Molecular insights into the lipid-carbohydrates metabolism switch under the endurance effort in Arabian horses
Purpose: RNA-sequencing (RNA-seq) was used to identify the changes in gene expression profile to describe the metabolism adaptation at the whole transcriptome of blood  to endurance effort. Samples from ten Arabian horses were taken before and after a 120km long endurance ride. Overall design: Methods: The blood samples from each horse were taken twice – before and after the run,  by jugular venipuncture into Tempus™ Blood RNA Tube (Applied Biosystems, Thermo Fisher Scientific, Waltham, MA, USA) and stored at -20°C (n=20) until analysis. RNA was isolated with PureLink™ RNA mini kit. The quality and quantity of obtained libraries (the TruSeq RNA Kit v2 kit (Illumina, San Diego, CA, USA) ) were assessed using Qubit 2.0 (Qubit™ dsDNA BR AssayKit, Invitrogen, Waltham, MA, USA ) and TapeStation 2200 (D100 screencaps, Agilent). Next, the cDNA libraries were sequenced on NextSeq 500 Illumina platform (Illumina, San Diego, CA, USA) and NextSeq 500/550 High Output KIT v 2.5 (75 cycles) according to protocol. The qPCR validation was used to confirm RNA-seq data. Results: The results, including KEGG (Kyoto Encyclopaedia of Genes and Genomes) and GO (Gene Ontology) analyses allowed us to describe the most significantly up-regulated – ARV1, DGAT2, LIPE, APOA2, MOGAT1, MOGAT2, GYS1, GYS2 and down-regulated - ACACA, ACACB, FADS1, FADS2 genes involved in carbohydrate and lipid metabolism. Also, the increased expression of RAF1, KRAS and NRAS genes involved in the Insulin pathway and PI3K-Akt was shown. The general insight into processes described  in the present study may confirm the thesis of the lipid-carbohydrates metabolism switch and provide the basis for further research. Conclusions: In our research, we aimed to unravel the complexity of metabolic changes at the molecular level in Arabian horses after exposure to heavy endurance physical effort. The presented results have shown significant changes in carbohydrate and fat metabolism reflected a modification of several pathways critical for the maintenance of homeostasis as well as energy use and production. The general insight into the processes described in the present study seems to confirm the thesis of the lipid-carbohydrates metabolism switch under the endurance effort and allow us to propose the panel of marker genes critical for endurance predisposition. The whole transcriptome sequencing of equine blood was performed for 10 samples using NextSeq 550High Output KIT v2.5 (Illumina) into 75 single-end cycles.