such as tensile, compressive, and shear loading, under periodic boundary conditions. The predicted results agree well with the experimental results. Furthermore, we investigate how brain tissue elasticity varies with its microstructural features. It is revealed that the large variability in brain tissue elasticity stems from the volume fraction of axonal fiber, the aspect ratio of axonal fiber, and the distribution of axonal fiber orientation. The volume fraction has the greatest impact on the mechanical behaviors of brain tissue, followed by the distribution of axonal fiber orientation, then the aspect ratio. This study provides critical insights for understanding the microstructural origins of the large variability in brain tissue elasticity.

Experimentally characterizing the spatially varying anisotropic mechanical property of cancellous bone via a Bayesian calibration method.

Traditional experimental tests for characterizing bone's mechanical properties usually hypothesize a uniaxial stress condition without quantitatively evaluating the influence of spatially varying principal material orientations, which cannot accurately predict the mechanical properties distribution of bones in vivo environment. In this study, a Bayesian calibrating procedure was developed using quantified multiaxial stress to investigate cancellous bone's local anisotropic elastic performance around joints as the spatial variation of main bearing orientations. First, the bone cube specimens from the distal femur of sheep are prepared using traditional anatomical axes. The multiaxial stress state of each bone specimen is calibrated using the actual principal material orientations derived from fabric tensor at different anatomical locations. Based on the calibrated multiaxial stress state, the process of identifying mechanical properties is described as an inverse problem. Then, a Bayesian calibration procedure based on a surrogate constitutive model was developed via multiaxial stress correction to identify the anisotropic material parameters. Finally, a comparison between the experiment and simulation results is discussed by applying the optimal model parameters obtained from the Bayesian probability distribution. Compared to traditional uniaxial methods, our results prove that the calibration based on the spatial variation of the main bearing orientations can significantly improve the accuracy of characterizing regional anisotropic mechanical responses. Moreover, we determine that the actual mechanical property distribution is influenced by complicated mechanical stimulation. This study provides a novel method to evaluate the spatially varying mechanical properties of bone tissues enduring complex mechanical loading accurately and effectively. It is expected to provide more realistic mechanical design targets in vivo for a personalized artificial bone prosthesis in clinical treatment.

Analyses of widely targeted metabolic profiling reveals mechanisms of metabolomic variations during Tibetan sheep (Ovis aries) testis development.

In mammals, the testis is the organ with the highest transcriptional activity. After gene transcription, translation, and post-translational protein modification, the transcriptional results are finally presented at the metabolic level. Metabolites not only essential for cell signaling and energy transfer, but also directly influenced by the physiological and pathological changes in tissues and accurately reflect the physiological changes. The fact that the testes are oxygen-deprived organs can explain why Sertoli cells and germ cells may use distinctive metabolic pathways to obtain energy in their different stages of development. Therefore, studying metabolic changes during testis development can better elucidate metabolic profile of the testis, which is essential to revealing characteristic metabolic pathways. The present study applied a widely targeted UPLC-MS/MS-based metabolomics approach with large-scale detection, identification and quantification to investigate the widespread metabolic changes during Tibetan sheep testis development. Firstly, a total of 847 metabolites were detected in the sheep testis, and their changes along with the three testis-development stages were further investigated. The results indicated that those metabolites were clustered into amino acids and their derivatives, carbohydrates and their derivatives, organic acids and their derivatives, benzene and substituted derivatives, alcohols and amines, lipids, nucleotides and their derivatives, bile acids, coenzymes and vitamins, hormones and hormone-related compounds, etc. Among them, the most abundant metabolites in the testis were amino acids and lipid metabolites. The results showed that most of the lipids, carbohydrates with their derivatives, as well as alcohol and amines metabolites were high in sexually immature sheep while organic acids, amino acids and nucleotides showed a continuously increasing trend along with testis development stages. Among them, the content of metabolites with antioxidant effects increased along with testis development, while those related with energy synthesis was downregulated with age. Further correlation analyses of each metabolite-metabolite pair emphasized the cross talk between differential metabolisms across testis development, suggesting a significant correlation between lipids and other metabolites. Finally, based on KEGG pathway analysis, we found that the metabolic pathways in Tibetan sheep testis development were mainly clustered into energy metabolism, gonadal development, and anti-oxidative stress. Reactive oxygen species (ROS) are by-products of normal cellular metabolism and are inevitable during testicular energy metabolism. Thus, the anti-oxidative stress function is a key process in maintaining the normal physiological function of testis. These results contributed to a broader view of the testis metabolome and a comprehensive analysis on metabolomic variation among different testis-development stages, providing a theoretical basis for us to understand the sheep testis metabolic mechanism.

The regulatory role of BMP4 in testicular Sertoli cells of Tibetan sheep.

This study aimed to determine the regulatory mechanism of bone morphogenetic protein 4 (BMP4) gene in the testes of Tibetan sheep and its role in the blood-testis barrier (BTB). First, we cloned BMP4 gene for bioinformatics analysis, and detected the mRNA and protein expression levels of BMP4 in the testes of Tibetan sheep pre-puberty (3-mo-old), during sexual maturity (1-yr-old), and in adulthood (3-yr-old) by qRT-PCR and Western blot. In addition, the subcellular localization of BMP4 was analyzed by immunohistochemical staining. Next, BMP4 overexpression and silencing vectors were constructed and transfected into primary Sertoli cells (SCs) to promote and inhibit the proliferation of BMP4, respectively. Then, CCK-8 was used to detect the proliferation effect of SCs. The expression of BMP4 and downstream genes, pathway receptors, tight junction-related proteins, and cell proliferation and apoptosis-related genes in SCs were studied using qRT-PCR and Western blot. The results revealed that the relative expression of BMP4 mRNA and protein in testicular tissues of 1Y group and 3Y group was dramatically higher than that of 3M group (P < 0.01), and BMP4 protein is mainly located in SCs and Leydig cells at different development stages. The CDS region of the Tibetan sheep BMP4 gene was 1,229 bp. CCK-8 results demonstrated that the proliferation rate of BMP4 was significantly increased in the overexpression group (pc-DNA-3.1(+)-BMP4; P < 0.05). In addition, the mRNA and protein expressions of SMAD5, BMPR1A, and BMPR1B and tight junction-related proteins Claudin11, Occludin, and ZO1 were significantly increased (P < 0.05). The mRNA expression of cell proliferation-related gene Bcl2 was significantly enhanced (P < 0.05), and the expression of GDNF was enhanced (P > 0.05). The mRNA expression of apoptosis-related genes Caspase3 and Bax decreased significantly (P < 0.05), while the mRNA expression of cell cycle-related genes CyclinA2 and CDK2 increased significantly (P < 0.05). It is worth noting that the opposite results were observed after transfection with si-BMP4. In summary, what should be clear from the results reported here is that BMP4 affects testicular development by regulating the Sertoli cells and BTB, thereby modulating the spermatogenesis of Tibetan sheep.

The fertility of male Tibetan sheep is mainly affected by testicular development and spermatogenesis. In these processes, Sertoli cells (SCs) play a central role and are regulated by a variety of genes and factors. BMP4 is mainly distributed in Sertoli cells, and its expression level increases with age. Overexpression of the BMP4 gene in Tibetan sheep testis SCs revealed elevated expression of BMP4 protein and its downstream genes SMAD5, pathway receptor proteins BMPR1A and BMPR1B; followed by elevated expression levels of cell proliferation-related genes and decreased expression levels of apoptosis-related genes. Meanwhile, the expression of tight junction proteins was also elevated. These results indicate that BMP4 affects testicular development by regulating the Sertoli cells and blood­testis barrier, thereby affecting the spermatogenesis of Tibetan sheep.

Finite element analysis of conductive hearing loss caused by fixation and detachment of ligament and tendon in the middle ear.

BACKGROUND AND OBJECTIVE: The fixation of ligament and tendon of the middle ear often occurs after chronic otitis media surgery. However, there are relatively few studies on the effect of ligament and tendon on sound transmission in the human middle ear. Here, the finite element model and lumped parameter model are used to study the effect of ligament and tendon fixation and detachment on sound transmission in human ear. METHODS: In this paper, the finite element model including the external auditory canal, middle ear and simplified inner ear is used to calculate and compare the middle ear frequency response of the normal and tympanosclerosis under pure tone stimulation. In addition, the lumped parametric model is taken into account to illustrate the effect of ligament and tendon stiffness on the human ear transmission system. RESULTS: The results indicate that the motion of the tympanic membrane and stapes is reduced by ligament and tendon fixation. Although ligament and tendon detachment have a limited effect in the piston-motion direction, the stability of motion in the plane perpendicular to the piston-motion direction is significantly reduced. Most significantly, the ligament and tendon fixation cause a hearing effect of about 18 dB, which is greater in the plane perpendicular to the piston-motion direction after ligament and tendon detachment than in the piston-motion direction. CONCLUSIONS: In this study, the calculation accuracy of the lumped parameter and the finite element model is studied, and the effect of ligament and tendon on hearing loss is further explored through the finite element model with high calculation accuracy, which is helpful to understand the role of ligament and tendon in the sound transmission mechanism of the human middle ear. The study of ligament and tendon on conductive hearing loss provides a reference for clinical treatment of tympanosclerosis.

Identification and Functional Assignment of Genes Implicated in Sperm Maturation of Tibetan Sheep.

While traveling through the epididymis, immature sheep spermatozoa undergo a sequence of processes that ultimately give them the capacity to swim and fertilize an egg. Different gene expression patterns may be found in the epididymal caput, corpus, and cauda, conferring variant or unique biological roles during epididymis development and sperm maturation. To search for candidate genes associated with ovine sperm maturation and assess their possible modulating mechanisms, we characterized gene expression in each epididymal segment derived from pre- and post-pubertal Tibetan sheep by RNA sequencing. Compared with pre-puberty, 7730 (3724 upregulated and 4006 downregulated), 7516 (3909 upregulated and 3607 downregulated), and 7586 (4115 elevated and 3471 downregulated) genes were found to be differentially expressed in the post-pubertal caput, corpus, and cauda epididymis, respectively, and real-time quantitative PCR verified the validity of the gathered expression patterns. Based on their functional annotations, most differential genes were assigned to the biological processes and pathways associated with cellular proliferation, differentiation, immune response, or metabolic activities. As for the post-pubertal epididymis, 2801, 197, and 186 genes were specifically expressed in the caput, corpus, and cauda, respectively. Functional annotation revealed that they were mainly enriched to various distinct biological processes associated with reproduction (including the caput binding of sperm to the zona pellucida; fertilization in the caput and corpus; and meiosis in the caput and cauda) and development (such as cell differentiation and developmental maturation in the caput; cell proliferation and metabolism in the corpus; and regulation of tube size and cell division/cell cycle in the cauda). Additionally, we focused on the identification of genes implicated in immunity and sperm maturation, and subsequent functional enrichment analysis revealed that immune-related genes mainly participated in the biological processes or pathways associated with the immune barrier (such as JAM3 and ITGA4/6/9) and immunosuppression (such as TGFB2, TGFBR1, TGFBR2, and SMAD3), thus protecting auto-immunogenic spermatozoa. Additionally, sperm maturation was mostly controlled by genes linked with cellular processes, including cell growth, proliferation, division, migration, morphogenesis, and junction. Altogether, these results suggest that most genes were differentially expressed in developmental epididymal regions to contribute to microenvironment development and sperm maturation. These findings help us better understand the epididymal biology, including sperm maturation pathways and functional differences between the epididymal regions in Tibetan sheep and other sheep breeds.

A Grey BP Neural Network-Based Model for Prediction of Court Decision Service Rate.

The judgment service rate is an important index to reflect the fairness of the judgment of legal cases in a certain area, which is of great significance to verify the accuracy of a court judgment. In this paper, a grey neural network model combining grey system theory and BP neural network algorithm is proposed to predict the index. Analyze the judgment service rate of the court judgment system, and build a prediction system based on the completion rate, completion rate, plaintiff satisfaction, defendant satisfaction, litigation time, property preservation cycle, document delivery time, implementation information disclosure rate, and other key indicators. Through example analysis, it is proved that the combined model of the grey prediction model and BP neural network has a small error and good simulation effect on the prediction of court decision-making service rate, which can better promote the development of court and society.

Yeast Products Mediated Ruminal Subenvironmental Microbiota, and Abnormal Metabolites and Digestive Enzymes Regulated Rumen Fermentation Function in Sheep.

Yeast products (YP) are commonly used as rumen regulators, but their mechanisms of action are still unclear. Based on our previous studies, we questioned whether yeast products would have an impact on rumen solid-associated (SA) and liquid-associated (LA) microorganisms and alter rumen fermentation patterns. Thirty 3-month-old male sheep weighing 19.27 ± 0.45 kg were selected and randomized into three groups for 60 days: (1) basal diet group (CON group), (2) basal diet add 20 g YP per day (low YP, LYP group) and (3) basal diet add 40 g YP per day (high YP, HYP group). The results demonstrated that the addition of YP increased rumen cellulase activity, butyrate and total volatile fatty acid (TVFA) concentrations (p < 0.05), while it decreased rumen amylase activity and abnormal metabolites, such as lactate, lipopolysaccharides (LPS) and histamine (HIS) (p < 0.05). Metagenomic analysis of rumen microorganisms in three groups revealed that YP mainly influenced the microbial profiles of the SA system. YP increased the relative abundance of R. flavefaciens and decreased methanogens in the SA system (p < 0.05). With the addition of YP, the abundance of only a few lactate-producing bacteria increased in the SA system, including Streptococcus and Lactobacillus (p < 0.05). However, almost all lactate-utilizing bacteria increased in the LA system, including Megasphaera, Selenomonas, Fusobacterium and Veillonella (p < 0.05). In addition, YP increased the abundance of certain GHs family members, including GH43 and GH98 (p < 0.05), but decreased the abundance of some KEGG metabolic pathways involved in starch and sucrose metabolism, biosynthesis of antibiotics and purine metabolism, among others. In conclusion, the addition of YP to high-concentrate diets can change the abundance of major functional microbiota in the rumen, especially in the solid fraction, which in turn affects rumen fermentation patterns and improves rumen digestibility.

miR-1285-3p targets TPI1 to regulate the glycolysis metabolism signaling pathway of Tibetan sheep Sertoli cells.

Glycolysis in Sertoli cells (SCs) can provide energy substrates for the development of spermatogenic cells. Triose phosphate isomerase 1 (TPI1) is one of the key catalytic enzymes involved in glycolysis. However, the biological function of TPI1 in SCs and its role in glycolytic metabolic pathways are poorly understood. On the basis of a previous research, we isolated primary SCs from Tibetan sheep, and overexpressed TPI1 gene to determine its effect on the proliferation, glycolysis, and apoptosis of SCs. Secondly, we investigated the relationship between TPI1 and miR-1285-3p, and whether miR-1285-3p regulates the proliferation and apoptosis of SCs, and participates in glycolysis by targeting TPI1. Results showed that overexpression of TPI1 increased the proliferation rate and decreased apoptosis of SCs. In addition, overexpression of TPI1 altered glycolysis and metabolism signaling pathways and significantly increased amount of the final product lactic acid. Further analysis showed that miR-1285-3p inhibited TPI1 by directly targeting its 3'untranslated region. Overexpression of miR-1285-3p suppressed the proliferation of SCs, and this effect was partially reversed by restoration of TPI1 expression. In summary, this study shows that the miR-1285-3p/TPI1 axis regulates glycolysis in SCs. These findings add to our understanding on the regulation of spermatogenesis in sheep and other mammals.

Effects of Pgam1-mediated glycolysis pathway in Sertoli cells on Spermatogonial stem cells based on transcriptomics and energy metabolomics.

Spermatogenesis is a complex process involving a variety of intercellular interactions and precise regulation of gene expression. Spermatogenesis is sustained by a foundational Spermatogonial stem cells (SSCs) and in mammalian testis. Sertoli cells (SCs) are the major component of SSC niche. Sertoli cells provide structural support and supply energy substrate for developing germ cells. Phosphoglycerate mutase 1 (Pgam1) is a key enzyme in the glycolytic metabolism and our previous work showed that Pgam1 is expressed in SCs. In the present study, hypothesized that Pgam1-depedent glycolysis in SCs plays a functional role in regulating SSCs fate decisions. A co-culture system of murine SCs and primary spermatogonia was constructed to investigate the effects of Pgam1 knockdown or overexpression on SSCs proliferation and differentiation. Transcriptome results indicated that overexpression and knockdown of Pgam1 in SCs resulted in up-regulation of 458 genes (117 down-regulated, 341 up-regulated) and down-regulation of 409 genes (110 down-regulated, 299 up-regulated), respectively. Further analysis of these DEGs revealed that GDNF, FGF2 and other genes that serve key roles in SSCs niche maintenance were regulated by Pgam1. The metabolome results showed that a total of 11 and 16 differential metabolites were identified in the Pgam1 gene overexpression and knockdown respectively. Further screening of these metabolites indicated that Sertoli cell derived glutamate, glutamine, threonine, leucine, alanine, lysine, serine, succinate, fumarate, phosphoenolpyruvate, ATP, ADP, and AMP have potential roles in regulating SSCs proliferation and differentiation. In summary, this study established a SCs-SSCs co-culture system and identified a list of genes and small metabolic molecules that affect the proliferation and differentiation of SSCs. This study provides additional insights into the regulatory mechanisms underlying interactions between SCs and SSCs during mammalian spermatogenesis.

Metagenomic Analysis Revealed Differences in Composition and Function Between Liquid-Associated and Solid-Associated Microorganisms of Sheep Rumen.

The rumen microbiota plays a key role in the utilization of plant materials by ruminants, yet little is known about the key taxa and their genetic functions of the rumen sub-environment involved in the ruminal degradation process. Understanding the differences in the composition and function of ruminal microbiota in the liquid-associated (LA) and solid-associated (SA) systems is needed to further study and regulate rumen function and health. In this study, rumen contents of nine sheep were collected to separate LA and SA systems with elution and centrifugal precipitation. Metagenome sequencing was used to investigate the differences in microbial composition and genetic functions of LA and SA systems, with special emphasis on their degradational potential toward carbohydrates. Results showed that the dominant species composition was similar between the two systems, but SA microorganisms had a higher relative abundance than LA microorganisms in all taxa. The concentration of fiber-degrading bacteria, such as Ruminococcus, Treponema, and Fibrobacter, was higher and Prevotella was lower in the SA vs. LA system. Additionally, SA microorganisms dominated in cellulose degradation, while LA microorganisms were more important in starch utilization based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) Orthology (KO)'s functional categories and Carbohydrate-Active Enzymes (CAZymes). In general, SA microorganisms are more abundant and important in metabolic functions than LA, such as carbohydrate and amino acid metabolisms. In summary, the key differential biomarkers between LA and SA systems were Prevotella, Ruminococcus, Treponema, and Fibrobacter. Ruminal microbes degraded carbohydrates synergistically with SA, thus, more focusing on cellulose and hemicellulose, while LA is more important to starch.