GSE193831
Exploring differentially expressed genes related to metabolism by RNA-Seq in porcine embryonic fibroblast after insulin treatment
Purpose: In this study,RNA-Seq technology was used to analyse the effects of insulin on glucose and lipid metabolism of fibroblasts in pigs. We focus mainly on the cellular mechanisms of insulin signalling and on new insights into pathways of primary relevance to insulin's metabolic effects. Methods: porcine embryonic fibroblast(PEF) mRNA profiles of control group and insulin treated group were generated by deep sequencing, in triplicate,using Illumina HiSeq 2000. Paired-end clean reads were aligned to the reference genome using Hisat2. Featurecounts was used to count the read numbers mapped to each gene. Then, the expected number of fragments per kilobase of transcript sequence per million base pairs sequenced (FPKM) was calculated for each gene based on the length of the gene and the read counts mapped to the gene. qRT–PCR validation was performed using TB Green assays. Results: A total of 439,950,662 clean reads filtered from 458,016,778 paired-end raw reads were identified, Of these sequence reads, 94.89%–97.20% was successfully aligned with the Sus scrofa. After insulin treated, a total of 801 and 1176 genes were differentially expressed after 48 and 72 h, respectively. Of these, 272 up-regulated genes and 264 down-regulated genes were common to both time points. The functions of the differentially expressed genes were annotated by a GO analysis, the biological processes related lipid metabolism and cell cycle were dominant. Via KEGG enrichment analysis, we found the DEGs were significantly enriched in  IL-17 signaling pathway, PI3K-Akt signaling pathway, pyruvate metabolism, and others pathway related to lipid metabolism. These results elucidate the transcriptomic response to insulin in PEF. Conclusions: Our data support a role of the insulin on lipid and glucose metabolism, show an association among insulin, glycolipid metabolism and cell cycle in PEF . Our study identified some important genes associated with metabolic and these genes can be used to increase our understanding of the mechanism of relieving the metabolic disturbance induced by insulin. Additionally, PEF treated with insulin would provides a useful cell model for glycolipid metabolism disorder such as diabetes. Overall design: Prior to add insulin,PEF lines were thawed and cultured,The cells were collected at 48 hours and 72 hours after the addition of insulin. The control samples were taken at 0 hours and three biological replicates were collected at each time point. Then the PEF mRNA profiles of control samples and insulin treated were generated by deep sequencing.