摘要
本研究通过比较高、低质量蛋壳力学特性、超微结构和蛋鸡的子宫组织形态、基因表达差异,挖掘调控产蛋后期蛋壳质量的重要候选基因和信号通路。选用80周龄海兰褐蛋鸡12只,6只产鸡蛋蛋壳质量相对好[蛋壳强度为(3.54±0.11)kg/cm^(2),蛋壳厚度为(385.06±12.46)mm]的为高质量组(HQE组),6只产鸡蛋蛋壳质量相对差[蛋壳强度为(2.54±0.07)kg/cm^(2),蛋壳厚度为(256.17±18.36)mm]的为低质量组(LQE组),每组每天收集6枚鸡蛋,累计收集5 d。鸡蛋称重后采用蛋壳强度仪测定蛋壳强度,螺旋测微仪测定蛋壳厚度,计算蛋壳比例;采用扫描电子显微镜观察蛋壳的超微结构,测定蛋壳有效层厚度、乳突层厚度和乳突宽度,计算乳突层厚度比例;运用组织切片技术比较2组蛋鸡子宫组织形态差异;运用转录组测序(RNA-seq)技术,比较2组子宫样本之间的差异表达基因(DEGs),并对其进行GO和KEGG富集分析。结果显示:与LQE相比,HQE组蛋壳强度、蛋壳厚度、蛋壳重和蛋壳比例显著增加(P<0.05),蛋壳有效层厚度、乳突层厚度和乳突层厚度比例显著增加(P<0.05),蛋鸡子宫黏膜组织管状腺细胞排布更紧密,子宫黏膜绒毛高度显著增加(P<0.05)。以LQE组为对照组,在HQE组子宫黏膜中筛选到303个DEGs,其中103个表达上调,200个表达下调;GO功能分析显示DEGs主要富集在DNA复制、免疫应答等生物过程;KEGG分析发现DEGs主要富集在细胞因子-细胞因子受体互作、Th17细胞分化、肿瘤坏死因子(TNF)、Toll样受体等与免疫、炎症反应相关的信号通路,其中信号转导和转录激活因子1(STAT1)、CC趋化因子受体2(CCR2)、CC趋化因子配体4(CCL4)、CC趋化因子配体20(CCL20)、白细胞介素18受体1(IL18R1)等基因在以上通路中反复富集,且在HQE组显著下调。由以上结果可知,产蛋后期蛋壳力学特性和超微结构变差的同时伴随着蛋鸡子宫组织形态变差,功能分析显示子宫黏膜组织上细胞因子-细胞因子受体互作等信号通路可能通过调控子宫免疫应答、炎症反应相关基因的表达在蛋壳矿化过程中发挥重要作用,该结果可为精准改善产蛋后期蛋壳质量提供理论依据。
This study aimed to identify important candidate genes and signaling pathways associated with eggshell quality by comparing the mechanical property,ultrastructure of higher and lower quality eggshells as well as the uterine morphology and gene expression of laying hens.Twelve Hy-Line brown laying hens aged 80-weeks were selected in the experiment,and 6 hens with higher eggshell quality[with(3.54±0.11)kg/cm^(2) eggshell strength and(385.06±12.46)mm eggshell thickness]were selected as the high quality group(HQE group),and 6 hens with lower eggshell quality[with(2.54±0.07)kg/cm^(2) eggshell strength and(256.17±18.36)mm eggshell thickness]were selected as the low quality group(LQE group).Each group collected 6 eggs a day for 5 days.After weighing of the egg,the eggshell strength was measured with an eggshell strength meter,the eggshell thickness was determined by a spiral micrometer,and then eggshell ratio was calculated;the ultrastructure of eggshell such as the effective layer thickness,mammillary layer thickness and mammillary width were observed with a scanning electron microscope,and the mammillary thickness layer ratio was calculated;histological section technique was used to compare the uterine morphological differences between two groups;RNA-seq technology was employed to compare the differentially expressed genes(DEGs)of uterine samples between the two groups,and these DEGs were analyzed for GO and KEGG enrichment.The results showed that,in comparison with the LQE group,the eggshell strength,eggshell thickness,eggshell weight and eggshell ratio in HQE group were significantly increased(P<0.05),as well as the eggshell effective layer thickness,mammillary layer thickness and mammillary layer thickness ratio(P<0.05).In respect to uterine morphology,the mucosal tubular gland cells of uterine in HQE group were more tightly arranged,and the height of the villi of the uterine mucosa was significantly higher than in LQE group(P<0.05).The LQE group was selected as the control group,and 303 DEGs were identified,including 103 up-regulated DEGs and 200 down-regulated DEGs.GO analysis suggested the DEGs were mainly enriched in DNA replication and immune response and other related biological processes.KEGG analysis indicated enriched pathways primarily associated with immune and inflammatory response,including cytokine-cytokine receptor interaction,Th17 cell differentiation,tumor necrosis factor(TNF)and Toll-like receptor signaling pathways,among which signal Transducer and activator of transcription 1(STAT1),CC chemokine receptor 2(CCR2),CC chemokine ligand 4(CCL4),CC chemokine ligand 20(CCL20)and interleukin 18 receptor 1(IL18R1)were repeatedly enriched in these pathways,and these genes were significantly down-regulated in the HQE group.From the above results,it can be concluded that the deterioration of mechanical properties and ultrastructure of eggshell in late laying period was accompanied by the deterioration of uterine tissue morphology.By functional analysis,signaling pathways such as cytokine-cytokine receptor interaction in uterine mucosa might play an important role in the process of eggshell mineralization by regulating the expression of genes related to uterine immune and inflammatory responses.These results provided theoretical basis for improving eggshell quality in the late laying period of hens.
作者
邵丹
瞿源
刘良基
童海兵
SHAO Dan;QU Yuan;LIU Liangji;TONG Haibing(Jiangsu Institute of Poultry Science,Yangzhou 225125,China)
出处
《动物营养学报》
北大核心
2025年第2期1221-1233,共13页
CHINESE JOURNAL OF ANIMAL NUTRITION
基金
现代农业产业技术体系专项资金项目(CARS-40-S23)。
关键词
蛋鸡
产蛋后期
蛋壳
转录组测序
差异表达基因
信号通路
laying hens
late laying period
eggshell
RNA-seq
differentially expressed gene
signaling pathway
