基于RNA-Seq分析褪黑激素对绒山羊皮肤基因表达的影响

为了研究褪黑激素（melatonin,MT）对绒山羊皮肤毛囊基因表达的影响,筛选与皮肤毛囊发育相关的重要基因．选取处于皮肤毛囊发育生长起始点（5月）的内蒙古绒山羊罕山型个体6只,分别作为埋植组（皮下埋植MT,n=3）和对照组（不埋植MT,n=3）,以绒山羊个体皮肤组织为试验材料,对其进行RNA—Seq测序,利用生物信息学方法对转录组数据进行分析．在测序数据质量控制的基础上,对差异表达基因进行筛选、功能注释和富集分析。结果表明,共筛选获得了462个在埋植组与对照组中差异表达的基因,其中,上调表达208个,下调表达254个;KEGG富集性分析表明差异表达基因主要参与：代谢途径、不饱和脂肪酸生物合成、脂肪酸代谢、癌症信号通路、鳞状细胞癌信号通路、黑色素合成、Wnt信号通路以及补体和凝血级联反应等,Wnt信号可能在褪黑激素调控皮肤毛囊发育中发挥重要作用．、研究结果为进一步阐明褪黑激素影响毛囊生长的分子机理提供了依据。     

鸡皮肤毛囊生长差异基因的生物信息学分析

【目的】 筛选鸡胚胎早期发育期间背部皮肤上皮和间充质中调控毛囊生长的关键基因、生物过程和信号通路,为了解鸡胚胎早期发育期间毛囊生长的分子机制提供参考。【方法】 从GEO数据库中下载基因芯片GSE62882数据集,采用R语言limma包对数据集进行差异分析,分别筛选出胚胎期第7和9天皮肤上皮和间充质差异基因以及皮肤上皮和间充质共同上下调基因,通过STRING在线数据库对差异基因和共同上下调基因进行蛋白互作网络分析,用Cytoscape 3.8.2软件进行可视化,运用R语言中的clusterProfiler程序包对差异基因和共同上下调基因进行GO功能和KEGG通路富集分析。【结果】 皮肤上皮在胚胎期第7和9天共筛选出626个差异基因,获得189条相互作用关系、10个生物过程和4条信号通路;皮肤间充质在胚胎期第7和9天共筛选出690个差异基因,获得326条相互作用关系、10个生物过程和3条信号通路;皮肤上皮和间充质在胚胎期第7和9天共同上调基因26个,共同下调基因48个,共同上下调基因存在34条相互作用关系,主要富集在Wnt信号通路上。【结论】 本研究从鸡胚胎期第7和9天皮肤上皮和间充质中筛选到共同上调基因26个,共同下调基因48个,这些基因主要富集在Wnt信号通路上,为探究上皮和间充质在毛囊生长发育中的作用机制提供理论参考。     

血小板衍生因子A的生理功能研究进展

血小板衍生因子A（platele-derived growth factor A,PDGFA）是PDGF家族中的一种多肽,具有促进细胞生长、刺激细胞与细胞间质相互作用、影响毛囊生长的功能。从PDGFA的蛋白结构、主要基因表达通路Wnt、SHH、BMP和在皮肤、肺部以及绒毛生长中的作用进行综述,探讨PDGFA在毛囊生长和周期维持中发挥的重要作用,以期为进一步研究绒毛生长机理提供参考。     

鸡毒支原体MG-HY株感染鸡气管的转录组学分析

旨在进一步探究鸡毒支原体（MG）感染SPF雏鸡后气管基因组转录水平的变化,筛选出MG感染后参与气管黏膜炎性损伤的差异表达基因和调控通路。使用MG-HY株菌液按0.2 mL·羽^-1经点眼滴鼻感染SPF雏鸡,感染后收集气管组织利用RNA-Seq技术进行测序分析。转录组分析结果显示,在MG感染期间RNA-seq共筛选出3 112个显著（P<0.01）差异表达基因（DEGs）,其中,1 646个上调基因,1 466个下调基因。GO功能分析显示,差异基因主要涉及生物调节、刺激的反应、多细胞生物过程、细胞成分组织或生物发生等生物过程。KEGG-Pathway分析发现差异基因参与黏膜免疫信号传导通路,例如：细胞因子-细胞因子受体相互作用,细胞黏附分子（CAMs）,细胞外基质（ECM）受体相互作用、紧密连接、PPAR信号通路和MAPK信号通路等,表明这些基因参与了MG诱导的雏鸡气管炎症反应和损伤。使用qRT-PCR验证与黏膜免疫相关的13个差异表达的基因,其结果与转录组分析一致。本研究为进一步阐明MG感染导致宿主气管黏膜上皮损伤和黏膜免疫机制提供了基础。     

Transcriptome sequencing of black and white hair follicles in the giant panda

With the completion of the draft assembly of the giant panda genome sequence, RNA sequencing technology has been widely used in genetic research on giant pandas. We used RNA-seq to examine black and white hair follicle samples from adult pandas. By comparison with the giant panda genome, 75 963 SNP loci were labeled, 2426 differentially expressed genes (DEGs) were identified, and 2029 new genes were discovered, among which 631 were functionally annotated. A cluster analysis of the DEGs showed that they were mainly related to the Wnt signaling pathway, ECM–receptor interaction, the p53 signaling pathway, and ribosome processing. The enrichment results showed that there were significant differences in the regulatory networks of hair follicles with different colors during the transitional stage of hair follicle resting growth, which may play a regulatory role in melanin synthesis during growth. In conclusion, our results provide new insights and more data support for research on the color formation in giant pandas.     

Combined microRNAome and transcriptome analysis of follicular phase and luteal phase in porcine ovaries

The aim of this study was to explore the expression difference of miRNAs and mRNAs between the follicular phase (FP) and luteal phase (LP) in porcine ovaries and provide a theoretical basis for the research on mammalian reproductive regulation. RNA‐Seq and miRNA‐Seq were used to identify differentially expressed genes (DEGs) and miRNAs (DEMs) between the FP and LP in ovaries of six sows (3‐year‐old Yorkshire pigs with similar weights and same parities). Bioinformatic analysis was used to screen potential genes and miRNAs related to porcine ovarian function. Real‐time qualitative PCR was used to validate the sequencing results. RNA‐Seq results showed that 3,078 genes were up‐regulated, and 1,444 genes were down‐regulated in the LP compared with the FP, and DEGs were significantly enriched in 242 Gene Ontology (GO) terms and 33 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. miRNA‐Seq identified 112 DEMs, of which 25 were up‐regulated and 87 were down‐regulated in the LP compared with the FP. We obtained 186 intersection genes (IGs) between the 4,522 DEGs and 2,444 target genes predicted from the 112 DEMs. After constructing a miRNA‐gene‐pathway network, we identified key miRNAs and genes including miR‐17‐3p, miR‐214, miR‐221‐5p, miR‐125b, FGF1, YWHAG, YWHAZ, FDFT1 and DHCR24, which are enriched in Hippo and PI3K‐Akt signalling pathways, and various metabolic pathways. These results indicate that these key genes and miRNAs may play important roles in the developmental transition from FP to LP in porcine ovaries and represent candidate targets for further study.     

藏鸡与白羽肉鸡垂体转录组差异表达研究

试验旨在通过对藏鸡和白羽肉鸡垂体转录组测序和生物信息学分析,筛选出与鸡生长发育相关的差异表达基因（differentially expression genes,DEGs）及信号通路。本研究选用42日龄健康藏鸡和白羽肉鸡各3只,分别采集垂体组织利用Illumina HiSeq 2000平台进行转录组mRNA测序,对筛选到的DEGs筛选DEGs,GO和KEGG数据库功能注释和富集分析,实时荧光定量PCR验证随机挑选的DEGs表达水平。结果显示,藏鸡和白羽肉鸡分别获得126 094 302和125 666 442条clean reads。与白羽肉鸡相比,藏鸡垂体组织中共有DEGs 392个,其中196个为上调基因,196个为下调基因（P<0.05）,其中包括生长激素（GH）基因、生长激素释放激素受体（GHRHR）基因和胰岛素样生长因子2 mRNA结合蛋白1（IGF2BP1）基因。GO和KEGG富集分析发现,DEGs显著富集于细胞黏附分子信号通路和神经活性配体-受体相互作用信号通路等细胞通讯相关的信号通路,GH、GHRHR和IGF2BP1基因也显著富集于神经活性配体-受体相互作用信号通路。实时荧光定量PCR结果显示,转录组测序结果准确可靠。综上研究,本研究初步揭示了影响藏鸡和白羽肉鸡生长发育速度差异的关键候选基因和信号通路,为了解鸡垂体组织调节生长发育的分子机制提供了理论依据。     

Pyridoxine regulates hair follicle development via the PI3K/Akt,Wnt and Notch signalling pathways in rex rabbits

This study was conducted to evaluate the effect of pyridoxine on the development of hair follicles in Rex rabbits and the underlying molecular mechanism. Two hundred 3-month-old Rex rabbits were randomly divided into 5 groups and fed diets supplemented with 0, 5, 10, 20, or 40 mg/kg pyridoxine. The hair follicle density on the dorsal skin and the gene and protein expression levels of components of the phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB or Akt), Wnt, Notch and bone morphogenetic protein (BMP) signalling pathways were measured. In addition, free hair follicles were isolated from Rex rabbits and cultured with pyridoxine in vitro to measure hair shaft growth. Furthermore, dermal papilla cells (DPC) were isolated from the skin of Rex rabbits and cultured with pyridoxine in vitro to measure the gene and protein expression levels of components of the PI3K/Akt, Wnt, Notch and BMP signalling pathways. The results showed that the addition of dietary pyridoxine significantly increased the total follicle density, secondary follicle density, and secondary-to-primary ratio (S/P, P < 0.05), that the growth ratio of hair stems was promoted by pyridoxine in basic culture medium, and that the growth length of tentacle hair follicles cultured in the pyridoxine group was longer than that in the control group (P < 0.05). In addition, pyridoxine changed the DPC cycle progression and promoted cell proliferation, and appropriate concentrations of pyridoxine (10 and 20 μmol/L) significantly inhibited cell apoptosis (P < 0.05). Pyridoxine significantly affected the gene expression of components of the PI3K/Akt, Wnt and Notch signalling pathways in the skin and DPC of Rex rabbits (P < 0.05), increased the levels of phosphorylated catenin beta 1 (CTNNB1) and Akt, and decreased the level of phosphorylated glycogen synthase kinase 3 beta (GSK-3β) (P < 0.05). Therefore, the molecular mechanism by which pyridoxine promotes hair follicle density in Rex rabbits probably occurs through activation of the PI3K/Akt, Wnt and Notch signalling pathways, prolonging hair follicle growth and delaying the onset of telogen.     

鸡Wnt3a基因组织表达差异与SNPs对鸡毛囊密度性状的遗传效应

Wnt家族成员3a （Wnt family member 3a,Wnt3a）基因是Wnt信号通路的重要成员,Wnt信号通路在动物皮肤毛囊的发生发育中具有重要作用。试验以Wnt3a为候选基因,旨在探讨鸡Wnt3a基因的组织表达差异与单核苷酸多态性（single nucleotide polymorphisms,SNPs）位点对鸡皮肤毛囊密度性状的遗传效应。用实时荧光定量PCR检测Wnt3a基因在鸡不同组织中的表达差异;用PCR扩增直接测序法对Wnt3a基因的SNPs位点进行筛查,验证和分析不同SNPs位点基因型背部毛囊密度的差异;用实时荧光定量PCR技术检测不同品种鸡皮肤组织中Wnt3a基因mRNA表达差异。结果表明,Wnt3a mRNA在皮肤组织中表达量最高,显著高于其他组织（P<0.05）,其次是肝脏、睾丸、卵巢和下丘脑,心脏、胸肌、垂体和脾脏等组织表达量较低。在Wnt3a基因第2和3外显子区各筛选到1个SNP位点：g.2587569 G>A和g.2555812 T>C;在第3内含子区筛选到1个SNP位点：g.2555377 T>C。卡方检验结果显示,3个位点均极显著偏离Hardy-Weinberg平衡。群体遗传参数分析发现,g.2555812 T>C位点为中度多态,有效等位基因数为1.7,遗传变异程度较g.2587569 G>A和g.2555377 T>C位点高。SNP位点分型与毛囊密度的关联分析结果表明,g.2555377 T>C位点CC基因型个体的毛囊密度显著高于TT和CT基因型个体（P<0.05）。在毛囊密度具有显著差异的不同品种鸡的背部皮肤组织中,Wnt3a mRNA表达差异显著（P<0.05）。该研究结果为进一步深入解析Wnt3a在鸡皮肤毛囊生长发育中的作用、筛选与毛囊密度性状相关的辅助育种标记提供了基础资料。     

羊胎儿期皮肤毛囊发育及调控机制的研究概述

中国绵山羊养殖历史悠久,品种资源丰富,在国民经济中起重要作用。羊毛按纤维类型可分为同质毛和异质毛,同质毛的特点是被毛由同一类型的羊毛纤维组成,其特点是羊毛的纤维直径、长度、卷曲等外表特征基本相同,异质毛的特点是被毛中兼有绒毛、有髓毛、无卷曲和少卷毛等不同类型的羊毛纤维,其化学、物理性能均不相同。毛囊是控制哺乳动物被毛生长的重要器官,是唯一具有周期性发育并可终生再生的器官,控制着被毛发生发育与自然脱落。毛囊依据其形成时间和结构可分为初级毛囊(primary follicles)和次级毛囊(secondary follicles),初级毛囊生长髓质发达的粗毛,次级毛囊产生无髓毛的绒毛。有髓毛较粗长且弯曲少,多用于加工粗纺织品、毛毯、地毯、毡制品;无髓毛直径一般不超40 μm,弯曲多,是毛纺工业的优质原料。毛囊的发育受到不同信号通路和基因的影响,如Wnt信号通路、骨形态发生蛋白质(BMP)信号通路、成纤维细胞生长因子(FGF)信号通路、音猬因子(SHH)信号通路、Notch信号通路等以及角蛋白家族基因、BMPs家族基因、同源异型盒基因(Hox)等。作者对国内外对绵山羊胎儿期皮肤毛囊发育及调控机制进行综述,以期为提高以绵山羊被毛产量及改善被毛品质为主要育种目标的选育提供参考。     

不同生理阶段边鸡卵巢转录谱的构建及卵泡发育相关基因的分析

本研究通过分析不同生理阶段边鸡卵巢转录表达谱,旨在找出与卵泡发育相关的基因并进行验证,以揭示边鸡就巢性强的遗传机制.构建代表不同生理期即70日龄(70 d,卵巢发育期)、165日龄(165 d,开产期)、220日龄(220 d,产蛋高峰期)和330日龄(330 d,就巢期)的边鸡卵巢基因表达谱,用转录组表达定量(RSE...     

Heat stress decreased hair follicle population in rex rabbits

The aim of this study was conducted to investigate the effect of heat stress on the hair follicle population and related signalling pathways in rex rabbits. Forty‐eight rabbits were randomly divided into two groups: one group in a high ambient environment (32 ± 2°C, heat stress) and the other group with normal temperature (20 ± 2°C, control). The results show that heat stress decreased the body weight gain and feed conversion rate, rabbit hair length and hair follicle density (p < 0.05). Besides, heat stress suppressed the gene expression of noggin, insulin‐like growth factor 1 (IGF‐1) and IGF‐1 receptor and protein expression of phosphorylated mechanistic target of rapamycin (mTOR) in rabbit skin (p < 0.05), while stimulated significantly the gene expression of bone morphogenetic protein 2 (BMP2) and BMP4 (p < 0.05). Heat exposure did not alter significantly the gene expression of alkaline phosphatase, versican and hepatocyte growth factor compared with the control (p > 0.05). In conclusion, noggin‐BMP, IGF‐1 and mTOR signalling pathways may be associated with the process of heat stress‐repressing hair follicle development.     

Expression profiles and associations of FGF1 and FGF10 with intramuscular fat in Tibetan chicken

ABSTRACT

1. FGF1 and FGF10, two paracrine members of the fibroblast growth factor (FGF) gene family, play critical roles in the development, structural and metabolic remodelling of adipose tissue.

2. The objective of this study was to investigate the expression profiles of FGF1 and FGF10 genes in breast muscle and thigh muscle in 5 developmental stages (1, 81, 119, 154 and 210 d old) in Tibetan chickens. The possible relationships between expression of these genes and intramuscular fat (IMF) content were analysed in Tibetan chickens.

3. Expression profiles showed that FGF1 and FGF10 mRNA were ubiquitously expressed in various tissues of 154-d-old Tibetan chickens. Lung tissue contained the highest amount of FGF1 and FGF10 mRNA while breast muscle and thigh muscle exhibited lower levels of FGF1 and FGF10 mRNA in both males and females compared with other tissues (P < 0.05). Temporal expression of FGF1 and FGF10 in breast and thigh muscle showed similar tendencies in males and females, respectively, with peaks in thigh muscle at 119-d-old and breast muscle in 1-d-old males and females, respectively.

4. Correlation analysis suggested that gender had an influence on the relationships of FGF1 and FGF10 expression with IMF content in thigh muscle. The RNA levels of FGF1 and FGF10 genes in male thigh muscle were positively related to IMF content of Tibetan chickens (P < 0.01), while the correlations were shown to be negative in female thigh muscle (P > 0.05).

5. These results provide a basis for functional elucidation of FGF1 and FGF10 genes on adipocyte development and intramuscular fat deposition, as well as selective breeding and resource exploration of local poultry breeds.     

EDA基因及其信号通路在动物皮肤毛囊发育中作用研究进展

EDA基因的突变会引起多种外胚层附属物的减少或是缺乏,像头发、牙齿、汗腺、皮脂腺和乳腺等外胚层附属物的生长发育,都需要EDA的参与。EDA信号通路在多种类型毛囊的发生、毛干的形成和皮脂腺的形态发生中发挥作用,它可控制胚胎上皮细胞的命运,调控毛囊细胞的分化过程。作者简述了EDA基因及其信号通路与动物毛囊发育的关系,介绍了EDA基因的功能和蛋白分子结构,以及它主要的两种转录体EDA1和EDA2的功能与作用机制,EDA1-EDAR系统在皮肤衍生结构的发育中发挥主要作用,EDA1和EDAR的突变均会导致HED。阐述了EDA基因信号通路调控毛囊组织生长发育机理以及与多个信号通路的关联,EDA信号可控制毛囊生长期到退行期的转换,对毛囊循环周期中退行期毛囊角质细胞的细胞凋亡起调控作用。EDA-EDAR系统是对皮肤附属器官发育有重要作用的Wnt信号通路的一个重要的效应器,其紧接着出现在Wnt诱导信号的下游。它依靠下游的NF-κB通路的活动来调控靶基因,通过刺激NF-κB调整Wnt、SHH、FGF、和TGF-β信号通路的效应器和抑制因子的转录来发挥作用,调控上皮细胞和间充质细胞还包括组织内的相互作用。结论认为EDA信号通路参与毛囊细胞的发育与分化调控,对动物毛囊生长发育起重要影响作用。     

Meat quality in broilers affected with inflammation of the feather follicles

Broilers with feather follicle inflammation and birds free of this disorder were selected from the broiler chickens kept on plastic (bralen-)coated metallic slats. Both groups of broilers were killed on a sanitary slaughter line and the samples of breast and thigh muscles were analyzed for the basic composition and characteristics of the metabolism of nitrogenous and lipidic components. The content of individual amino acids in the muscles and the proportion of fatty acids in the intramuscular fat of broilers were determined for the evaluation of nutritive value. The samples of the affected spots of skin and samples of organs (liver) were subjected to microbiological examination. The resultant finding represented the common mesophilous microflora. No substantial statistically significant differences in chemical characteristics were found between the two groups. It follows from the results that the inflammation of feather follicles is a local skin disorder with no effect on the quality of the meat.     

Genome-wide Association Study of Teats Number Traits in Kele Pigs

Teats number is one of the most important reproductive traits,and closely related to the economic benefit in pig industry.In order to reveal the underlying genetics of left teats number,right teats number and total teats number traits,a genome-wide association study（GWAS）was performed.Samples of DNA were collected to genotyping for 22 Kele pigs using the Illumina Porcine SNP 60K Chip.The GWAS was performed using a mixed-effects model and linear regression approach.When a genome-wide threshold was determined using the Bonferroni method（P<2.06E-5),4 single nucleotide polymorphism（SNP）markers were potentially associated with left teats number,right teats number and total teat number.However,3 SNPs were significant associated and 18 SNPs were potentially associated in chromosomes level.304 Ensembl genes were retrieved around 1 cM of the associated SNPs.The candidate genes in Wnt and Fgf signaling pathway（BTRC,FGF5,FGF8,BMP3,RASGEF1B and HMGB3）might have effect on target traits.These results provided valuable information about the selective breeding for Kele pigs.     

可乐猪乳头数性状全基因组关联分析

乳头数性状是猪最重要的繁殖性状之一,与养猪业的经济效益密切相关。本试验以Illunima Porcine SNP 60K芯片对22头可乐猪进行基因分型,通过Plink 1.07软件,基于混合线性模型对左乳头数、右乳头数和总乳头数性状进行全基因组关联分析。经过严格的质量控制和多重检验之后,共鉴定出全基因组水平潜在关联的SNPs位点4个（P<2.06E-5）;染色体水平显著关联位点3个,潜在关联位点18个;在关联SNP位点可能连锁的上、下游1 cM区间内检索到304个基因,其中Wnt及Fgf信号通路中的候选基因BTRC、FGF5、FGF8和BMP3、RASGEF1B、HMGB3可能影响猪的乳头数性状或繁殖性状。通过全基因组关联分析策略发掘出的关联SNP位点及潜在候选基因,将为可乐猪的选育保种奠定基础。     

Effects of Wnt10b on dermal papilla cells via the canonical Wnt/β‐catenin signalling pathway in the Angora rabbit

Wnt10b is a member of Wnt family that plays a variety of roles in biological functions, including those in the development of hair follicles. To investigate the effect of Wnt10b on hair growth in the Angora rabbit and to determine the underlying molecular mechanism, we cultured dermal papilla (DP) cells with exogenous Wnt10b in vitro. We observed the expressions of downstream critical gene β‐catenin and lymphoid enhancer‐binding factor 1 (LEF1) in Wnt/β‐catenin pathway. The levels of β‐catenin mRNA and protein were higher in the Wnt10b group of DP cells than in the Control group, and the mRNA level of LEF1 in the Wnt10b group was higher than in the Control group. Moreover, translocation of β‐catenin from cytoplasm to nucleus was activated in the Wnt10b group. Furthermore, the mRNA levels of the hair follicle‐regulatory genes, insulin‐like growth factor‐1 (IGF‐1) and alkaline phosphatase (ALP), and the protein activity of ALP was also upregulated in the Wnt10b group compared to their corresponding levels in the Control group. These data suggest that Wnt10b could activate the canonical Wnt/β‐catenin signalling pathway to induce DP cells in the Angora rabbit. In addition, the proliferation of DP cells was significantly promoted when cultured with Wnt10b for 48 and 72 hr, suggesting that Wnt10b plays a pivotal role in the proliferation and maintenance of DP cells in vitro. In conclusion, this study demonstrates that Wnt10b may promote hair follicle growth in Angora rabbit through the canonical Wnt/β‐catenin signalling pathway that promotes the proliferation of DP cells.