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Single Cell RNA-sequencing of murine back skin interfollicular epidermis basal cells

ABSTRACT: The ability of the skin to grow in response to stretching has, for decades, been exploited in reconstructive surgery. The response of epidermal cells to stretching has been studied in vitro. However, it remains unclear how mechanical forces affect epidermal cell behaviour in vivo. Here, we develop a mouse model in which the consequences of stretching on skin epidermis can be studied. Using a multidisciplinary approach that combines clonal analysis with quantitative modelling and single-cell RNA-seq, we show that stretching induces skin expansion by a transient bias in the renewal activity of epidermal stem cells, while a second subpopulation of basal progenitors remains committed to differentiation. Transcriptional and chromatin profiling identifies how cell states and gene regulatory networks are modulated by stretching. Using pharmacological inhibitors and mouse mutants, we define the mechanisms that control stretch-mediated tissue expansion.

ORGANISM(S): Mus musculus

PROVIDER: GSE146637 | GEO | 2020/05/18

REPOSITORIES: GEO

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Mechanisms of stretch-mediated skin expansion at single cell resolution [ATAC-seq]

Project description:The ability of the skin to expand in response to stretching has, for decades, been exploited in reconstructive surgery. Several studies have investigated the response of stretching epidermal cells in vitro. However, it remains unclear how mechanical forces affect epidermal stem cell behaviour in vivo. Here, we develop a mouse model in which the temporal consequences of the stretching the skin epidermis can be studied. Using a multidisciplinary approach that combines clonal analysis and mathematical modelling, we show that mechanical force induces skin expansion by promoting the renewal of epidermal stem cells. This occurs through a structured response in which cell fates are coordinated locally by stem cells that switch between states primed for renewal or differentiation. Transcriptional and chromatin profiling identifies the gene regulatory networks modulated by mechanical force. Using a combination of pharmacological inhibition and several conditional gene loss-of-function mouse mutants, we dissect the signalling pathways that control force-mediated tissue expansion.

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Project description:With gene expression profiling it was aimed to identify the differentially expressed genes associated with the regulation of the cytoskeleton to investigate the stretch-induced cell alignment mechanism. A whole genome microarray based analysis of the stretch-induced gene expression changes was done. Gene expression was measured at the beginning of the alignment process showing first reoriented cells after 5 h stretching and at the end after 24 h, where nearly all cells are aligned. Cyclic mechanical stretching of cells results in cellular alignment perpendicular to the stretch direction regulating cellular response. This stress response is assumed to be an adaptation mechanism to reduce extensive stretching but also acts as architectural restructuring changing performance and biomechanics of the tissue. Gene expression profiling of control vs. stretched primary human dermal fibroblasts after 5 h and 24 h demonstrated the regulation of differentially expressed genes associated with metabolism, differentiation and morphology. Primary human dermal fibroblasts from ten donors were cultured on Bioflex culture plates and stretched for 5h and 24 h or left untreated to avoid changes according to cell culturing. Each of the subject provided 4 samples (control/treated and 5hrs/24hrs) resulting in 40 samples total.

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Cyclic mechanical stretch-induced gene expression

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Project description:The renewing human epidermis constantly senses and adapts to a wide range of mechanical cues that are ubiquitous throughout life. The mechanisms of how mechanical forces are responded by interfollicular epidermal stem cells (IFESCs) and are transmitted directly into nucleus to modify gene expression was discovered to be mediated by transcriptional coactivators YAP. Interestingly, YAP, a critical modulator in controlling organ size by regulating both cell proliferation and apoptosis, was also found to induce self-renewal or amplification of epidermal progenitor/stem cells, and balance epidermal growth and differentiation. Because growth regulation imposed by different stimuli is achieved through distinct YAP target genes even in a single cell type. This study used chromatin immunoprecipitation (ChIP) coupled with human promoter tiling microarray analysis (ChIP-on-chip) to profile YAP at promoters genome-wide. The goal of the study was to identify the target genes occupied by YAP in IFESCs under the mechanical stretch condition.

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Interfollicular epidermal differentiation is gradualistic rather than stepwise with GRHL3 controlling progression from stem to transition cell states

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