NCAPG Regulates Myogenesis in Sheep, and SNPs Located in Its Putative Promoter Region Are Associated with Growth and Development Traits.

Previously, NCAPG was identified as a candidate gene associated with sheep growth traits. This study aimed to investigate the direct role of NCAPG in regulating myogenesis in embryonic myoblast cells and to investigate the association between single-nucleotide polymorphisms (SNPs) in its promoter region and sheep growth traits. The function of NCAPG in myoblast proliferation and differentiation was detected after small interfering RNAs (siRNAs) knocked down the expression of NCAPG. Cell proliferation was detected using CCK-8 assay, EdU proliferation assay, and flow cytometry cell cycle analysis. Cell differentiation was detected via cell immunofluorescence and the quantification of myogenic regulatory factors (MRFs). SNPs in the promoter region were detected using Sanger sequencing and genotyped using the improved multiplex ligation detection reaction (iMLDR®) technique. As a result, a notable decrease (p < 0.01) in the percentage of EdU-positive cells in the siRNA-694-treated group was observed. A significant decrease (p < 0.01) in cell viability after treatment with siRNA-694 for 48 h and 72 h was detected using the CCK-8 method. The quantity of S-phase cells in the siRNA-694 treatment group was significantly decreased (p < 0.01). After interfering with NCAPG in myoblasts during induced differentiation, the relative expression levels of MRFs were markedly (p < 0.05 or p < 0.01) reduced compared with the control group on days 5-7. The myoblast differentiation in the siRNA-694 treatment group was obviously suppressed compared with the control group. SNP1, SNP2, SNP3, and SNP4 were significantly (p < 0.05) associated with all traits except body weight measured at birth and one month of age. SNP5 was significantly (p < 0.05) associated with body weight, body height, and body length in six-month-old sheep. In conclusion, interfering with NCAPG can inhibit the proliferation and differentiation of ovine embryonic myoblasts. SNPs in its promoter region can serve as potential useful markers for selecting sheep growth traits.

Association between DNA Methylation in the Core Promoter Region of the CUT-like Homeobox 1 (CUX1) Gene and Lambskin Pattern in Hu Sheep.

CUT-like homeobox 1 (CUX1) has been proven to be a key regulator in sheep hair follicle development. In our previous study, CUX1 was identified as a differential expressed gene between Hu sheep lambskin with small wave patterns (SM) and straight wool patterns (ST); however, the exact molecular mechanism of CUX1 expression has been obscure. As DNA methylation can regulate the gene expression, the potential association between CUX1 core promotor region methylation and lambskin pattern in Hu sheep was explored in the present study. The results show that the core promoter region of CUX1 was present at (-1601-(-1) bp) upstream of the transcription start site. A repressive region (-1151-(-751) bp) was also detected, which had a strong inhibitory effect on CUX1 promoter activity. Bisulfite amplicon sequencing revealed that no significant difference was detected between the methylation levels of CUX1 core promoter region in SM tissues and ST tissues. Although the data demonstrated the differential expression of CUX1 between SM and ST probably has no association with DNA methylation, the identification of the core region and a potential repressive region of CUX1 promoter can enrich the role of CUX1 in Hu sheep hair follicle development.

Reproductive performance and productivity of local and Dorper x local crossbred ewes under community-based management system, Ethiopia.

This study evaluated the reproductive and productivity of local and Dorper crossbred ewes in a community-based management system. We analyzed data collected from 2013 to 2021, taking into account different factors such as dam-breed, location, type of birth, season, and year of lambing. Lambing was observed all year-round, but the majority (35%) occurred in September, October, and December. This suggests that pasture availability, which is influenced by climatic-factors, may play a role in the seasonality of lambing. Litter-size at birth and weaning did not show any significant difference. Age at first lambing varied between breeds (P < 0.001), while lambing-interval and annual reproductive rates were unaffected by dam-breed (P > 0.05). The annual number of lambing per year significantly varied based on location and lambing seasons (P < 0.001), with a higher frequency during the major rainy-season compared to the dry-season (1.58vs1.42), highlighting the influence of feed availability. Productivity indices of ewes were calculated. Location and season of lambing had a significant impact on annual ewe productivity, while the ewe genotype showed no significant influence on productivity indices, except for the weight of lambs produced per kilogram of metabolic weight (0.84vs0.72 lambs per kg ewe and year; P < 0.01: 2.02vs1.77 kg lamb per kg0.75 ewe and year), where local ewes outperformed Dorper crossbred ewes. The difference in annual-productivity indices between local and Dorper crossbred ewes was more evident when considering the postpartum weight, as the ewes exhibited higher postpartum weights. However, both ewe genotypes produced comparable lamb weights per year (20.91vs20.16 kg lamb weaned per ewe and year for local and Dorper crossbred ewes, respectively). In summary, under low-input conditions, Dorper crossbred ewes demonstrated comparable reproductive performances and productivity traits to local ewes. Nevertheless, breed and environmental factors identified in this study should be taken into account to enhance sheep productivity in both local and Dorper crossbred ewes.

Using the community-based breeding program (CBBP) model as a collaborative platform to develop the African Goat Improvement Network-Image collection protocol (AGIN-ICP) with mobile technology for data collection and management of livestock phenotypes.

Introduction: The African Goat Improvement Network Image Collection Protocol (AGIN-ICP) is an accessible, easy to use, low-cost procedure to collect phenotypic data via digital images. The AGIN-ICP collects images to extract several phenotype measures including health status indicators (anemia status, age, and weight), body measurements, shapes, and coat color and pattern, from digital images taken with standard digital cameras or mobile devices. This strategy is to quickly survey, record, assess, analyze, and store these data for use in a wide variety of production and sampling conditions. Methods: The work was accomplished as part of the multinational African Goat Improvement Network (AGIN) collaborative and is presented here as a case study in the AGIN collaboration model and working directly with community-based breeding programs (CBBP). It was iteratively developed and tested over 3 years, in 12 countries with over 12,000 images taken. Results and discussion: The AGIN-ICP development is described, and field implementation and the quality of the resulting images for use in image analysis and phenotypic data extraction are iteratively assessed. Digital body measures were validated using the PreciseEdge Image Segmentation Algorithm (PE-ISA) and software showing strong manual to digital body measure Pearson correlation coefficients of height, length, and girth measures (0.931, 0.943, 0.893) respectively. It is critical to note that while none of the very detailed tasks in the AGIN-ICP described here is difficult, every single one of them is even easier to accidentally omit, and the impact of such a mistake could render a sample image, a sampling day's images, or even an entire sampling trip's images difficult or unusable for extracting digital phenotypes. Coupled with tissue sampling and genomic testing, it may be useful in the effort to identify and conserve important animal genetic resources and in CBBP genetic improvement programs by providing reliably measured phenotypes with modest cost. Potential users include farmers, animal husbandry officials, veterinarians, regional government or other public health officials, researchers, and others. Based on these results, a final AGIN-ICP is presented, optimizing the costs, ease, and speed of field implementation of the collection method without compromising the quality of the image data collection.

The African Goat Improvement Network: a scientific group empowering smallholder farmers.

The African Goat Improvement Network (AGIN) is a collaborative group of scientists focused on genetic improvement of goats in small holder communities across the African continent. The group emerged from a series of workshops focused on enhancing goat productivity and sustainability. Discussions began in 2011 at the inaugural workshop held in Nairobi, Kenya. The goals of this diverse group were to: improve indigenous goat production in Africa; characterize existing goat populations and to facilitate germplasm preservation where appropriate; and to genomic approaches to better understand adaptation. The long-term goal was to develop cost-effective strategies to apply genomics to improve productivity of small holder farmers without sacrificing adaptation. Genome-wide information on genetic variation enabled genetic diversity studies, facilitated improved germplasm preservation decisions, and provided information necessary to initiate large scale genetic improvement programs. These improvements were partially implemented through a series of community-based breeding programs that engaged and empowered local small farmers, especially women, to promote sustainability of the production system. As with many international collaborative efforts, the AGIN work serves as a platform for human capacity development. This paper chronicles the evolution of the collaborative approach leading to the current AGIN organization and describes how it builds capacity for sustained research and development long after the initial program funds are gone. It is unique in its effectiveness for simultaneous, multi-level capacity building for researchers, students, farmers and communities, and local and regional government officials. The positive impact of AGIN capacity building has been felt by participants from developing, as well as developed country partners.

CRABP2 Promotes the Proliferation of Dermal Papilla Cells via the Wnt/ß-Catenin Pathway.

In our previous study of Hu sheep hair follicles, we found that CRABP2 was highly expressed in DPCs, which suggested that CRABP2 may influence the number of DPCs. In the present study, we aimed to understand the effect of CRABP2 in Hu sheep dermal papilla cells (DPCs). First, we explored the influence of CRABP2 on the ability of Hu sheep DPCs' proliferation. Based on the results obtained from some experiments, such as CCK-8, EDU, qPCR, and Western blot experiment, we found that the overexpression of CRABP2 facilitated the proliferation of DPCs compared to the negative control group. Then, we also detected the effect of CRABP2 on the Wnt/ß-catenin pathway based on the important function of the Wnt/ß-catenin pathway in hair follicles. The results showed that CRABP2 could activate the Wnt/ß-catenin pathway in DPCs, and it rescues the proliferation of DPCs when the Wnt/ß-catenin pathway was inhibited. In summary, our findings indicate that CRABP2 is a vital functional gene in the proliferation of Hu sheep DPCs. Our study will be of great use for revealing the roles of CRABP2 in the hair follicles of Hu sheep.

Genetic diversity and within-breed variation in three indigenous Ethiopian sheep based on whole-genome analysis.

The objective of this work was to study genetic diversity by comparing whole genome sequence data of Rutana, Gumuz and Washera sheep found in Amhara and Benishanguel gumuz regional states of Ethiopia. We employed variant calling format tools version 0.1.15 to calculate some genetic diversity indices such as observed heterozygosity, expected heterozygosity, inbreeding coefficient, and nucleotide diversity. The results revealed that, observed heterozygosity ranged from 0.33 in Gumuz to 0.34 in Rutana and Washera sheep. Expected heterozygosity ranged from 0.37 in Rutana to 0.38 in Gumuz and Washera sheep. Expected heterozygosity was found to be higher than observed heterozygosity. Higher inbreeding coefficient (0.12) was recorded for Gumuz sheep compared to 0.09 of Rutana and Washera sheep. Mean nucleotide diversity values were 0.0029, 0.0030 and 0.0028 for Gumuz, Rutana and Washera sheep, respectively. Higher values of nucleotide diversity were recorded. Population structure analysis using principal component analysis revealed no clear separation between Gumuz, Rutana and Washera sheep populations with possibility of gene flow attributed to geographical location proximity. The smaller population size, closed breeding system, genetic drift and uncontrolled (non-random) mating might lead to higher rate of inbreeding in Gumuz, Rutana and Washera sheep, requiring timely intervention. This intervention helps to prevent inbreeding depression and extinction of these valuable breeds of sheep, which helps in sustaining the livelihood of sheep keepers in lowlands and highlands. Nevertheless, the whole-genome analysis revealed high within-breed variation. Uncovered areas of studies like mapping quantitative trait loci, identifying genes underpinning productivity traits such as carcass quantity and meat quality could be carried out on diversified sheep resources identified by the current study. Identifying the genomic regions and biological pathways that contribute to explaining variability in these traits is of great importance for selection purposes. Designing conservation-based within-breed sheep selective breeding programs are recommended considering economically important traits into account.

Going to scale-From community-based to population-wide genetic improvement and commercialized sheep meat supply in Ethiopia.

Community-based breeding programs (CBBPs) have shown, at pilot scale, to be effective and beneficial in achieving genetic progress and in improving livelihoods of smallholder communities. In Ethiopia 134 sheep and goat CBBPs were operational producing their own improved rams and bucks. Based on experience the implementation of further programs is possible with appropriate private and public support. A different challenge is the efficient dissemination of the improved genetics produced in current CBBPs to create population-wide economic impact. We present a framework applied to the Ethiopian Washera sheep breed to meet this challenge. We propose the establishment of a genetic improvement structure that supports a meat commercialization model based on the integration of community-based breeding program cooperatives, client communities and complementary services such as fattening enterprises. We calculated that the recently established 28 community-based breeding programs in the Washera breeding tract can provide genetically improved rams to 22% of the four million head. To reach the whole population 152 additional CBBPs are needed. We simulated the genetic improvements obtainable in the current 28 CBBPs assuming realized genetic progress in CBBPs of a similar breed and calculated the expected additional lamb carcass meat production after 10 years of selection to be 7 tons and the accumulated discounted benefit 327 thousand USD. These benefits could be increased if the CBBPs are linked to client communities by providing them with improved rams: additional meat production would be 138 tons with a value of 3,088 thousand USD. The total meat production of the existing Washera CBBPs was calculated at 152 tons and the joint meat production of CBBPs if integrated with client communities would be 3,495 tons. A full integration model, which includes enterprises purchasing lambs for fattening, can produce up to 4,255 tons of meat. We conclude that Washera CBBPs cooperatives can benefit from a higher level of organization to produce population-wide genetic improvement and economic benefits. Unlike in the dairy and chicken industries, for low input sheep and goat smallholder systems the proposed commercialization model puts breeder cooperatives at the center of the operation. Cooperatives need to be capacitated and supported to become fully functional business ventures.

Marker-assisted introgression of fecundity mutation into Barbarine sheep breed under low input production system.

Fecundity gene introgression has become an economical method to improve sheep prolificacy in developing countries. The FecXBar and FecGH mutations are variants of the BMP15 and GDF9 sheep genes, respectively, identified in Tunisian Barbarine sheep prolific line "W-INRAT," created through a prolificacy-based selection program. The first mutation caused increased prolificacy in heterozygous state and sterility in homozygous ewes. The aim of this work was to increase the number of effective carriers by the introgression of fecundity mutation into non-carrier conventional flocks based on a marker-assisted breeding program. The genotyping was carried out to follow up on the segregation of prolificacy mutation. The conventional ewes, raised in state farms of Tunisia and inseminated by "W-INRAT" rams, yielded 100% carriers females at heterozygous state. These females were selected to be inseminated with conventional rams and produced carriers' lambs with 66.7% males and 33.3% females. The prolific males will be kept in breeding centers to disseminate the fecundity mutation in commercial flocks. This approach facilitates the dissemination of fecundity genes and contributes to livelihood improvement in communities raising Barbarine sheep.

How to succeed in implementing community-based breeding programs: Lessons from the field in Eastern and Southern Africa.

Breeding programs involving either centralized nucleus schemes and/or importation of exotic germplasm for crossbreeding were not successful and sustainable in most Africa countries. Community-based breeding programs (CBBPs) are now suggested as alternatives that aim to improve local breeds and concurrently conserve them. Community-based breeding program is unique in that it involves the different actors from the initial phase of design up until implementation of the programs, gives farmers the knowledge, skills and support they need to continue making improvements long into the future and is suitable for low input systems. In Ethiopia, we piloted CBBPs in sheep and goats, and the results show that they are technically feasible to implement, generate genetic gains in breeding goal traits and result in socio-economic impact. In Malawi, CBBPs were piloted in local goats, and results showed substantial gain in production traits of growth and carcass yields. CBBPs are currently being integrated into goat pass-on programs in few NGOs and is out-scaled to local pig production. Impressive results have also been generated from pilot CBBPs in Tanzania. From experiential monitoring and learning, their success depends on the following: 1) identification of the right beneficiaries; 2) clear framework for dissemination of improved genetics and an up/out scaling strategy; 3) institutional arrangements including establishment of breeders' cooperatives to support functionality and sustainability; 4) capacity development of the different actors on animal husbandry, breeding practices, breeding value estimation and sound financial management; 5) easy to use mobile applications for data collection and management; 6) long-term technical support mainly in data management, analysis and feedback of estimated breeding values from committed and accessible technical staff; 7) complementary services including disease prevention and control, proper feeding, and market linkages for improved genotypes and non-selected counterparts; 8) a system for certification of breeding rams/bucks to ensure quality control; 9) periodic program evaluation and impact assessment; and 10) flexibility in the implementation of the programs. Lessons relating to technical, institutional, community dynamics and the innovative approaches followed are discussed.

Integration of the Microbiome, Metabolome and Transcriptome Reveals Escherichia coli F17 Susceptibility of Sheep.

Escherichia coli (E. coli) F17 is one of the most common pathogens causing diarrhea in farm livestock. In the previous study, we accessed the transcriptomic and microbiomic profile of E. coli F17-antagonism (AN) and -sensitive (SE) lambs; however, the biological mechanism underlying E. coli F17 infection has not been fully elucidated. Therefore, the present study first analyzed the metabolite data obtained with UHPLC-MS/MS. A total of 1957 metabolites were profiled in the present study, and 11 differential metabolites were identified between E. coli F17 AN and SE lambs (i.e., FAHFAs and propionylcarnitine). Functional enrichment analyses showed that most of the identified metabolites were related to the lipid metabolism. Then, we presented a machine-learning approach (Random Forest) to integrate the microbiome, metabolome and transcriptome data, which identified subsets of potential biomarkers for E. coli F17 infection (i.e., GlcADG 18:0-18:2, ethylmalonic acid and FBLIM1); furthermore, the PCCs were calculated and the interaction network was constructed to gain insight into the crosstalk between the genes, metabolites and bacteria in E. coli F17 AN/SE lambs. By combing classic statistical approaches and a machine-learning approach, our results revealed subsets of metabolites, genes and bacteria that could be potentially developed as candidate biomarkers for E. coli F17 infection in lambs.

Effect of CUX1 on the Proliferation of Hu Sheep Dermal Papilla Cells and on the Wnt/ß-Catenin Signaling Pathway.

CUT-like homeobox 1 protein (CUX1), also called CUX, CUTL1, and CDP, is a member of the DNA-binding protein homology family. Studies have shown that CUX1 is a transcription factor that plays an important role in the growth and development of hair follicles. The aim of this study was to investigate the effect of CUX1 on the proliferation of Hu sheep dermal papilla cells (DPCs) to reveal the role of CUX1 in hair follicle growth and development. First, the coding sequence (CDS) of CUX1 was amplified by PCR, and then CUX1 was overexpressed and knocked down in DPCs. A Cell Counting Kit-8 (CCK8), 5-ethynyl-2-deoxyuridine (EdU), and cell cycle assays were used to detect the changes in the proliferation and cell cycle of DPCs. Finally, the effects of overexpression and knockdown of CUX1 in DPCs on the expression of WNT10, MMP7, C-JUN, and other key genes in the Wnt/ß-catenin signaling pathway were detected by RT-qPCR. The results showed that the 2034-bp CDS of CUX1 was successfully amplified. Overexpression of CUX1 enhanced the proliferative state of DPCs, significantly increased the number of S-phase cells, and decreased the number of G0/G1-phase cells (p < 0.05). CUX1 knockdown had the opposite effects. It was found that the expression of MMP7, CCND1 (both p < 0.05), PPARD, and FOSL1 (both p < 0.01) increased significantly after overexpression of CUX1 in DPCs, while the expression of CTNNB1 (p < 0.05), C-JUN, PPARD, CCND1, and FOSL1 (all p < 0.01) decreased significantly. In conclusion, CUX1 promotes proliferation of DPCs and affects the expression of key genes of the Wnt/ß-catenin signaling pathway. The present study provides a theoretical basis to elucidate the mechanism underlying hair follicle development and lambskin curl pattern formation in Hu sheep.

The identification and validation of target genes of IGFBP3 protein in sheep skeletal muscle cells.

This study aimed to identify the target genes of IGFBP3（insulin growth factor binding protein）protein and to investigate its target genes effects on the proliferation and differentiation of Hu sheep skeletal muscle cells. IGFBP3 was an RNA-binding protein that regulates mRNA stability. Previous studies have reported that IGFBP3 promotes the proliferation of Hu sheep skeletal muscle cells and inhibits differentiation, but the downstream genes that bind to it have not been reported yet. We predicted the target genes of IGFBP3 through RNAct and sequencing data, and verified by qPCR and RIP（RNA Immunoprecipitation）experiments, and demonstrated GNAI2（G protein subunit alpha i2）as one of the target gene of IGFBP3. After interference with siRNA, we carried out qPCR, CCK8, EdU, and immunofluorescence experiments, and found that GNAI2 can promote the proliferation and inhibit differentiation of Hu sheep skeletal muscle cells. This study revealed the effects of GNAI2 and provided one of the regulatory mechanisms of IGFBP3 protein underlying sheep muscle development.

Expression analysis of TLR signaling pathway genes under lipopolysaccharide-induced and E. coli F17-infected sheep intestinal epithelial cells.

Escherichia coli (E. coli) F17 is one of the main pathogens causing diarrhea in young livestock. The specific F17 fimbriae and lipopolysaccharide (LPS) in the surface components of E. coli F17 induces immune activation via interacting with the intestinal epithelial cells (IECs)-expressed innate immune toll-like receptors (TLRs) signaling pathway. In this study, the expression patterns of eight canonical genes from the TLR signaling pathway (IL-6, IL-8, IL-1ß, TLR4, MyD88, CD14, TNF-α and TRAF6) were analyzed in LPS-induced IECs, E. coli F17-infected IECs and ileum tissue of E. coli F17-infected lambs. The results showed that increased expression levels of all the studied genes were observed following post-LPS-induced and E. coli F17-infected treatment, with TLR4 having the highest up-regulated expression multiple (compared to NC, fold change = 17.94 and 20.11, respectively), and CD14 having the lowest up-regulated expression multiple (fold change = 2.68 and 1.59, respectively), and higher expression levels of all the studied TLR signaling pathway genes were observed in ileum tissue of E. coli F17 antagonistic (AN) lambs than in E. coli F17 sensitive (SE) lambs. Furthermore, when compared to LPS-induced IECs, E. coli F17-infected IECs showed a more pronounced increase in the expression of IL6, TLR4 and TNF-α, indicating the different roles of these genes in the IECs resistance to E. coli F17 infection. Our results demonstrate that the TLR signaling pathway likely promotes immune activation and provide the first evidence that TLRs have a significant potential to protect against E. coli F17 infections.

Effects of YAP1 on proliferation and differentiation of Hu sheep skeletal muscle satellite cells in vitro.

This study aimed to understand the expression level of YAP1 in the skeletal muscle of Hu sheep and to reveal the regulatory mechanism of YAP1 on Hu sheep skeletal muscle satellite cells (SMSCs). Previous research by our group has found that YAP1 may affect the growth and development of Hu sheep skeletal muscle. In the present study, we found the expression of YAP1 in the skeletal muscle is higher than in other tissues of Hu sheep. Then, we detected the effect of YAP1 on proliferation and differentiation in Hu sheep SMSCs. According to the results of qPCR, CCK-8, EDU, and Western blot, compared to the group of negative control, overexpression of YAP1 promoted the proliferation and inhibited the differentiation of SMSCs according to the results of qPCR, CCK-8, EDU, Western blot, while the interference of YAP1 was on the contrary. Overall, our study suggests that YAP1 is an important functional molecule in the growth and development of skeletal muscle by regulating the proliferation and differentiation of SMSCs. These findings are of great use for understanding the roles of YAP1 in the skeletal muscle of Hu sheep.

Whole-genome resequencing of Dorper and Hu sheep to reveal selection signatures associated with important traits.

Dorper and Hu sheep exhibit different characteristics in terms of reproduction, growth, and meat quality. Comparison of the genomes of two breeds help to reveal important genomic information. In this study, whole genome resequencing of 30 individuals (Dorper, DB and Hu sheep, HY) identified 15,108,125 single nucleotide polymorphisms (SNPs). Population differentiation (Fst) and cross population extended haplotype homozygosity (XP-EHH) were performed for selective signal analysis. In total, 106 and 515 overlapped genes were present in both the Fst results and XP-EHH results in HY vs DB and in DB vs HY, respectively. In HY vs DB, 106 genes were enriched in 12 GO terms and 83 KEGG pathways, such as ATP binding (GO:0005524) and PI3K-Akt signaling pathway (oas04151). In DB vs HY, 515 genes were enriched in 109 GO terms and 215 KEGG pathways, such as skeletal muscle cell differentiation (GO:0035914) and MAPK signaling pathway (oas04010). According to the annotation results, we identified a series of candidate genes associated with reproduction (UNC5C, BMPR1B, and GLIS1), meat quality (MECOM, MEF2C, and MYF6), and immunity (GMDS, GALK1, and ITGB4). Our investigation has uncovered genomic information for important traits in sheep and provided a basis for subsequent studies of related traits.

Litter size, litter weight, and lamb survivability of Doyogena sheep managed under community-based breeding program in Ethiopia.

Increasing litter size and weight during the marketing time by decreasing mortality among lambs per ewe is the objective of the community-based breeding program (CBBP). This study aims to find out litter size, litter weight per ewe, and preweaning lamb mortality of Doyogena sheep managed under CBBP. The study analyzed the data records of 4530 animals for 8 years from 2013 to 2020. A logistic regression procedure was used to analyze pre-weaning lamb survival rates. Results showed that the overall least-squares means of litter size at birth (LSB), litter size at weaning (LSW), total litter weight at birth (TLWB), and total litter weight at weaning (TLWW) were 1.57 ± 0.02 lambs, 1.50 ± 0.02 lambs, 5.24 ± 0.09 kg, and 24.14 ± 0.69 kg respectively. The incidence of pre-weaning lamb mortality was 4.72%. Year and breeder cooperative vary were observed for pre-weaning lamb losses. Females had lower odds of survival as compared to males. Odds of survival were low for triplets, quadruplets, and low birth weight (≤2 kg) born lambs. The ongoing selection program which aims to improve reproductive and growth performance had a positive influence on the survival rate of lambs. Improvement of the environment in the flock, special care for multiple-born and care for small lambs would lead to further lamb survival improvements.

Preliminary study of melatonin in the proliferation and apoptosis of Hu sheep dermal papilla cells in vitro.

Previous studies have shown that melatonin has a certain regulatory effect on the growth of sheep wool. However, the mechanism of melatonin action remains unknown. In the present study, we aimed to understand the role of exogenous melatonin in the dermal papilla cells of Hu sheep. To confirm the optimal melatonin treatment regimen for Hu sheep dermal papilla cells, we detected the cell viability by exposing them to different concentrations of melatonin and different treatment times. The results showed that cell viability was best when dermal papilla cells were treated with 1000 pg/ml of melatonin for 48 h. According to the results of qPCR, CCK-8, EDU, Western blot, and Flow cytometry analysis, we found that 1000 pg/ml melatonin promoted the proliferation and inhibited the apoptosis of dermal papilla cells compared with the exogenous melatonin blank group (control group). Furthermore, we also found that 1000 pg/ml of melatonin promoted the cell cycle progress of dermal papilla cells according to the results of qPCR and Flow cytometry analysis. Overall, our findings showed that melatonin plays an important role in the dermal papilla cells of Hu sheep.

New Insight into the Role of the Leucine Aminopeptidase 3 (LAP3) in Cell Proliferation and Myogenic Differentiation in Sheep Embryonic Myoblasts.

Previous genome-wide association studies (GWAS) have found that LAP3 may have the potential function to impact sheep muscle development. In order to further explore whether LAP3 expression has an important role in the development of sheep embryonic myoblasts, we conducted the spatiotemporal expression profile analysis of LAP3 at the tissue and cellular level. Then we used small interfering RNA and eukaryotic recombinant vectors to perform gain/loss-of-function analysis of LAP3. CCK-8 detection, EdU staining, and flow cytometry were used to investigate the impact of LAP3 knockdown or overexpression on the proliferation of embryonic myoblasts. In addition, cell phenotype observation, MyHC indirect immunofluorescence, and quantitative detection of the expression changes of myogenic regulatory factors (MRFs) were used to explore the effect of LAP3 on myogenic differentiation. The results showed that the LAP3 expression level in muscle tissue of fetuses was significantly higher than that in newborn lambs and adult sheep, and its expression level on day 3 of differentiation was also significantly higher than that in the proliferation phase and other differentiation time points. LAP3 silencing could significantly increase cell viability and EdU-positive cells, as well as prolonging the length of S phase of myoblasts to promote proliferation, while the results were reversed when LAP3 was overexpressed. Moreover, LAP3 silencing significantly hindered myotube formation and down-regulated the expression levels of MRFs from day 5 to day 7 of terminal differentiation, while the results were reversed when LAP3 was highly expressed. Overall, our results suggested that the expression of LAP3 impacts on the development of sheep embryonic myoblasts which provides an important theoretical basis for molecular breeding of meat production in sheep.

The Effect of EGR1 on the Proliferation of Dermal Papilla Cells.

Early growth response factor 1 (EGR1) is a zinc-finger transcription factor that plays a vital role in the development of hair follicles. According to our previous studies, EGR1 is a transcriptional promoter of the bone morphogenetic protein 7 (BMP7), a candidate gene involved in the proliferation of dermal papilla cells. Since hair follicles are the basis of lambskin pattern formation and dermal papilla cells (DPCs) act on hair follicle growth, in order to elucidate the role of EGR1 and hair follicles, this study aimed to investigate the biological role of EGR1 in DPCs. In our study, the EGR1 coding sequence (CDS) region was firstly cloned by polymerase chain reaction, and bioinformatics analysis was performed. Then, the function of EGR1 was detected by 5-ethynyl-2'-deoxyuridine (EDU) and Cell Counting Kit-8 (CCK8), and Western blot (WB) was conducted to analyze the cellular effect of EGR1 on DPCs. The proliferative effect of EGR1 on DPCs was also further confirmed by detecting its expression by qPCR and WB on marker genes of proliferation, including PCNA and CDK2. The sequence of the EGR1 CDS region of a lamb was successfully cloned, and its nucleic acid sequence was analyzed and found to be highly homologous to Rattus norvegicus, Mus musculus, Bos taurus and Homo sapiens. Predictive analysis of the protein encoded by EGR1 revealed that it is an extra-membrane protein, and not a secretory protein, with subcellular localization in the nucleus and cytoplasm. The proliferative effect of DPCs was significantly stronger (p < 0.01) in EGR1 up-regulated DPCs compared to the controls, while the opposite result was observed in EGR1 down-regulated DPCs. Markers of proliferation including PCNA and CDK2 also appeared to be differentially upregulated in EGR1 gene overexpression compared to the controls, with the opposite result in EGR1 gene downregulation. In summary, our study revealed that EGR1 promotes the proliferation of DPCs, and we speculate that EGR1 may be closely associated with hair follicle growth and development.