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Terminally differentiated keratinocytes are critical for epidermal function and are surrounded by involucrin (IVL). Increased IVL expression is associated with a near-selective sweep in European populations compared with those in Africa. This positive selection for increased IVL in the epidermis identifies human adaptation outside of Africa. The functional significance is unclear. We hypothesize that IVL modulates the environmentally sensitive vitamin D receptor (VDR) in the epidermis. We investigated VDR activity in Ivl ‒/‒ and wild-type mice using vitamin D agonist (MC903) treatment and comprehensively determined the inflammatory response using single-cell RNA sequencing and associated skin microbiome changes using 16S bacterial phylotyping. VDR activity and target gene expression were reduced in Ivl ‒/‒ mouse skin, with decreased MC903-mediated skin inflammation and significant reductions in CD4+ T cells, basophils, macrophages, monocytes, and type II basal keratinocytes and an increase in suprabasal keratinocytes. Coinciding with the dampened MC903-mediated inflammation, the skin microbiota of Ivl ‒/‒ mice was more stable than that of the wild-type mice, which exhibited an MC903-responsive increase in Bacteroidetes and a decrease in Firmicutes. Together, our studies in Ivl ‒/‒ mice identify a functional role for IVL to positively impact VDR activity and suggest an emerging IVL/VDR paradigm for adaptation in the human epidermis., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Squamous cell carcinoma in situ (SCCIS) is a prevalent precancerous lesion that can progress to cutaneous squamous cell carcinoma. Although SCCIS is common, its pathogenesis remains poorly understood. To better understand SCCIS development, we performed laser captured microdissection of human SCCIS and the adjacent epidermis to isolate genomic DNA and RNA for next-generation sequencing. Whole-exome sequencing identified UV-signature mutations in multiple genes, including NOTCH1-3 in the epidermis and SCCIS and oncogenic TP53 mutations in SCCIS. Gene families, including SLFN genes, contained UV/oxidative-signature disruptive epidermal mutations that manifested positive selection in SCCIS. The frequency and distribution of NOTCH and TP53 mutations indicate that NOTCH mutations may precede TP53 mutations. RNA sequencing identified 1,166 differentially expressed genes; the top five enriched gene ontology biological processes included (i) immune response, (ii) epidermal development, (iii) protein phosphorylation, (iv) regulation of catalytic activity, and (v) cytoskeletal regulation. The NEURL1 ubiquitin ligase, which targets Notch ligands for degradation, was upregulated in SCCIS. NEURL1 protein was found to be elevated in SCCIS suggesting that increased levels could represent a mechanism for downregulating Notch during UV-induced carcinogenesis. The data from DNA and RNA sequencing of epidermis and SCCIS provide insights regarding SCCIS formation., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Despite critical functions in cutaneous health and disease, it is unclear how resident skin microbial communities are altered by topical antimicrobial interventions commonly used in personal and clinical settings. Here we show that acute exposure to antiseptic treatments elicits rapid but short-term depletion of microbial community diversity and membership. Thirteen subjects were enrolled in a longitudinal treatment study to analyze the effects of topical treatments (i.e., ethanol, povidone-iodine, chlorhexidine, and water) on the skin microbiome at two skin sites of disparate microenvironment: forearm and back. Treatment effects were highly dependent on personalized and body site-specific colonization signatures, which concealed community dynamics at the population level when not accounted for in this analysis. The magnitude of disruption was influenced by the identity and abundance of particular bacterial inhabitants. Lowly abundant members of the skin microbiota were more likely to be displaced, and subsequently replaced, by the most abundant taxa prior to treatment. Members of the skin commensal family Propionibactericeae were particularly resilient to treatment, suggesting a distinct competitive advantage in the face of disturbance. These results provide insight into the stability and resilience of the skin microbiome, while establishing the impact of topical antiseptic treatment on skin bacterial dynamics and community ecology., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Identifying previously unknown proteins or detecting the presence of known proteins in research samples is critical to many experiments conducted in life sciences, including dermatology. Sensitive protein detection can help elucidate new intervention targets and mechanisms of disease, such as in autoimmune blistering skin diseases, atopic eczema, or other conditions. Historically, peptides from highly purified single proteins were sequenced, with many limitations, by stepwise degradation from the N-terminus to the C-terminus with subsequent identification by UV absorbance spectroscopy of the released amino acids (i.e., Edman degradation). Recently, however, the availability of comprehensive protein databases from different species (derived from high-throughput next-generation sequencing of those organisms' genomes) and sophisticated bioinformatics analysis tools have facilitated the development and use of mass spectrometry for identification and global analysis of proteins, summarized as mass spectrometry-based proteomics. Mass spectrometry is an analytical technique measuring the mass (m)-to-charge (z) ratio of ionized biological molecules such as peptides. Proteins can be identified by correlating peptide-derived experimental mass spectrometry spectra with theoretical spectra predicted from protein databases. Here we briefly describe how this technique works, how it can be used for identification of proteins, and how this knowledge can be applied in elucidating human biology and disease., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Microbial burden of chronic wounds is believed to play an important role in impaired healing and the development of infection-related complications. However, clinical cultures have little predictive value of wound outcomes, and culture-independent studies have been limited by cross-sectional design and small cohort size. We systematically evaluated the temporal dynamics of the microbiota colonizing diabetic foot ulcers, a common and costly complication of diabetes, and its association with healing and clinical complications. Dirichlet multinomial mixture modeling, Markov chain analysis, and mixed-effect models were used to investigate shifts in the microbiota over time and their associations with healing. Here we show, to our knowledge, previously unreported temporal dynamics of the chronic wound microbiome. Microbiota community instability was associated with faster healing and improved outcomes. Diabetic foot ulcer microbiota were found to exist in one of four community types that experienced frequent and nonrandom transitions. Transition patterns and frequencies were associated with healing time. Exposure to systemic antibiotics destabilized the wound microbiota, rather than altering overall diversity or relative abundance of specific taxa. This study provides evidence that the dynamic wound microbiome is indicative of clinical outcomes and may be a valuable guide for personalized management and treatment of chronic wounds., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Culture-independent studies to characterize skin microbiota are increasingly common, due in part to affordable and accessible sequencing and analysis platforms. Compared to culture-based techniques, DNA sequencing of the bacterial 16S ribosomal RNA (rRNA) gene or whole metagenome shotgun (WMS) sequencing provides more precise microbial community characterizations. Most widely used protocols were developed to characterize microbiota of other habitats (i.e., gastrointestinal) and have not been systematically compared for their utility in skin microbiome surveys. Here we establish a resource for the cutaneous research community to guide experimental design in characterizing skin microbiota. We compare two widely sequenced regions of the 16S rRNA gene to WMS sequencing for recapitulating skin microbiome community composition, diversity, and genetic functional enrichment. We show that WMS sequencing most accurately recapitulates microbial communities, but sequencing of hypervariable regions 1-3 of the 16S rRNA gene provides highly similar results. Sequencing of hypervariable region 4 poorly captures skin commensal microbiota, especially Propionibacterium. WMS sequencing, which is resource and cost intensive, provides evidence of a community's functional potential; however, metagenome predictions based on 16S rRNA sequence tags closely approximate WMS genetic functional profiles. This study highlights the importance of experimental design for downstream results in skin microbiome surveys., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Extracellular matrix proteins play a critical role in dermal wound healing by mediating matrix-cell interactions and re-establishing the dermal architecture and environment. Fibulin-5 is an elastin-binding protein essential for elastic fiber development in vivo, and it has recently been shown to inhibit angiogenesis in vitro. Here, we use mice deficient for the fibulin-5 gene (fbln5) to examine the role of fibulin-5 and the effect of the loss of elastic fibers in dermal wound healing. Fbln5 is upregulated in the granulation tissue 14 days after full-thickness wounding in wild-type mice, before the formation of elastic fibers. Although wounded fbln5(-/-) skin showed enhanced neovascularization compared to the wild-type skin, no difference in the rate of wound closure was observed between mutant and wild-type mice. In addition, a breaking strength test revealed that there was no difference in breaking stress or strain between wild-type and fbln5(-/-) wounded skin. These results suggest that fibulin-5 and elastic fibers are not directly involved in short-term wound healing. Clearly, the long-term effect of the absence of fibulin-5 on the function and integrity of regenerated skin needs to be further addressed.