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Gene expression profile of human mesenchymal stromal cells (MSCs) exposed to hypoxic and pseudohypoxic preconditioning

ABSTRACT: We performed gene expression profiling by RNA sequencing (RNA-Seq) to fully characterize the gene regulation effect induced by exposure of MSCs to two hypoxic preconditioning methods. We determined the gene expression profile of 6 populations of human MSCs isolated from bone marrow and preconditioned for 6h in 2% O2 (hypoxia) or with 40μM Vadadustat (pseudohypoxia), compared to control cells cultured in 21% O2. There were 5 populations isolated from males and 1 population isolated from a female. The mean age of the BM-MSCs donors was 50.5 years and the median was 44 years. RNA-Sequencing was performed using the Illumina platform. For each library an average of 16 - 20 million read pairs were generated. Quality control checks of the sequencing raw data was conducted with FastQC, while adapter-trimming was performed with BBDUK2. The relative expression of transcripts were quantified for each donor using the Salmon program v0.8.1. Fastq files were mapped to the Homo_sapiens.GRCh37 reference genome using HISAT2 software version 2.1.0. The tximport pipeline was used to import transcript abundance datasets for the differential gene expression analysis for three groups: Control vs. Vadadustat, Control vs. Hypoxia and Vadadustat vs. Hypoxia by DESeq2 software version 1.14.1. Our results provide insight into the transcriptomic effects of these two methods of hypoxic preconditioning of MSCs.

ORGANISM(S): Homo sapiens

PROVIDER: GSE180371 | GEO | 2021/08/04

REPOSITORIES: GEO

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Project description:Hypoxic-ischemic (HI) injury in the developing brain is a common cause of disability in children, and there are no effective treatments at this time. Exposure to sublethal hypoxic conditions (hypoxic preconditioning) 24 hours prior to hypoxic-ischemic insult is protective in the developing rat model. We have observed protective effects on brain histopathology and on long-term sensory-motor behavioral tasks. Changes in gene expression are thought to underlie this protective effect. By comparing gene expression in rats subjected to hypoxic preconditioning or sham conditioning at several time points from 0 to 24 hrs after preconditioning, we should gain insight into the mechanisms underlying these neuroprotective effects and may identify targets for therapeutic intervention. The aim of this study is to determine the effect of hypoxic preconditioning on global gene expression, and, in littermates, to examine the effect of hypoxic preconditioning 24 h prior to hypoxic-ischemic insult on brain histopathology. We hypothesize that changes in gene expression underlie the protective effect of hypoxic preconditioning against subsequent hypoxic-ischemic insult. Gene expression will be examined in two groups, 1) preconditioned and 2) sham controls, at 4 time points. On postnatal day 6, preconditioned animals are exposed to normothermic hypoxia for 3 hrs (8.0% oxygen, 36 degrees C), and sham animals are simultaneouosly exposed to normoxia at 36 degrees C. Animals are then returned to their dams until euthanized at 4 time points (0h, 2h, 8h, and 24h later). Five brains/group/timepoint will be used, with an equal number of males and females in each group. Brains are removed and dissected on ice. Cerebral cortex is dissected from both hemispheres and rapidly frozen on dry ice. Total RNA is isolated using the QIAGEN RNeasy Protect Maxi Kit. Littermates of these animals will be exposed to hypoxic preconditioning or sham preconditioning and subjected to hypoxic-ischemic injury 24 h later. These animals are euthanized at postnatal day 14 for histopathologic evaluation of injury.

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Gene expression profile of human mesenchymal stromal cells (MSCs) exposed to hypoxic and pseudohypoxic preconditioning

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