miR-138-5p promotes chicken granulosa cell apoptosis via targeting SIRT1.

Granulosa cell (GC) apoptosis is the main trigger of follicular atresia. MicroRNAs (miRNAs) are 18-22 nt RNAs whose function is primarily determined by their extended seed region and are considered to be involved in the biological functions of follicular development, including follicular atresia, folliculogenesis, and oogenesis. MiR-138-5p is known to act on chicken GCs. In this study, we found that miR-138-5p was enriched in reproductive organs, such as the uterus and ovaries. To examine whether miR-138-5p could regulate the biological process of GCs, miR-138-5p was examined by transfection of cells with a mimic or inhibitor of miR-138-5p. Expression levels of caspase-3 and caspase-9 mRNA and protein were markedly increased or decreased after transfection of the mimic or inhibitor, respectively. Furthermore, following miR-138-5p inhibition, SIRT1, one of the target genes of miR-138-5p, was found to increase the mRNA, which is correlated with the increased levels of BCL2 expression, an anti-apoptotic gene in the chicken GCs. These results suggest that miR-138-5p promotes apoptosis in chicken GCs by targeting SIRT1.

Whole transcriptome analysis reveals the key genes and noncoding RNAs related to follicular atresia in broilers.

Broodiness, a maternal behavior, is accompanied by the atresia of follicles and the serious degradation of poultry reproductive performance. The comparison of follicles between brooding and laying hens is usually an ideal model for exploring the regulation mechanism of follicle atresia. In this study, we selected three brooding hens and three laying hens to collect their follicles for whole transcriptome sequencing. The results demonstrated different expression patterns between the follicles of brooding hens and laying hens. In the top 10 differentially expressed genes with the highest expression, MMP10 was relatively low expressed in the follicles of brooding hens, but other nine genes were relatively highly expressed, including LRR1, RACK1, SPECC1L, ABHD2, COL6A3, RPS17, ATRN, BIRC6, PGAM1 and SPECC1L. While miR-21-3p, miR-146a-5p, miR-142-5p and miR-1b-3p were highly expressed in the follicles of brooding hen, miR-106-5p, miR-451, miR-183, miR-7, miR-2188-5p and miR-182-5p were lowly expressed in brooding hen. In addition, we identified 124 lncRNAs specifically expressed in the follicles of brooding hens and 147 lncRNAs specifically expressed in the follicles of laying hens. Our results may provide a theoretical basis for further exploration of the molecular mechanism of broodiness in broilers.

Whole-genome resequencing reveals aberrant autosomal SNPs affect chicken feathering rate.

Previous studies have shown that the feather growth rate of chicks is determined by two alleles located on the sex chromosome Z; however, in chicken production, feathering is usually not consistently controlled by the sex chromosome. To identify whether the feathering rate is related to autosomal inheritance, whole-genome resequencing was performed in eight chickens with slow- and fast-feathering rate. A total of 54,984 autosomal single nucleotide polymorphisms (SNPs) were identified, including 393 and 376 exonic SNPs in slow-feathering and fast-feathering chickens, respectively. Mutated genes were mainly involved in response to stimuli and growth and reproduction processes. Mutated genes related to slow-feathering rate were mainly involved in wingless-type MMTV integration site signaling pathway and mitogen-activated protein kinase signaling pathway, whereas mutated genes associated with fast-feathering rate were primarily enriched in autophagy, calcium signaling pathway, extracellular matrix-receptor interaction, and Focal adhesion processes. Importantly, two SNPs, involved in feather development, were found in the exonic regions of Wnt signaling genes. These results shed new light on the relationship between genetic mutation and feather growth rate from the perspective of autosomal inheritance and may have economic significance in chicken breeding.

Effects of Lactobacillus plantarum on growth traits, slaughter performance, serum markers and intestinal bacterial community of Daheng broilers.

Probiotics are expected to be an ideal alternative for antibiotics in the poultry industry. This study aimed to investigate the effect of Lactobacillus plantarum on growth traits, slaughter performance, serum markers and intestinal bacterial community of Daheng broilers. A total of 2400 healthy one-day-old Daheng broilers were randomly divided into 5 groups with 6 replicates per group and 40 individuals per replicate. Birds in control group were fed a basal diet, and others were fed basal diets supplemented with 105 , 106 , 107 and 108  CFU/kg Lactobacillus plantarum, respectively. It turned out that adding Lactobacillus plantarum to diet could significantly improve the serum immune performance of broilers (p < 0.05), enhance the antioxidant capacity to a certain extent (p > 0.05), but had no significant effect on growth traits and slaughter performance. Moreover, Lactobacillus plantarum could improve the diversity of intestinal bacterial community, but with the increase of addition concentration, the diversity would gradually decrease. In conclusion, Lactobacillus plantarum can be used as feed additive in broiler production, but whether it is more effective than antibiotics needs further investigation.

Small RNA sequencing of pectoral muscle tissue reveals microRNA-mediated gene modulation in chicken muscle growth.

Sichuan mountainous black-bone (SMB) chicken is a small-sized black-feathered chicken breed with low amount of meat, while Dahen (DH) chicken has a larger body size and a faster growth rate. MicroRNAs (miRNAs) are involved in various physiological processes, but their role in chicken muscle growth remains unclear. We aimed to investigate the miRNAs and pathways participating in the muscle growth of chicken. MiRNA profiles of four SMB chickens and four DH chickens were detected by small RNA sequencing. A total of 994 known miRNAs were identified, among which gga-miR-1a-3p, gga-miR-148-3p and gga-miR-133a-3p exhibited the highest enrichment in both breeds of chickens. Thirty-two miRNAs were differently expressed between SMB and DH chickens. The differently expressed miRNAs were mainly associated with fatty acid metabolism, immunity and MAPK activation-related processes. Kyoto encyclopaedia of genes and genomes (KEGG) analysis showed that miRNAs were involved in the immunity-related and MAPK signalling pathways. Moreover, miR-204 was downregulated in DH chicken compared with SMB chicken, and significantly inhibited the expression of MAP3K13, which is involved in the MAPK pathway. It was confirmed through luciferase reporter assays that miR-204 specifically inhibited the activity of MAP3K13. Our results helped demonstrate the potential molecular mechanisms of muscle growth in chickens and provide valuable information for chicken breeding.

Population genomics identifies patterns of genetic diversity and selection in chicken.

BACKGROUND: There are hundreds of phenotypically distinguishable domestic chicken breeds or lines with highly specialized traits worldwide, which provide a unique opportunity to illustrate how selection shapes patterns of genetic variation. There are many local chicken breeds in China. RESULTS: Here, we provide a population genome landscape of genetic variations in 86 domestic chickens representing 10 phenotypically diverse breeds. Genome-wide analysis indicated that sex chromosomes have less genetic diversity and are under stronger selection than autosomes during domestication and local adaptation. We found an evidence of admixture between Tibetan chickens and other domestic population. We further identified strong signatures of selection affecting genomic regions that harbor genes underlying economic traits (typically related to feathers, skin color, growth, reproduction and aggressiveness) and local adaptation (to high altitude). By comparing the genomes of the Tibetan and lowland fowls, we identified genes associated with high-altitude adaptation in Tibetan chickens were mainly involved in energy metabolism, body size maintenance and available food sources. CONCLUSIONS: The work provides crucial insights into the distinct evolutionary scenarios occurring under artificial selection for agricultural production and under natural selection for success at high altitudes in chicken. Several genes were identified as candidates for chicken economic traits and other phenotypic traits.

Comparative transcriptome analysis reveals regulators mediating breast muscle growth and development in three chicken breeds.

Objective: The goal of this study was to investigate the mechanisms of muscle growth and development of three chicken breeds. Participants: Eighteen chickens, including three different breeds with different growth speeds (White Broiler, Daheng, and Commercial Layers of Roman), were used. Methods: Total RNA from breast muscle of these chickens was subjected to a gene expression microarray. Differentially expressed genes (DEGs) were screened and functional enrichment analysis was performed using DAVID. Seven DEGs were confirmed by quantitative reverse transcription PCR. Results: Overall, 8,398 DEGs were found among the different lines. The DEGs between each two lines that were unique for a developmental stage were greater than those that were common during all stages. Functional analysis revealed that DEGs across the entire developmental process were primarily involved in positive cell proliferation, growth, cell differentiation, and developmental processes. Genes involved in muscle regulation, muscle construction, and muscle cell differentiation were upregulated in the faster-growing breed compared to the slower-growing breed. DEGs including myosin heavy chain 15 (MYH15), myozenin 2 (MYOZ2), myosin-binding protein C (MYBPC3), insulin-like growth factor 2 (IGF2), apoptosis regulator (BCL-2), AP-1 transcription factor subunit (JUN), and AP-1 transcription factor subunit (FOS) directly regulated muscle growth or were in the center of the protein-protein interaction network. Pathways, including the extracellular matrix (ECM)-receptor interaction, mitogen-activated protein kinase (MAPK) signaling pathway, and focal adhesion, were the most enriched DEGs between lines or within lines under different developmental stages. Conclusions: Genes involved in muscle construction and cell differentiation were differentially expressed among the three breeds.

A comparison of arterial spin labeling perfusion MRI and DCE-MRI in human prostate cancer.

Perfusion MRI has the potential to provide pathophysiological biomarkers for the evaluating, staging and therapy monitoring of prostate cancer. The objective of this study was to explore the feasibility of noninvasive arterial spin labeling (ASL) to detect prostate cancer in the peripheral zone and to investigate the correlation between the blood flow (BF) measured by ASL and the pharmacokinetic parameters K(trans) (forward volume transfer constant), kep (reverse reflux rate constant between extracellular space and plasma) and ve (the fractional volume of extracellular space per unit volume of tissue) measured by dynamic contrast-enhanced (DCE) MRI in patients with prostate cancer. Forty-three consecutive patients (ages ranging from 49 to 86 years, with a median age of 74 years) with pathologically confirmed prostate cancer were recruited. An ASL scan with four different inversion times (TI = 1000, 1200, 1400 and 1600 ms) and a DCE-MRI scan were performed on a clinical 3.0 T GE scanner. BF, K(trans), kep and ve maps were calculated. In order to determine whether the BF values in the cancerous area were statistically different from those in the noncancerous area, an independent t-test was performed. Spearman's bivariate correlation was used to assess the relationship between BF and the pharmacokinetic parameters K(trans), kep and ve. The mean BF values in the cancerous areas (97.1 ± 30.7, 114.7 ± 28.7, 102.3 ± 22.5, 91.2 ± 24.2 ml/100 g/min, respectively, for TI = 1000, 1200, 1400, 1600 ms) were significantly higher (p < 0.01 for all cases) than those in the noncancerous regions (35.8 ± 12.5, 42.2 ± 13.7, 53.5 ± 19.1, 48.5 ± 13.5 ml/100 g/min, respectively). Significant positive correlations (p < 0.01 for all cases) between BF and the pharmacokinetic parameters K(trans), kep and ve were also observed for all four TI values (r = 0.671, 0.407, 0.666 for TI = 1000 ms; 0.713, 0.424, 0.698 for TI = 1200 ms; 0.604, 0.402, 0.595 for TI = 1400 ms; 0.605, 0.422, 0.548 for TI = 1600 ms). It can be seen that the quantitative ASL measurements show significant differences between cancerous and benign tissues, and exhibit strong to moderate correlations with the parameters obtained using DCE-MRI. These results show the promise of ASL as a noninvasive alternative to DCE-MRI.

Characterization of the expression profile of calpain-3 (CAPN3) gene in chicken.

Calpain-3 is a skeletal muscle-specific protease and participates in the regulation of myogenesis. In this study, we quantified the expression of calpain-3 (CAPN3) mRNA in a Chinese local chicken breed (Sichuan Mountainous Black-boned chicken [MB]), to discern the tissue and ontogenic expression pattern. Meanwhile, we compared the CAPN3 mRNA expression pattern in MB chicken at 10 weeks with a commercial meat type chicken line (S01) of the same age to identify the unique expression pattern under different genetic background. A real time quantitative PCR (qRT-PCR) assay was developed for an accurate measurement of its expression in various tissues from chickens at different ages (0, 2, 4, 6, 8, 10, and 12 weeks). Expression of the CAPN3 mRNA was detected in the selected tissues, regardless of age. The breast muscle and leg muscle tissues had a significantly higher expression than the other tissues from the same individual (P < 0.01). Overall, the CAPN3 mRNA level exhibited a "rise-decline" developmental change in detected tissues except for brain. The S01 chicken had a higher expression of the CAPN3 mRNA in detected tissues than the MB chicken at 10 weeks. The present expression data of chicken CAPN3 gene may provide some information to shed light on the tissue and ontogenic expression pattern during chicken development.

Characterization of the expression profiles of calpastatin (CAST) gene in chicken.

The calpain system, a Ca(2+)-activated protease family, plays an important role in postmortem tenderization of skeletal muscle due to its involvement in the degradation of important myofibrillar and associated proteins, as well as in cytoskeletal remodeling and regulation of muscle growth. In this study, we quantified the expression of calpastatin (CAST) in two Chinese chicken breeds (mountainous black-bone chicken breed (MB) and a commercial meat type chicken breed (S01)), to discern the tissue and age-related specific expression pattern and its potential role on muscle tissue metabolism. Real-time quantitative PCR (RT-qPCR) assay was developed for accurate measurement of CAST mRNA levels in various tissues from chicken with different ages (0, 2, 4, 6, 8, 10, and 12 week). CAST mRNA was detected in collected organs. The heart and leg muscle tissues had the highest expression of CAST than other tissues from the same chicken (P < 0.01). Age-related expression pattern of CAST gene was evident in breast muscle, liver, and brain tissues (P < 0.05), but not in heart and leg muscle tissues (P > 0.05). Overall, the CAST mRNA level exhibited a "rise-decline-rise-decline" developmental change in breast muscle and liver, with the highest expression at 2 weeks and the lowest expression at 8 weeks. The S01 chicken had significantly higher expression of CAST in breast muscle and heart than the MB chicken (P < 0.05) at 10 weeks. Our results suggested the CAST expression may be related to muscle fiber development.

Integration analysis of metabolome and transcriptome profiles revealed the age-dependent dynamic change in chicken meat.

To explore the chemical composition of chicken meat during different growth and development periods, the dynamic alterations of the metabolite composition were determined using LC-MS/MS-based metabolomics. Together, 573 metabolites were identified in chicken meat from five age stages. Generally, pentadecanoic acid, stearic acid, creatine, carnosine, IMP, L-histidine and L-isoleucine presented an upward trend with age, while anserine, DHA, L-aspartic acid, LPA 18:1 and LPI 18:1 decreased with age. The main pathways of chicken meat metabolism affected by age were fructose and mannose metabolism, arachidonic acid metabolism, steroid hormone biosynthesis, riboflavin metabolism, biosynthesis of unsaturated fatty acids, and linoleic acid metabolism. Using transcriptomic profiling data, we conducted Pearson correlation analysis between gene expression and metabolite profile data in each age comparison. Integration analysis of metabolome and transcriptome would be helpful to understand the biological processes underlying the development of meat quality and explore valuable biomarkers for specific metabolite accumulation.

miRNA sequencing analysis of healthy and atretic follicles of chickens revealed that miR-30a-5p inhibits granulosa cell death via targeting Beclin1.

BACKGROUND: The egg production performance of chickens is affected by many factors, including genetics, nutrition and environmental conditions. These factors all play a role in egg production by affecting the development of follicles. MicroRNAs (miRNAs) are important non-coding RNAs that regulate biological processes by targeting genes or other non-coding RNAs after transcription. In the animal reproduction process, miRNA is known to affect the development and atresia of follicles by regulating apoptosis and autophagy of granulosa cells (GCs). RESULTS: In this study, we identified potential miRNAs in the atretic follicles of broody chickens and unatretic follicles of healthy chickens. We identified gga-miR-30a-5p in 50 differentially expressed miRNAs and found that gga-miR-30a-5p played a regulatory role in the development of chicken follicles. The function of miR-30a-5p was explored through the transfection test of miR-30a-5p inhibitor and miR-30a-5p mimics. In the study, we used qPCR, western blot and flow cytometry to detect granulosa cell apoptosis, autophagy and steroid hormone synthesis. Confocal microscopy and transmission electron microscopy are used for the observation of autophagolysosomes. The levels of estradiol (E2), progesterone (P4), malondialdehyde (MDA) and superoxide dismutase (SOD) were detected by ELISA. The results showed that miR-30a-5p showed a negative effect on autophagy and apoptosis of granulosa cells, and also contributed in steroid hormones and reactive oxygen species (ROS) production. In addition, the results obtained from the biosynthesis and dual luciferase experiments showed that Beclin1 was the target gene of miR-30a-5p. The rescue experiment conducted further confirmed that Beclin1 belongs to the miR-30a-5p regulatory pathway. CONCLUSIONS: In summary, after deep miRNA sequencing on healthy and atretic follicles, the results indicated that miR-30a-5p inhibits granulosa cell death by inhibiting Beclin1.

miR-23b-3p inhibits chicken granulosa cell proliferation and steroid hormone synthesis via targeting GDF9.

MicroRNAs (miRNAs) are ∼22 nt RNAs that direct post-transcriptional repression of mRNA targets in diverse eukaryotic lineages. Granulosa cells (GCs) are the earliest differentiated follicular somatic cells. From the initiation of primordial follicles, their differentiation and growth are closely related to the development of follicles. The research on follicular development mostly focused on the granular layer, as well as the hormone synthesis induced by granulosa cell differentiation before and after follicular selection. In this study, we evaluated the effects of miR-23b-3p on chicken granulosa cells, including granulosa cell proliferation and steroid hormone synthesis. Elevated expression of miR-23b-3p significantly inhibited granulosa cell proliferation and steroid hormone synthesis, but did not affect apoptosis. Furthermore, it was observed that the forecast growth differentiation factor 9 (GDF9) is a target gene of miR-23b-3p and miR-23b-3p can down-regulate expression of GDF9. Overall, this study demonstrated that miR-23b-3p can regulate the proliferation and steroid hormone synthesis of chicken granulosa cells by inhibiting the expression of GDF9.

Fibromodulin is involved in autophagy and apoptosis of granulosa cells affecting the follicular atresia in chicken.

Follicular atresia is an important cause of reproductive decline in egg-laying hens. Therefore, a better understanding of the regulation mechanism of follicle atresia in poultry is an important measure to maintain persistent high egg performance. However, how the role of the regulatory relationship between autophagy and apoptosis in the intrafollicular environment affects the follicular atresia of chickens is remain unclear. The objective of this study was to explore the regulatory molecular mechanisms in regard to follicular atresia. 20 white leghorn layers (32-wk-old) were equally divided into 2 groups. The control group was fed freely, and the experimental group induced follicular atretic by fasting for 5 d. The results showed that the expression of prolactin (PRL) levels was significantly higher in the fasted hens, while the levels of luteinizing hormone (LH) and follicle stimulating hormone (FSH) were lower. Most importantly, RNA sequencing, qPCR, and Western blotting detected significantly elevated levels of autophagy and apoptosis markers in atresia follicles. Interestingly, we found that fibromodulin (FMOD) levels was significantly lower in follicles from fasted hens and that this molecule had an important regulatory role in autophagy. FMOD silencing significantly promoted autophagy and apoptosis in granulosa cells, resulting in hormonal imbalance. FMOD was found to regulate autophagy via the transforming growth factor beta (TGF-ß) signaling pathway. Our results suggest that the increase in autophagy and the imbalance in internal homeostasis cause granulosa cell apoptosis, leading to follicular atresia in the chicken ovary. This finding could provide further insight into broodiness in chicken and provide avenues for further improvements in poultry production.

Galacto-oligosaccharides and xylo-oligosaccharides affect meat flavor by altering the cecal microbiome, metabolome, and transcriptome of chickens.

Studies have shown that prebiotics can affect meat quality; however, the underlying mechanisms remain poorly understood. This study aimed to investigate whether prebiotics affect the flavor of chicken meat via the gut microbiome and metabolome. The gut content was collected from chickens fed with or without prebiotics (galacto-oligosaccharides or xylo-oligosaccharides) and subjected to microbiome and metabolome analyses, whereas transcriptome sequencing was performed using chicken breast. Prebiotic supplementation yielded a slight improvement that was not statistically significant in the growth and production performance of chickens. Moreover, treatment with prebiotics promoted fat synthesis and starch hydrolysis, thus increasing meat flavor by enhancing lipase and α-amylase activity in the blood of broiler chickens. The prebiotics altered the proportions of microbiota in the gut at different levels, especially microbiota in the phyla Bacteroidetes and Firmicutes, such as members of the Alistipes, Bacteroides, and Faecalibacterium genera. Furthermore, the prebiotics altered the content of cecal metabolites related to flavor substances, including 8 types of lysophosphatidylcholine (lysoPC) and 4 types of amino acid. Differentially expressed genes (DEGs) induced by prebiotics were significantly involved in fatty acid accumulation processes, such as lipolysis in adipocytes and the adipocytokine signaling pathway. Changes in gut microbiota were correlated with metabolites, for example, Bacteroidetes and Firmicutes were positively and negatively correlated with lysoPC, respectively. Finally, DEGs interacted with cecal metabolites, especially meat-flavor-related amino acids and their derivatives. The findings of this study integrated and incorporated associations among the gut microbiota, metabolites, and transcriptome, which suggests that prebiotics affect the flavor of chicken meat.

RNA sequencing reveals lncRNA-mediated non-mendelian inheritance of feather growth change in chickens.

BACKGROUND: Long non-coding RNAs (lncRNAs) play an essential role in biological processes. However, the expression patterns of lncRNAs that regulate the non-Mendelian inheritance feather phenotypes remain unknown. OBJECTIVE: This study aimed to compare the expression profiles of lncRNAs in the follicles of the late-feathering cocks (LC) and late-feathering hens (LH) that followed genetic rules and the early-feathering hen (EH) and early-feathering cock (EC) that did not conform to the genetic laws. METHODS: We performed RNA sequencing and investigated the differentially expressed lncRNAs (DElncRNAs) between the early- and late-feathering chickens, which function by cis-acting or participate in the competing endogenous RNA (ceRNA) network. RESULTS: A total of 53 upregulated and 43 downregulated lncRNAs were identified in EC vs. LC, and 58 upregulated and 109 downregulated lncRNAs were identified in EH vs. LH. The target mRNAs regulated by lncRNAs in cis were enriched in the pentose phosphate pathway, TGF-ß signaling pathway and Jak-STAT signaling pathway in EC vs. LC and were associated with the TGF-ß signaling pathway, Wnt signaling pathway, p53 signaling pathway and Jak-STAT signaling pathway in EH vs. LH. In addition, the lncRNA-mediated ceRNA regulatory pathways of hair follicle formation were mainly enriched in the TGF-ß signaling pathway, Wnt signaling pathway, melanogenesis, and calcium signaling pathways. The levels of ENSGALG00000047626 were significantly higher in the late-feathering chickens than in the early-feathering chickens, which regulated the expression of SSTR2 by gga-miR-1649-5p. CONCLUSION: This study provides a novel molecular mechanism of lncRNA's response to the feather rate that does not conform to the genetic laws in chickens.

Comparative Analysis of Skeletal Muscle DNA Methylation and Transcriptome of the Chicken Embryo at Different Developmental Stages.

DNA methylation is a key epigenetic mechanism involved in embryonic muscle development and plays an important role in early muscle development. In this study, we sought to investigate the effects of genome-wide DNA methylation by combining the expression profiles of the chicken embryonic muscle. Genome-wide DNA methylation maps and transcriptomes of muscle tissues collected from different embryonic development points (E7, E11, E17, and D1) were used for whole-genome bisulfite sequencing (WGBS) and RNA sequencing, respectively. We found that the differentially methylated genes (DMGs) were significantly associated with muscle organ development, regulation of skeletal muscle satellite cell proliferation, and actin filament depolymerization. Furthermore, genes TBX1, MEF2D, SPEG, CFL2, and TWF2 were strongly correlated with the methylation-caused expression switch. Therefore, we chose the CFL2 gene to explore its function in skeletal muscle satellite cells, and the in vitro experiments showed that CFL2 acts as a negative regulator of chicken skeletal muscle satellite cell proliferation and can induce cell apoptosis. These results provide valuable data for future genome and epigenome studies of chicken skeletal muscle and may help reveal the molecular mechanisms of potential economic traits.

Long Non-coding RNA Expression Profile in Broiler Liver with Cadmium-Induced Oxidative Damage.

Cadmium pollution is serious heavy metal pollution in environmental pollution and impacts on livestock productivity. However, the effect and mechanisms of cadmium toxicity on the broiler remain unclear. This study aimed to explore the liver oxidative damage and reveal the related long non-coding RNA (lncRNA) expression patterns in the broiler liver with cadmium exposure. The broilers were fed with diets containing CdCl2 and detected the oxidative stress indexes in the liver tissues. Transcriptome sequencing of broiler liver was performed to identify cadmium exposure-related differentially expressed lncRNAs (DElncRNAs). The functions and pathways of DElncRNAs were analyzed by GO and KEGG. The sequencing results were verified by the quantitative real-time polymerase chain reaction. Cadmium exposure induced tissue structure disorder, focal hemorrhage, and irregular hepatocytes in the broiler liver, and significantly decreased GSH level and enzyme activities, and increased MDA expression in the liver. A total of 74 DElncRNAs were obtained in cadmium group compared with the control group, which were enriched in the GO terms, including intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator, branched-chain amino acid biosynthetic process. The enriched KEGG pathways, including lysine biosynthesis, valine, leucine and isoleucine biosynthesis, and pantothenate and CoA biosynthesis, were related to oxidative stress. PCR analysis indicated that the changes in ENSGALG00000053559, ENSGALG00000053926, and ENSGALG00000054404 expression were consistent with sequencing. Our results provide novel lncRNAs involved in oxidative stress in the broiler liver with cadmium exposure.

miR-21-5p Regulates the Proliferation and Differentiation of Skeletal Muscle Satellite Cells by Targeting KLF3 in Chicken.

The proliferation and differentiation of skeletal muscle satellite cells (SMSCs) play an important role in the development of skeletal muscle. Our previous sequencing data showed that miR-21-5p is one of the most abundant miRNAs in chicken skeletal muscle. Therefore, in this study, the spatiotemporal expression of miR-21-5p and its effects on skeletal muscle development of chickens were explored using in vitro cultured SMSCs as a model. The results in this study showed that miR-21-5p was highly expressed in the skeletal muscle of chickens. The overexpression of miR-21-5p promoted the proliferation of SMSCs as evidenced by increased cell viability, increased cell number in the proliferative phase, and increased mRNA and protein expression of proliferation markers including PCNA, CDK2, and CCND1. Moreover, it was revealed that miR-21-5p promotes the formation of myotubes by modulating the expression of myogenic markers including MyoG, MyoD, and MyHC, whereas knockdown of miR-21-5p showed the opposite result. Gene prediction and dual fluorescence analysis confirmed that KLF3 was one of the direct target genes of miR-21-5p. We confirmed that, contrary to the function of miR-21-5p, KLF3 plays a negative role in the proliferation and differentiation of SMSCs. Si-KLF3 promotes cell number and proliferation activity, as well as the cell differentiation processes. Our results demonstrated that miR-21-5p promotes the proliferation and differentiation of SMSCs by targeting KLF3. Collectively, the results obtained in this study laid a foundation for exploring the mechanism through which miR-21-5p regulates SMSCs.

Whole genome re-sequencing identifies unique adaption of single nucleotide polymorphism, insertion/deletion and structure variation related to hypoxia in Tibetan chickens.

BACKGROUND: The adaptation to hypoxia in high altitude areas has great research value in the field of biological sciences. Tibetan chicken has unique adaptability to high-altitude, low pressure and anoxic conditions, and served as a biological model to search for genetic diversity of hypoxia adaption. METHODS: The whole genome re-sequencing technology was conducted to investigate the genetic diversity. RESULTS: In this study, we obtained quantity genetic resource, contained 5164926 single nucleotide polymorphisms (SNPs), 237504 Insertion/Deletion (InDel), 55606 structural variation types in all chromosomes of Tibetan chicken. Moreover, 17154 non-synonymous mutations, 45763 synonymous mutations, 258 InDel mutations and 9468 structural mutations were detected in coding sequencing (CDS) region. Furthermore, SNPs occur in 591 genes, including HIF1A, VEGF, MAPK 8/9/10/11, PPARA/D/G, NOTCH2, and ABCs, which were involved in 14 hypoxia-related pathways, such as VEGF signaling pathway, MAPK signaling pathway, PPAR signaling pathway and Notch signaling pathway. Among them, 19 genes with non-synonymous SNP variation in CDS were identified. Moreover, structure variation in CDS also occurred in the mentioned above genes with SNPs. CONCLUSIONS: This study provides useful targets for clarifying the hypoxia adaptability of the domestication of chickens in Tibetan and may help breeding efforts to develop improved breeds for the highlands.