College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China.
College of Science, Gansu Agricultural University, Lanzhou 730070, China.
Poult Sci. 2024 May;103(5):103582. doi: 10.1016/j.psj.2024.103582. Epub 2024 Feb 22.
Small peptides are nutrients and bioactive molecules that have dual regulatory effects on nutrition and physiology. They are of great significance for maintaining the intestinal health and production performance of broilers. We here cultured the primary small intestinal epithelial cells (IEC) of chicken in a medium containing L-Leu (Leu) and L-Leu-L-Leu (Leu-Leu) for 24 h. The untreated cells were considered as the control group. The growth, proliferation, and apoptosis of IEC were examined. By combining RNA-seq and label-free sequencing technology, candidate genes, proteins, and pathways related to the growth, proliferation, and apoptosis of IEC were screened. Immunofluorescence detection revealed that the purity of the isolated primary IEC was >90%. The Leu-Leu group significantly promoted IEC growth and proliferation and significantly inhibited IEC apoptosis, and the effect was better than those of the Leu and control groups. Using transcriptome sequencing, four candidate genes, CCL20, IL8L1, IL8, and IL6, were screened in the Leu group, and one candidate gene, IL8, was screened in the Leu-Leu group. Two candidate genes, IL6 and RGN, were screened in the Leu-Leu group compared with the Leu group. Nonquantitative proteomic marker sequencing results revealed that through the screening of candidate proteins and pathways, found one growth-related candidate protein PGM3 and three proliferation-related candidate proteins RPS17, RPS11, and RPL23, and two apoptosis-related candidate proteins GPX4 and PDPK1 were found in the Leu-Leu group compared with Leu group. In short, Leu-Leu could promote IEC growth and proliferation and inhibit IEC apoptosis. On combining transcriptome and proteome sequencing technologies, multiple immune- and energy-related regulatory signal pathways were found to be related to IEC growth, proliferation, and apoptosis. Three candidate genes of IL8, IL6, and RGN were identified, and six candidate proteins of PGM3, RPS17, RPS11, RPL23, GPX4, and PDPK1 were involved in IEC growth, proliferation, and apoptosis. The results provide valuable data for preliminarily elucidating small peptide-mediated IEC regulation pathways, improving the small peptide nutrition theoretical system, and establishing small peptide nutrition regulation technology.
小肽是一种具有营养和生物活性双重调节作用的物质,对维持肉鸡的肠道健康和生产性能具有重要意义。我们在此使用含有 L-Leu(Leu)和 L-Leu-L-Leu(Leu-Leu)的培养基培养鸡的原代小肠上皮细胞(IEC)24 小时。未处理的细胞作为对照组。检测 IEC 的生长、增殖和凋亡情况。结合 RNA-seq 和无标记测序技术,筛选与 IEC 生长、增殖和凋亡相关的候选基因、蛋白质和途径。免疫荧光检测显示,分离的原代 IEC 的纯度>90%。Leu-Leu 组显著促进 IEC 的生长和增殖,显著抑制 IEC 的凋亡,效果优于 Leu 组和对照组。通过转录组测序,在 Leu 组中筛选出 4 个候选基因 CCL20、IL8L1、IL8 和 IL6,在 Leu-Leu 组中筛选出 1 个候选基因 IL8。与 Leu 组相比,Leu-Leu 组筛选出 2 个候选基因 IL6 和 RGN。非定量蛋白质组标志物测序结果显示,通过筛选候选蛋白和途径,在 Leu-Leu 组中发现一个与生长相关的候选蛋白 PGM3 和三个与增殖相关的候选蛋白 RPS17、RPS11 和 RPL23,以及两个与凋亡相关的候选蛋白 GPX4 和 PDPK1。总之,Leu-Leu 可促进 IEC 的生长和增殖,抑制 IEC 的凋亡。结合转录组和蛋白质组测序技术,发现多个与免疫和能量相关的调节信号通路与 IEC 的生长、增殖和凋亡有关。鉴定出 IL8、IL6 和 RGN 三个候选基因,PGM3、RPS17、RPS11、RPL23、GPX4 和 PDPK1 六个候选蛋白参与 IEC 的生长、增殖和凋亡。这些结果为初步阐明小肽介导的 IEC 调节途径提供了有价值的数据,完善了小肽营养理论体系,建立了小肽营养调控技术。