Inflammatory Memory Sensitizes Skin Epithelial Stem Cells to Tissue Damage

Skin has been utilized as a model system to study many biological processes including wound healing, inflammation, self‑renewal and cancer. Skin is a self-renewal barrier which is maintained by epithelial stem cells (EpSCs) and its main role is to provide a defense against environmental assaults.

In this study Naik et al investigate how skin inflammation affects the EpSCs to allow them to speed up barrier restoration following the subsequent tissue damage.

Topical application of IMQ elicited hyper‑thickening and parakeratosis within epidermis and upper hair follicles, accompanied by increased proliferation and apoptosis of EpSCs. The phenotypic effects subside over time, usually within 30 days (D30) post-inflammation treatment.

EpSCs were lineage tracked for up to 180 days post-inflammation employing a tamoxifen-inducible – marker-based fate mapping system using Rosa‑LSL‑YFP reporter mice. In this assay tamoxifen-induced CreER was driven by the Krt14 (skin epithelial stem cell marker) or Krt10 (skin epithelial differentiation marker). Lineage tracking confirmed that EpSCs were long-lived and were able to survive the inflammatory assault as shown by the persistence of YFP+ cells in the Krt14 promoter driven reporter mice after homeostatic restoration.

Post-inflammation recovered skin was then challenged with secondary assaults, such as wounding. Post-inflamed wounded skin healed more rapidly than skin that was not exposed to inflammation in the first place. This phenomenon was observed even if other inflammatory stimuli were employed, suggesting a quicker response to assaults when already sensitized.

Inflammation-experienced skin displayed accelerated wound healing measured by epidermal thickening and re-epithelization. These effects were not due to increased proliferation or myoblast-mediated dermal contraction, but most likely a result of increased keratinocytes migration as shown in ex vivo explant assay of keratinocyte outgrowth.

Analysis of wounding at sites distal to inflammation-experienced skin demonstrated no enhanced wound healing suggesting that systemic signaling was not involved. The apparent sensitivity to the inflammation stimuli was also not dependent on the memory of the skin's resident innate or adaptive immune cells as they returned to normal levels by D30 post-inflammation. Depletion of skin-resident macrophages prior to wounding also had no obvious impact on the wound repair in inflammation-experienced skin.

Purified populations of EpSCs from IMQ-treated skin tissue at peak response (D6) and after inflammation symptoms have resolved (D30 and D180) were analyzed by assay for Transposase Accessible Chromatin with high-throughput sequencing (ATAC-seq) to identify accessible chromatin regions. IMQ-treated EpSCs had enrichment of peaks associated with genes involved in inflammation such as NF-κB, and STAT1/3.

30 days post-exposure to IMQ, most of the peaks had resolved. There were approximately 2000 peaks that persisted, some as long as 180 days post-treatment, suggestive of epigenetic imprinting. Analysis of peak-associated genes demonstrated enrichment of inflammation- and hyperproliferation-associated pathways, including apoptosis signaling, interleukin signaling, oxidative stress response and PI3 kinase pathways.

To test the relevance of the open chromatin domains post-inflammation in maintaining functional EpSCs memory, the authors evaluated the transcriptional response of EpSCs in accelerated wound repair in inflammation-experienced mice skin. Dramatic transcriptomic changes were observed in IMQ-treated EpSCs relative to controls, which correlated well with genes featuring newly acquired (post IMQ treatment) open chromatin domains. Many of these transcripts had lost expression by day 30 after IMQ, however, 12 hrs post-wounding 52% of the genes upregulated were associated with genes that acquire the open chromatin peaks during and after IMQ treatment.

In addition the authors have identified rapid-response transcripts which serve as effectors of EpSCs memory, including Absent in Melanoma 2 (AIM2) and other downstream components of the AIM2 inflammasome such as Caspase-1 (CASP1) and IL1R1, all specifically required for enhancing of wound repair in inflammation-experienced skin.

In summary the authors present compelling evidence to show that EpSCs maintain memory of tissue assault following initial exposure to inflammation factors and how that affects skin barrier restoration following the subsequent tissue damage by wounding. This functional adaptation of the EpSCs does not require skin-resident macrophages or T cells and that the retention of inflammatory memory by the EpSCs is controlled by chromatin remodeling within those cells which is sustained over long term. The authors speculate that these epigenetic alterations that mobilize stem cells to heal wounds more rapidly are often associated with increased cancer susceptibility. The inflammation-induced rewiring of EpSCs unearthed through this research could have major implications for future therapeutics aimed at autoimmune and hyperproliferative disorders.

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Epistem's Skin Inflammation and Wound Healing Models

At Epistem we can offer a range of in vivo skin models including mouse models of psoriasis induced by imiquimod and a humanized skin graft model (dependent on tissue availability). For in vitro studies we offer growth studies in primary monolayer cultures, ex vivo culture of skin and a living skin equivalent culture for test compound screening. Epistem also provides several in vitro pre-clinical wound healing models including scratch assay, gel contraction assay, high throughput migration assay, 3D migration assay and model for angiogenesis, that are specifically designed to determine the effect of a test item on a specific process involved in the wound healing such as cell migration, cell proliferation or differentiation.

All our models are supported by GCLP‑compliant histology/IHC and genomic analytical services. We have extensive FACS experience and assays available for cytokine analysis on various tissues. We have in-house bioinformatics support for all of our gene expression studies and have extensive experience identifying biomarkers or biomarker signatures in a variety of tissues including skin.

“Inflammatory memory sensitizes skin epithelial stem cells to tissue damage” Naik et al, 2017. Nature, 2017 October 26; 550 (7677): 475-480.doi:10.1038/nature24271

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