Li Cong-Jun, Lin Shudai, Ranilla-García María Jose, Baldwin Ransom L
Animal Genomics and Improvement Laboratory, Agricultural Research Service, Beltsville Agricultural Research Center, USDA, Beltsville, MD, United States.
Department of Animal Genetics, Breeding and Reproduction, College of Animal Science, South China Agricultural University, Guangzhou, China.
Front Vet Sci. 2019 Jul 2;6:214. doi: 10.3389/fvets.2019.00214. eCollection 2019.
Previous research has demonstrated a positive relationship between dietary Metabolisable Energy Intake (MEI) and increased maintenance energy costs associated with the visceral tissues. Limitations in understanding this relationship include a lack of access to samples to assess regulatory control of the putative response gastrointestinal tissues to nutrients. This experiment was conducted with a single nutrient (starch hydrolysate) infused (7 d) directly into the intestine to mimic typical changes in post-ruminal starch delivery in dairy production settings. Duodenal epithelial samples collected via biopsy were evaluated using next-generation sequencing technology (RNA-Seq) to validate the use of this approach for the profiling and comparison of the transcriptome of cattle intestinal epithelial tissues. Samples of intestinal epithelial tissue were collected prior to and during the infusion of starch hydrolysate. Biopsies were collected on day 0 before and day 1, day 3, and day 7 during the infusion. Additionally, samples were collected on day 1 and day 7 after infusion was discontinued (Day 8 and Day14 of the experiment). Evaluation of RNA-seq data revealed dynamic changes in global gene expression during infusion. On day 7 of the infusion, 1490 genes were found to be differentially expressed (DE) compared to the day 0 control samples with FDR < 0.05, vs. 105 genes on day 1 and 246 genes on Day 3. However, on day 8, after infusion was terminated for 24 h, only 428 genes were identified as differentially expressed compared to day 0 and only 107 genes continued to be identified by Day 14. Thus, the apparent differential expression of these genes is putatively a result of the single nutrient infused. Further, performing function and pathway analysis of the identified DE genes using IPA, we observe changes in digestive system development, and function pathways are among the primary functions of the DE genes, as well as immune response elements. Finally, primary transcription regulators such as PTH, JUN, WNT, and TNFRSF11B were identified as the activated upstream regulators for specific future focus. Using a serial biopsy approach we are able to identify differentially expressed genes from cow duodenal epithelial tissue in response to a short-term perturbation with infused starch hydrolysate.
先前的研究表明，日粮可代谢能量摄入量（MEI）与内脏组织相关的维持能量成本增加之间存在正相关关系。理解这种关系的局限性包括无法获取样本以评估假定的胃肠道组织对营养物质反应的调节控制。本实验通过将单一营养素（淀粉水解物）直接注入肠道（7天）来模拟奶牛生产环境中瘤胃后淀粉输送的典型变化。通过活检收集的十二指肠上皮样本使用下一代测序技术（RNA测序）进行评估，以验证这种方法用于牛肠上皮组织转录组分析和比较的有效性。在注入淀粉水解物之前和期间收集肠上皮组织样本。在注入前的第0天以及注入期间的第1天、第3天和第7天进行活检。此外，在停止注入后（实验的第8天和第14天）的第1天和第7天收集样本。RNA测序数据评估显示注入期间全局基因表达存在动态变化。在注入的第7天，与第0天的对照样本相比，发现1490个基因差异表达（DE），错误发现率（FDR）<0.05，而第1天为105个基因，第3天为246个基因。然而，在注入终止24小时后的第8天，与第0天相比，仅鉴定出428个差异表达基因，到第14天仅继续鉴定出107个基因。因此，这些基因明显的差异表达推测是单一注入营养素的结果。此外，使用IPA对鉴定出的差异表达基因进行功能和通路分析，我们观察到消化系统发育和功能通路的变化是差异表达基因的主要功能之一，以及免疫反应元件。最后，确定了诸如甲状旁腺激素（PTH）、Jun原癌基因（JUN）、Wnt信号通路相关蛋白（WNT）和肿瘤坏死因子受体超家族成员11B（TNFRSF11B）等主要转录调节因子作为未来特定研究重点的激活上游调节因子。使用连续活检方法，我们能够识别奶牛十二指肠上皮组织中因注入淀粉水解物的短期扰动而差异表达的基因。
