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Although not completely understood, the role of the Hedgehog-GLI (HH-GLI) signaling pathway in melanoma and epithelial skin tumors has been reported before. In this study, we confirmed in various melanoma cell line models that keratin 16 (KRT16) and S100 Calcium-Binding Protein A7 (S100A7) are transcriptional targets of GLI Family Zinc Finger (GLI) proteins. Besides their important role in protecting and maintaining the epidermal barrier, keratins are somehow tightly connected with the S100 family of proteins. We found that stronger expression of KRT16 indeed corresponds to stronger expression of S100A7 in our clinical melanoma samples. We also report a trend regarding staining of GLI1, which corresponds to stronger staining of GLI3, KRT16, and S100A7 proteins. The most interesting of our findings is that all the proteins are detected specifically in the epidermis overlying the tumor, but rarely in the tumor itself. The examined proteins were also not detected in the healthy epidermis at the edges of the sample, suggesting that the staining is specific to the epidermis overlaying the tumor mass. Of all proteins, only S100A7 demonstrated a statistically significant trend regarding tumor staging and staining intensity. Results from our clinical samples prove that immune infiltration is an important feature of melanoma. Pigmentophages and tumor-infiltrating lymphocytes (TIL) demonstrate a significant association with tumor stage, while mononuclear cells are equally present in all stages. For S100A7, we found an association between the number of TILs and staining intensity. Considering these new findings presented in our study, we suggest a more detailed examination of the possible role of the S100A7 protein as a biomarker in melanoma.

Accumulating evidence indicates that microRNAs (miRNAs) have a vital effect on the pathogenesis of psoriasis. This study is conducted to investigate the potential involvement of miR-181a-5p and miR-181b-5p in the proliferation of HaCaT keratinocytes. Cell viability and proliferation were evaluated respectively in this study using the CCK-8 and the 5-ethynyl-2'-deoxyuridine (EdU) assays. The expression of Maternal Embryonic Leucine Zipper Kinase (MELK) and Keratin 16 (KRT16) mRNA and protein in tissues and cells was assessed using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The Luciferase reporter system analyzes the connection between miR-181a-5p/miR-181b-5p and MELK. The results showed that miR-181a/b-5p expression was downregulated in the psoriasis lesions and negatively regulated the proliferation of keratinocytes. MELK was directly targeted by miR-181a-5p/miR-181b-5p. In addition, HaCaT keratinocytes proliferation was inhibited by knockdown of MELK while promoted dramatically by MELK overexpression. Notably, miR-181a/b-5p mimics could attenuate the effects of MELK in keratinocytes. In conclusion, our research findings suggested miR-181a-5p and miR-181b-5p negatively regulate keratinocyte proliferation by targeting MELK, providing potential diagnostic biomarkers and therapeutic targets for psoriasis., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Psoriasis and chronic ulcers not only significantly impair quality of life but also pose a challenge in dermatological treatment. This study aimed to identify new therapeutic targets and biomarkers for psoriasis and chronic ulcers by comparing their gene expression profiles. The gene expression profiles of psoriatic, wound and chronic ulcer patients, as well as healthy controls, were determined via RNA extraction and next-generation sequencing of biopsies. In order to identify biomarkers, functional enrichment, differential expression analysis and machine learning algorithms were implemented. It is worth mentioning that the genes IL17A, TNF, KRT16, MMP9, and CD44 exhibited substantial correlations with the pathogenesis of the conditions being studied. As evidenced by their AUC-ROC values approaching 0.90, machine learning models accurately identified these biomarkers. The differential gene expression was consequently validated via qRT-PCR, which highlighted the increased expression of matrix remodelling enzymes and inflammatory cytokines. Additionally, genes essential for maintaining epidermis integrity and facilitating wound healing exhibited downregulation. These insights into the molecular mechanisms of psoriasis and chronic ulcers pave the way for the development of targeted therapies, offering hope for improved treatment strategies., (© 2023 The Authors. International Wound Journal published by Medicalhelplines.com Inc and John Wiley & Sons Ltd.)

Competing Interests: Conflicts of interest Professor O’Toole has received funding for the university from Palvella Therapeutics (consultancy), Timber Pharma (consultancy), and Kamari Pharma (research grant and consultancy). Dr Hansen has received support from Palvella as a principal investigator for a clinical study in pachyonychia. Drs Tan, Steele and Thomas and Ms Schwartz have no conflicts of interest to declare.

Nasopharyngeal carcinoma (NPC) refers to a malignancy initiating from the superior mucosal epithelium of the nasopharynx. Optimal therapies for NPC are still needed. In this investigation, we attempted to explore whether BarH-like homeobox 2 (BARX2), a well-known tumor suppressor, had anti-cancer properties on NPC, and the possible mechanisms. After searching for NPC-related databases, we determined BARX2 as one of the core genes in NPC. The results of RT-qPCR and immunohistochemistry or Western blot demonstrated that BARX2 was reduced in NPC patients and cells. Ectopic expression of BARX2 reverted the malignant phenotype of NPC cells. Mechanistically, BARX2 bound to the keratin 16 (KRT16) promoter to downregulate its expression. In addition, BARX2 was found to reduce the phosphorylation levels of MEK and ERK. Further KRT16 upregulation in cells overexpressing BARX2 promoted malignant aggressiveness of C666-1 and HNE3 cells and activated the Ras signaling pathway. BARX2 inhibited the growth and metastasis of tumors and suppressed the Ras signaling pathway in vivo . In conclusion, our findings indicate that BARX2 reverts malignant phenotypes of NPC cells by downregulating KRT16 in a Ras-dependent fashion. BARX2 might act as a possible therapeutic regulator for NPC.

Pachyonychia congenita (PC) is a genetic disorder of keratin that presents with nail dystrophy, painful palmoplantar keratoderma, and other clinical manifestations. We investigated the genotype‒structurotype‒phenotype correlations seen with mutations in keratin genes (keratin [K]6A, K6B, K6C, K16, K17) and utilized protein structure modeling of high-frequency mutations to examine the functional importance of keratin structural domains in PC pathogenesis. Participants of the International PC Research Registry underwent genetic testing and completed a standardized survey on their symptoms. Our results support previous reports associating oral leukokeratosis with K6A mutations and cutaneous cysts, follicular hyperkeratosis, and natal teeth with K17 mutations. Painful keratoderma was prominent with K6A and K16 mutations. Nail involvement was most common in patients with K6A mutation and least common in those with K6C mutation. Across keratin subtypes, patients with coil 2B mutations had the greatest impairment in ambulation, and patients with coil 1A mutations reported more emotional issues. Molecular modeling demonstrated that hotspot missense mutations in PC largely disrupted hydrophobic interactions or surface charge. The former may destabilize keratin dimers/tetramers, whereas the latter likely interferes with higher-order keratin filament formation. Understanding the pathologic alterations in keratin structure improves our knowledge of how PC genotype correlates with clinical phenotype, advancing insight into disease pathogenesis and therapeutic development., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Psoriasis is a chronic, inflammatory skin disease. It was reported that lncRNA Non-coding RNA-activated by DNA damage (NORAD) has potential regulatory effects on skin diseases. Our previous studies found that lncRNA NORAD was highly expressed and its potential target miR-26a was down-regulated in psoriasis model mice. Here, we aimed to investigate the role of NORAD in the development of psoriasis., Methods: IL-22/LPS (interleukin-22/lipopolysaccharide)-stimulated HaCaT (human immortalized keratinocytes) cell model and imiquimod-induced mouse model were established. Keratin 6 (K6), Keratin 16 (K16), Keratin 17 (K17), and Cell division cycle 6 (CDC6) levels were detected by western blot. Cell activity was detected by CCK-8, MTT, and EdU assays. Quantitative real-time PCR was performed to examine the levels of NORAD, miR-26a, CDC6, K6, K16, and K17. Haematoxylin-eosin staining was applied to observe the degree of skin thickening and hyperplasia. Fluorescence in situ hybridization detects the location of NORAD. RNA immunoprecipitation, RNA pull-down, and Luciferase test were performed to detect the interaction between NORAD and miR-26a., Results: In IL-22/LPS-stimulated HaCaT cells, NORAD, CDC6, and keratinocyte proliferation-related proteins (K6, K16, and K17) were up-regulated and miR-26a was down-regulated. Cell survival and proliferation were also increased. However, the results were reversed after interference with NORAD. Also, in vitro experiments revealed that NORAD negatively regulated miR-26a. In IL-22/LPS-stimulated HaCaT cells and skin of imiquimod-induced mice, we found that lower NORAD resulted in an increase of miR-26a and a decrease of CDC6, further decreased levels of keratinocyte proliferation-related proteins (K6, K16, and K17).

Mitochondria fulfill essential roles in ATP production, metabolic regulation, calcium signaling, generation of reactive oxygen species (ROS), and additional determinants of cellular health. Recent studies have highlighted a role for mitochondria during cell differentiation, including in skin epidermis. The observation of oxidative stress in keratinocytes from Krt16 null mouse skin, a model for pachyonychia congenita (PC)-associated palmoplantar keratoderma, prompted us to examine the role of Keratin (K) 16 protein and its partner K6 in regulating the structure and function of mitochondria. Electron microscopy revealed major anomalies in mitochondrial ultrastructure in late stage, E18.5, Krt6a/Krt6b null embryonic mouse skin. Follow-up studies utilizing biochemical, metabolic, and live imaging readouts showed that, relative to controls, skin keratinocytes null for Krt6a/Krt6b or Krt16 exhibit elevated ROS, reduced mitochondrial respiration, intracellular distribution differences, and altered movement of mitochondria within the cell. These findings highlight a novel role for K6 and K16 in regulating mitochondrial morphology, dynamics, and function and shed new light on the causes of oxidative stress observed in PC and related keratin-based skin disorders.

Background: Pachyonychia congenita (PC), a rare genodermatosis, primarily affects ectoderm-derived epithelial appendages and typically includes oral leukokeratosis, nail dystrophy and very painful palmoplantar keratoderma (PPK). PC dramatically impacts quality of life although it does not affect lifespan. PC can arise from mutations in any of the wound-repair-associated keratin genes KRT6A, KRT6B, KRT6C, KRT16 or KRT17. There is no cure for this condition, and current treatment options for PC symptoms are limited and palliative in nature., Objectives: This review focuses on recent progress made towards understanding the pathophysiology of PPK lesions, the most prevalent and debilitating of all PC symptoms., Methods: We reviewed the relevant literature with a particular focus on the Krt16 null mouse, which spontaneously develops footpad lesions that mimic several aspects of PC-associated PPK., Results: There are three main stages of progression of PPK-like lesions in Krt16 null mice. Ahead of lesion onset, keratinocytes in the palmoplantar (footpad) skin exhibit specific defects in terminal differentiation, including loss of Krt9 expression. At the time of PPK onset, there is elevated oxidative stress and hypoactive Keap1-Nrf2 signalling. During active PPK, there is a profound defect in the ability of the epidermis to maintain or return to normal homeostasis., Conclusions: The progress made suggests new avenues to explore for the treatment of PC-based PPK and deepens our understanding of the mechanisms controlling skin tissue homeostasis. What's already known about this topic? Pachyonychia congenita (PC) is a rare genodermatosis caused by mutations in KRT6A, KRT6B, KRT6C, KRT16 and KRT17, which are normally expressed in skin appendages and induced following injury. Individuals with PC present with multiple clinical symptoms that usually include thickened and dystrophic nails, palmoplantar keratoderma (PPK), glandular cysts and oral leukokeratosis. The study of PC pathophysiology is made challenging because of its low incidence and high complexity. There is no cure or effective treatment for PC. What does this study add? This text reviews recent progress made when studying the pathophysiology of PPK associated with PC. This recent progress points to new possibilities for devising effective therapeutics that may complement current palliative strategies., (© 2019 British Association of Dermatologists.)

Background: Pachyonychia congenita (PC), a rare autosomal dominant disorder, is featured by significant hypertrophic nail, palmoplantar keratoderma, and plantar pain. It is caused by the mutation of KRT6A, KRT6B, KRT6C, KRT16, or KRT17., Aims: To identify the gene mutation caused the PC in a Chinese family., Patients/methods: Genomic DNA was extracted from peripheral blood samples of five patients and six healthy individuals. Genomic DNA of three patients was sequenced by whole-exome sequencing (WES). Then, exons 6 of KRT16 of all samples were amplified by polymerase chain reaction (PCR), and PCR products were sequenced to identify potential mutations., Results: We identified the proline substitution mutation p.Leu421Pro (c.1262T>C) in the 2B domain of K16 that is associated with PC in a Chinese family. The same mutation was not found in the six healthy individuals of the family., Conclusions: The mutation found in this study is the first report in China. So far, 25 mutations in KRT16 have been reportedly associated with PC. Twenty-one mutations are located on exon 1, and four mutations on exon 6., (© 2019 Wiley Periodicals, Inc.)

The type I intermediate filament keratin 16 (KRT16 gene; K16 protein) is constitutively expressed in ectoderm-derived appendages and in palmar/plantar epidermis and is robustly induced when the epidermis experiences chemical, mechanical or environmental stress. Missense mutations at the KRT16 locus can cause pachyonychia congenita (PC, OMIM:167200) or focal non-epidermolytic palmoplantar keratoderma (FNEPPK, OMIM:613000), which each entail painful calluses on palmar and plantar skin. Krt16-null mice develop footpad lesions that mimic PC-associated PPK, providing an opportunity to decipher its pathophysiology, and develop therapies. We report on insight gained from a genome-wide analysis of gene expression in PPK-like lesions of Krt16-null mice. Comparison of this data set with publicly available microarray data of PPK lesions from individuals with PC revealed significant synergies in gene expression profiles. Keratin 9 (Krt9/K9), the most robustly expressed gene in differentiating volar keratinocytes, is markedly downregulated in Krt16-null paw skin, well-ahead of lesion onset, and is paralleled by pleiotropic defects in terminal differentiation. Effective prevention of PPK-like lesions in Krt16-null paw skin (via topical delivery of the Nrf2 inducer sulforaphane) involves the stimulation of Krt9 expression. These findings highlight a role for defective terminal differentiation and loss of Krt9/K9 expression as additional drivers of PC-associated PPK and highlight restoration of KRT9 expression as a worthy target for therapy. Further, we report on the novel observation that keratin 16 can localize to the nucleus of epithelial cells, implying a potential nuclear function that may be relevant to PC and FNEPPK., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Psoriasis is a multisystem disease affecting about 2% of the population, while keratin16 (KRT16) has been reported to participate in psoriasis. However, the specific mechanism of KRT16 in psoriasis was inadequately investigated. The objective of the study was to elucidate the mechanism by which siRNA-mediated silencing of KRT16 affects keratinocyte proliferation and vascular endothelial growth factor (VEGF) secretion in psoriasis through the extracellular signal-related kinase (ERK) signaling pathway. Psoriasis-related core gene KRT16 was screened out. Then, the expression of KRT16, VEGF, and ERK signaling pathway-related genes was detected in psoriatic patients. To further investigate the mechanism of KRT16, keratinocytes in psoriatic patients were treated with KRT16 siRNA or/and ERK inhibitor (PD98059) to detect the changes in related gene expression and cell survival. KRT16 was involved in psoriasis development. The expression levels of KRT16, p-ERK1/2, and VEGF in lesion tissues are significantly elevated. Keratinocytes treated with KRT16-siRNA and KRT16-siRNA + PD98059 exhibited reduced KRT16, p-ERK1/2, and VEGF expression. The cell survival rate in cells treated with KRT16-siRNA, PD98059, and KRT16-siRNA + PD98059 reduced significantly. These findings indicate that silencing KRT16 inhibits keratinocyte proliferation and VEGF secretion in psoriasis via inhibition of ERK signaling pathway, which provides a basic theory in the treatment of psoriasis., (© 2019 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia on behalf of Kaohsiung Medical University.)

Background: Targeting the c-Met signaling pathway has become a therapeutic strategy in multiple types of cancer. We unveiled a novel c-Met regulating mechanism that could be applied as a modality for oral squamous cell carcinoma (OSCC) therapy., Methods: Upregulation of keratin 16 (KRT16) was found by comparing isogenic pairs of low and high invasive human OSCC lines via microarray analysis. OSCC cells with ectopic expression or silencing of KRT16 were used to scrutinize functional roles and associated molecular mechanisms., Results: We observed that high KRT16 expression significantly correlated with poorer pathological differentiation, advanced stages, increased lymph nodes metastasis, and decreased survival rate from several Taiwanese OSCC patient cohorts. We further revealed that miR-365-3p could target ETS homologous factor (EHF), a KRT16 transcription factor, to decrease migration, invasion, metastasis and chemoresistance in OSCC cells via inhibition of KRT16. Under confocal microscopic examination, c-Met was found possibly partially associates with KRT16 through β5-integrin. Colocalization of these three proteins may facilitate c-Met and β5-integrin-mediated signaling in OSCC cells. Depletion of KRT16 led to increased protein degradation of β5-integrin and c-Met through a lysosomal pathway leading to inhibition of their downstream Src/STAT3/FAK/ERK signaling in OSCC cells. Knockdown of KRT16 enhanced chemosensitivity of OSCC towards 5-fluorouracil (5-FU). Various combination of c-Met inhibitor (foretinib), protein tyrosine kinase inhibitor (genistein), β5-integrin antibody, and 5-FU markedly augmented cytotoxic effects in OSCC cells as well as tumor killing effects in vitro and in vivo., Conclusions: Our data indicate that targeting a novel miR-365-3p/EHF/KRT16/β5-integrin/c-Met signaling pathway could improve treatment efficacy in OSCC.

Gallic acid (GA), a natural small molecule found in Radix Paeoniae Rubra, has a variety of favorable biological activities. However, the anti-psoriasis effect of GA has never been explored up to now. This study evaluates the protective effect of GA on psoriasis-like skin disease both in vitro and in vivo and explores the underlying mechanism. The results show that GA significantly decreases the mRNA and protein expression of keratin 16 and keratin 17 which are the markers of psoriasis. Additionally, GA obviously ameliorates psoriasis area and severity index scores and decreases the epidermal hyperplasia of psoriasis-like disease mice. The activity of Nrf2 which targets keratin 16 and keratin 17 is significantly downregulated by GA. Furthermore, the downregulation of keratin 16 and keratin 17 induced by GA was abolished by the Nrf2-overexpression in vitro. This study initially elucidates the anti-psoriasis effect and mechanism of GA which hints that GA would be a potential candidate for the treatment of psoriasis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Skin wound healing is a dynamic and complex process involving several mediators at the cellular and molecular levels. Lupeol, a phytoconstituent belonging to the triterpenes class, is found in several fruit plants and medicinal plants that have been the object of study in the treatment of various diseases, including skin wounds. Various medicinal properties of lupeol have been reported in the literature, including anti-inflammatory, antioxidant, anti-diabetic, and anti-mutagenic effects. We investigated the effects of lupeol (0.1, 1, 10, and 20 μg/mL) on in vitro wound healing assays and signaling mechanisms in human neonatal foreskin keratinocytes and fibroblasts. Results showed that, at high concentrations, Lupeol reduced cell proliferation of both keratinocytes and fibroblasts, but increased in vitro wound healing in keratinocytes and promoted the contraction of dermal fibroblasts in the collagen gel matrix. This triterpene positively regulated matrix metalloproteinase (MMP)-2 and inhibited the NF-κB expression in keratinocytes, suggesting an anti-inflammatory effect. Lupeol also modulated the expression of keratin 16 according to the concentration tested. Additionally, in keratinocytes, lupeol treatment resulted in the activation of Akt, p38, and Tie-2, which are signaling proteins involved in cell proliferation and migration, angiogenesis, and tissue repair. These findings suggest that lupeol has therapeutic potential for accelerating wound healing.

Background: Liver X receptors (LXRs) are involved in maintaining epidermal barrier and suppressing inflammatory responses in model systems. The LXR agonist VTP-38543 showed promising results in improving barrier function and inflammatory responses in model systems., Objective: To assess the safety, tolerability, cellular and molecular changes, and clinical efficacy of the topical VTP-38543 in adults with mild to moderate atopic dermatitis (AD)., Methods: A total of 104 ambulatory patients with mild to moderate AD were enrolled in this randomized, double-blind, vehicle-controlled trial between December 2015 and September 2016. VTP-38543 cream in 3 concentrations (0.05%, 0.15%, and 1.0%) or placebo was applied twice daily for 28 days. Pretreatment and posttreatment skin biopsy specimens were obtained from a subset of 33 patients. Changes in SCORing of Atopic Dermatitis, Eczema Area and Severity Index, Investigator's Global Assessment, and tissue biomarkers (by real-time polymerase chain reaction and immunostaining) were evaluated., Results: Topical VTP-38543 was safe and well tolerated. VTP-38543 significantly increased messenger RNA (mRNA) expression of epidermal barrier differentiation (loricrin and filaggrin, P = .02) and lipid (adenosine triphosphate-binding cassette subfamily G member 1 and sterol regulatory element binding protein 1c, P < .01) measures and reduced epidermal hyperplasia markers (thickness, keratin 16 mRNA). VTP-38543 nonsignificantly suppressed cellular infiltrates and down-regulated mRNA expression of several T H 17/T H 22-related (phosphatidylinositol 3, S100 calcium-binding protein A12) and innate immunity (interleukin 6) markers., Conclusion: Topical VTP-38543 is safe and well tolerated. Its application led to improvement in barrier differentiation and lipids. Longer-term studies are needed to clarify whether a barrier-based approach can induce meaningful suppression of immune abnormalities., Trial Registration: clinicaltrials.gov Identifier: NCT02655679., (Copyright © 2018 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

The ATF4 transcription factor is a key regulator of the adaptive integrated stress response (ISR) induced by various stresses and pathologies. Identification of novel transcription targets of ATF4 during ISR would contribute to the understanding of adaptive networks and help to identify novel therapeutic targets. We were previously searching for genes that display an inverse regulation mode by the transcription factors ATF4 and p53 in response to mitochondrial respiration chain complex III inhibition. Among the selected candidates the human genes for cytokeratine 16 (KRT16), anti-apoptotic protein Niban (FAM129A) and hexokinase HKDC1 have been found highly responsive to ATF4 overexpression. Here we explored potential roles of the induction of KRT16, FAM129A and HKDC1 genes in ISR. As verified by RT-qPCR, a dysfunction of mitochondrial respiration chain and ER stress resulted in a partially ATF4-dependent stimulation of KRT16, FAM129A and HKDC1 expression in the HCT116 colon carcinoma cell line. ISRIB, a specific inhibitor of ISR, was able to downregulate the ER stress-induced levels of KRT16, FAM129A and HKDC1 transcripts. An inhibition of ATF4 by RNAi attenuated the induction of KRT16, FAM129A and HKDC1 mRNAs in response to ER stress or to a dysfunctional mitochondrial respiration. The similar induction of the three genes was observed in another tumor-derived cervical carcinoma cell line HeLa. However, in HaCaT and HEK293T cells that display transformed phenotypes, but do not originate from patient-derived tumors, the ER stress-inducing treatments resulted in an upregulation of FAM129A and HKDC1, but not KRT16 transcripts, By a luciferase reporter approach we identified a highly active ATF4-responsive element within the upstream region of the KRT16 gene. The results suggest a conditional regulation of KRT16 gene by ATF4 that may be inhibited in normal cells, but engaged during cancer progression. Potential roles of KRT16, FAM129A and HKDC1 genes upregulation in adaptive stress responses and pathologies are discussed.

Psoriasis is a chronic inflammatory skin disease characterized by keratinocyte hyperproliferation of epidermis. Although hyperproliferation-associated keratins K6, K16, and K17 are considered to be the hallmarks of psoriasis, the molecular basis underlying the overexpression of these keratins remains unclear. Nrf2 regulates cell proliferation. Therefore, we investigated whether Nrf2 regulates keratinocyte proliferation via promoting expression of K6, K16, and K17 in psoriasis. We initially found that psoriatic epidermis exhibited elevated expression of Nrf2. Furthermore, Nrf2 promoted expression of K6, K16, and K17 in both HaCaT cells and primary human keratinocytes by binding to the ARE domains located in the promoter of these genes. Additionally, upon stimulation with IL-17 or IL-22, Nrf2 translocated to the nucleus and initiated expression of targeted keratins. In mice of imiquimod-induced psoriasis-like dermatitis, topical application of Nrf2 small interfering RNA alleviated the epidermal hyperplasia with reduced expression of these keratins. More importantly, Nrf2 promoted the proliferation of human keratinocytes through up-regulation of K6, K16, or K17. These data suggested that inflammatory cytokines promoted Nrf2 nuclear translocation in psoriatic epidermis, which led to elevated expression of K6, K16, and K17, thus promoting keratinocyte proliferation and contributing to the pathogenesis of psoriasis., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Epidermolysis bullosa simplex is a skin-blistering disorder caused by mutations in keratin (K)14 or K5. Treatment with nuclear factor (erythroid-derived 2)-like 2 inducer sulforaphane ameliorated skin blistering in Krt14-null mice, correlating with induction of K17. To be therapeutically useful for epidermolysis bullosa simplex, topical broccoli sprout extract (BSE), enriched for sulforaphane, would ideally induce the expression of homologous keratins (eg, K6, K17, K16) in the basal layer of human epidermis without impacting expression of defective keratins (K5/K14)., Objective: The purpose of this 1-week, randomized, split-body, single-blinded, placebo-controlled trial was to assess the impact of BSE on keratin expression., Methods: Five subjects (34-71 years old) applied BSE (500 nmol of sulforaphane/mL) or vehicle alone to the inner aspect of the arm daily. Expression of keratin, nuclear factor (erythroid-derived 2)-like 2, and other markers was assessed using reverse transcription-polymerase chain reaction and indirect immunofluorescence., Results: One subject (age 71 years) was excluded a posteriori because of poor tissue quality. Topical BSE activated nuclear factor (erythroid-derived 2)-like 2 and up-regulated K17 in the epidermis of all subjects, had variable effects on K16 and K6 expression, and did not alter expression of K14 or K5., Limitations: Small sample size is a limitation., Conclusion: BSE represents an attractive therapeutic candidate for K14-associated epidermolysis bullosa simplex., (Copyright © 2016 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.)

Palmoplantar keratoderma (PPK) are debilitating lesions that arise in individuals with pachyonychia congenita (PC) and feature upregulation of danger-associated molecular patterns and skin barrier regulators. The defining features of PC-associated PPK are reproduced in mice null for keratin 16 (Krt16), which is commonly mutated in PC patients. Here, we have shown that PPK onset is preceded by oxidative stress in footpad skin of Krt16-/- mice and correlates with an inability of keratinocytes to sustain nuclear factor erythroid-derived 2 related factor 2-dependent (NRF2-dependent) synthesis of the cellular antioxidant glutathione (GSH). Additionally, examination of plantar skin biopsies from individuals with PC confirmed the presence of high levels of hypophosphorylated NRF2 in lesional tissue. In Krt16-/- mice, genetic ablation of Nrf2 worsened spontaneous skin lesions and accelerated PPK development in footpad skin. Hypoactivity of NRF2 in Krt16-/- footpad skin correlated with decreased levels or activity of upstream NRF2 activators, including PKCδ, receptor for activated C kinase 1 (RACK1), and p21. Topical application of the NRF2 activator sulforaphane to the footpad of Krt16-/- mice prevented the development of PPK and normalized redox balance via regeneration of GSH from existing cellular pools. Together, these findings point to oxidative stress and dysfunctional NRF2 as contributors to PPK pathogenesis, identify K16 as a regulator of NRF2 activation, and suggest that pharmacological activation of NRF2 should be further explored for PC treatment.

Transcription factors are key molecules that finely tune gene expression in response to injury. We focused on the role of a transcription factor, Foxn1, whose expression is limited to the skin and thymus epithelium. Our previous studies showed that Foxn1 inactivity in nude mice creates a pro-regenerative environment during skin wound healing. To explore the mechanistic role of Foxn1 in the skin wound healing process, we analyzed post-injured skin tissues from Foxn1::Egfp transgenic and C57BL/6 mice with Western Blotting, qRT-PCR, immunofluorescence and flow cytometric assays. Foxn1 expression in non-injured skin localized to the epidermis and hair follicles. Post-injured skin tissues showed an intense Foxn1-eGFP signal at the wound margin and in leading epithelial tongue, where it co-localized with keratin 16, a marker of activated keratinocytes. This data support the concept that suprabasal keratinocytes, expressing Foxn1, are key cells in the process of re-epithelialization. The occurrence of an epithelial-mesenchymal transition (EMT) was confirmed by high levels of Snail1 and Mmp-9 expression as well as through co-localization of vimentin/E-cadherin-positive cells in dermis tissue at four days post-wounding. Involvement of Foxn1 in the EMT process was verified by co-localization of Foxn1-eGFP cells with Snail1 in histological sections. Flow cytometric analysis showed the increase of double positive E-cadherin/N-cadherin cells within Foxn1-eGFP population of post-wounded skin cells isolates, which corroborated histological and gene expression analyses. Together, our findings indicate that Foxn1 acts as regulator of the skin wound healing process through engagement in re-epithelization and possible involvement in scar formation due to Foxn1 activity during the EMT process.

Background: K1 and K2 are the main type II keratins in the suprabasal epidermis where each of them heterodimerizes with the type I keratin K10 to form intermediate filaments. In regions of the ears, tail, and soles of the mouse, only K2 is co-expressed with K10, suggesting that these keratins suffice to form a mechanically resilient cytoskeleton., Objective: To determine the effects of the suppression of both main keratins, K2 and K10, in the suprabasal plantar epidermis of the mouse., Methods: Krt2(-/-) Krt10(-/-) mice were generated by crossing Krt2(-/-) and Krt10(-/-) mice. Epidermal morphology of soles of hind-paws was examined macroscopically and histologically. Immunofluorescence analysis and quantitative PCR analysis were performed to analyze the expression of keratins in sole skin of wildtype and Krt2(-/-) Krt10(-/-) mice. Highly abundant proteins of the sole stratum corneum were determined by electrophoretic and chromatographic separation and subsequent mass spectrometry., Results: K2 and K10 are the most prominent suprabasal keratins in normal mouse soles with the exception of the footpads where K1, K9 and K10 predominate. Mice lacking both K2 and K10 were viable and developed epidermal acanthosis and hyperkeratosis in inter-footpad epidermis of the soles. The expression of keratins K1, K9 and K16 was massively increased at the RNA and protein levels in the soles of Krt2(-/-) Krt10(-/-) mice., Conclusions: This study demonstrates that the loss of the main cytoskeletal components of plantar epidermis, i.e. K2 and K10, can be only partly compensated by the upregulation of other keratins. The thickening of the epidermis in the soles of Krt2(-/-) Krt10(-/-) mice may serve as a model for pathomechanistic aspects of palmoplantar keratoderma., (Copyright © 2015. Published by Elsevier Ireland Ltd.)

Background: Impaired wound healing is a complication of diabetes and a serious problem in clinical practice. We previously found that whey protein (WP) was able to regulate wound healing normally in streptozotocin (STZ)-diabetic models. This subsequent study was designed to assess the effect of WP on heat shock protein-72 (Hsp72) and keratin16 (Krt16) expression during wound healing in diabetic rats., Methods: WP at a dosage of 100 mg/kg of body weight was orally administered daily to wounded normal and STZ-diabetic rats for 8 days., Results: At day 4, the WP-treated diabetic wound was significantly reduced compared to that in the corresponding control. Diabetic wounded rats developed severe inflammatory infiltration and moderate capillary dilatation and regeneration. Treated rats had mild necrotic formation, moderate infiltration, moderate to severe capillary dilatation and regeneration, in addition to moderate epidermal formation. Hsp72 and Krt16 densities showed low and dense activity in diabetic wounded and diabetic wounded treated groups, respectively. At day 8, WP-treatment of diabetic wounded animals revealed great amelioration with complete recovery and closure of the wound. Reactivity of Hsp72 and Krt16 was reversed, showing dense and low, or medium and low, activity in the diabetic wounded and diabetic wounded treated groups, respectively. Hsp72 expression in the pancreas was found to show dense reactivity with WP-treated diabetic wound rats., Conclusion: This data provides evidence for the potential impact of WP in the up-regulation of Hsp72 and Krt16 in T1D, resulting in an improved wound healing process in diabetic models.

Background: Pachyonychia congenita (PC) is a skin disorder resulting from mutations in keratin (K) proteins including K6a, K6b, K16, and K17. One of the major symptoms is painful plantar keratoderma. The pathogenic sequelae resulting from the keratin mutations remain unclear., Objective: To better understand PC pathogenesis., Methods: RNA profiling was performed on biopsies taken from PC-involved and uninvolved plantar skin of seven genotyped PC patients (two K6a, one K6b, three K16, and one K17) as well as from control volunteers. Protein profiling was generated from tape-stripping samples., Results: A comparison of PC-involved skin biopsies to adjacent uninvolved plantar skin identified 112 differentially-expressed mRNAs common to patient groups harboring K6 (i.e., both K6a and K6b) and K16 mutations. Among these mRNAs, 25 encode structural proteins including keratins, small proline-rich and late cornified envelope proteins, 20 are related to metabolism and 16 encode proteases, peptidases, and their inhibitors including kallikrein-related peptidases (KLKs), and serine protease inhibitors (SERPINs). mRNAs were also identified to be differentially expressed only in K6 (81) or K16 (141) patient samples. Furthermore, 13 mRNAs were identified that may be involved in pain including nociception and neuropathy. Protein profiling, comparing three K6a plantar tape-stripping samples to non-PC controls, showed changes in the PC corneocytes similar, but not identical, to the mRNA analysis., Conclusion: Many differentially-expressed genes identified in PC-involved skin encode components critical for skin barrier homeostasis including keratinocyte proliferation, differentiation, cornification, and desquamation. The profiling data provide a foundation for unraveling the pathogenesis of PC and identifying targets for developing effective PC therapeutics., (Copyright © 2015 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.)

Background: Previous research revealed heterogeneity in the perfusion intensity within clinically homogenous-appearing plaques, without differences in erythema. In addition, an increased perfusion was found within the perilesional skin. This raises the question whether the heterogeneity in perfusion found both inside and outside a lesion influences the expression levels of genes and proteins involved in the pathogenesis of psoriasis., Objectives: To correlate the perfusion intensity to mRNA and protein expression of genes associated with the pathogenesis of psoriasis and to visualize the dynamics of the perfusion intensity over time using laser Doppler perfusion imaging., Methods: Fourteen patients with plaque psoriasis were included. The superficial microcirculation and clinical local scores (single usability metric, SUM, scores) were analysed in one representative lesion every 2 weeks. After 8 weeks 4 biopsies were taken, one from a highly perfused area (hotspot) and one from a low perfusion area (coldspot) of the lesional skin, one biopsy from the highly perfused perilesional skin and one from the distant uninvolved skin., Results: Statistically significant differences in mRNA and protein expression, including IL-17 and TBX21/T-Bet, were found between hotspots and coldspots, and between the highly perfused perilesional and the uninvolved skin. Hotspots tend to remain on the same location during 8 weeks of follow-up., Conclusions: Within homogenous-appearing psoriatic plaques, there are remarkable differences in mRNA and protein levels, which are correlated with the perfusion intensity and can be detected by using laser Doppler perfusion imaging. In addition, differences in mRNA and protein expression between the highly perfused perilesional skin and the uninvolved skin were found, indicating that several biological changes occur well before clinical changes become manifest., (© 2015 S. Karger AG, Basel.)

Background: Pachyonychia congenita (PC) is a rare autosomal dominant keratinizing disorder characterized by severe, painful, palmoplantar keratoderma and nail dystrophy, often accompanied by oral leucokeratosis, cysts and follicular keratosis. It is caused by mutations in one of five keratin genes: KRT6A, KRT6B, KRT6C, KRT16 or KRT17., Objectives: To identify mutations in 84 new families with a clinical diagnosis of PC, recruited by the International Pachyonychia Congenita Research Registry during the last few years., Methods: Genomic DNA isolated from saliva or peripheral blood leucocytes was amplified using primers specific for the PC-associated keratin genes and polymerase chain reaction products were directly sequenced., Results: Mutations were identified in 84 families in the PC-associated keratin genes, comprising 46 distinct keratin mutations. Fourteen were previously unreported mutations, bringing the total number of different keratin mutations associated with PC to 105., Conclusions: By identifying mutations in KRT6A, KRT6B, KRT6C, KRT16 or KRT17, this study has confirmed, at the molecular level, the clinical diagnosis of PC in these families., (© 2014 The Authors. British Journal of Dermatology published by John Wiley & Sons Ltd on behalf of British Association of Dermatologists.)

Unlabelled: Pachyonychia congenita (PC), a rare autosomal dominant disorder characterized by hypertrophic nail dystrophy, is classified into two main clinical subtypes: PC-1 and PC-2. PC-1 is associated with mutations in the KRT6A or KRT16 genes, whereas PC-2 is linked to KRT6B or KRT17 mutations. Blood samples were collected from three generations of a new Chinese PC-1 family, including three PC patients and five unaffected family members. A novel missense mutation p.Leu128Pro (c.383T>C) was identified in a highly conserved helix motif in domain 1A of K16. The disease haplotype carried the mutation and cosegregated with the affection status. PolyPhen2 and SIFTS analysis rated the substitution as probably damaging; Swiss-Model analysis indicated that the structure of the mutant protein contained an unnormal α-helix. Overexpression of mutant protein in cultured cells led to abnormal cell morphology., Conclusion: The wider spectrum of KRT16 mutations suggests that changes in codons 125, 127, and 132 are most commonly responsible for PC-1 and that proline substitution mutations at codons 127 or 128 may produce more severe disease. This study extends the KRT16 mutation spectrum and adds new information on the clinical and genetic diversity of PC.

There are several animal models illustrating dry eye pathophysiology. Current study would like to establish an ex vivo tissue culture model for characterizing dry eye. Human conjunctival explants were cultured under airlift or submerged conditions for up to 2 weeks, and only airlifted conjunctival cultures underwent increased epithelial stratification. Starting on day 4, the suprabasal cells displayed decreased K19 expression whereas K10 keratin became evident in airlift group. Pax6 nuclear expression attenuated already at 2 days, while its perinuclear and cytoplasmic expression gradually increased. MUC5AC and MUC19 expression dramatically decreased whereas the full thickness MUC4 and MUC16 expression pattern disappeared soon after initiating the airlift condition. Real time PCR showed K16, K10 and MUC16 gene up-regulated while K19, MUC5AC, MUC19 and MUC4 down-regulated on day 8 and day 14. On day 2 was the appearance of apoptotic epithelial and stromal cells appeared. The Wnt signaling pathway was transiently activated from day 2 to day 10. The inflammatory mediators IL-1β, TNF-α, and MMP-9 were detected in the conditioned media after 6 to 8 days. In conclusion, airlifted conjunctival tissue cultures demonstrated Wnt signaling pathway activation, coupled with squamous metaplasia, mucin pattern alteration, apoptosis and upregulation of proinflammatory cytokine expression. These changes mimic the pathohistological alterations described in dry eye. This correspondence suggests that insight into the pathophysiology of dry eye may be aided through the use of airlifted conjunctival tissue cultures.

Purpose: Patients with triple-negative breast cancer (TNBC) and residual disease after neoadjuvant chemotherapy generally have worse outcome; however, some patients with residual tumor after neoadjuvant chemotherapy do not relapse. We hypothesize that there are subgroups of patients with chemoresistant TNBC with different prognosis., Experimental Design: Forty-nine chemoresistant cases from 111 patients with TNBC treated with neoadjuvant chemotherapy (M.D. Anderson Cancer Center, Houston, TX) constituted the discovery cohort, and 25 chemoresistant samples from 47 neoadjuvant chemotherapy-treated TNBC (The Methodist Hospital, Houston, TX) were chosen for validation. Extended validation was carried out in 269 operable TNBC predicted to be chemoresistant by expression pattern from published datasets., Results: We established a seven-gene prognostic signature using dChip and gene set enrichment analyses. In the independent validation cohort, the classifier predicted correctly with positive predictive value of 75.0% and negative predictive value (i.e., relapse-free survival; RFS) of 76.9% at 3 years. Those predicted to relapse had a HR of 4.67 [95% confidence interval (CI): 1.27-17.15] for relapse in 3 years. In extended validation, patients predicted not to relapse exhibited 3-year RFS of 78.9%, whereas the 3-year RFS was 48.5% for patients predicted to relapse, with HR of 2.61 (95% CI: 1.52-4.49). The TNBC subgroup that predicted to have relatively favorable prognosis was characterized by high expression of "luminal-like" genes [androgen-receptor (AR) and GATA3], whereas the subgroup with worse prognosis was characterized by expression of cancer stem-cell markers., Conclusion: We developed a clinically relevant signature for patients with chemoresistant TNBC. For these women, new therapeutic strategies like targeting AR activation or cancer stem cells may need to be developed., (©2013 AACR)

Background: Pachyonychia congenita (PC) is a group of autosomal dominant keratinizing disorders caused by a mutation in one of 4 keratin genes. Previous classification schemes have relied on data from case series and case reports. Most patients in these reports were not genetically tested for PC., Objective: We sought to clarify the prevalence of clinical features associated with PC., Methods: We surveyed 254 individuals with confirmed keratin mutations regarding their experience with clinical findings associated with PC. Statistical comparison of the groups by keratin mutation was performed using logistic regression analysis., Results: Although the onset of clinical symptoms varied considerably among our patients, a diagnostic triad of toenail thickening, plantar keratoderma, and plantar pain was reported by 97% of patients with PC by age 10 years. Plantar pain had the most profound impact on quality of life. Other clinical findings reported by our patients included fingernail dystrophy, oral leukokeratosis, palmar keratoderma, follicular hyperkeratosis, hyperhidrosis, cysts, hoarseness, and natal teeth. We observed a higher likelihood of oral leukokeratosis in individuals harboring KRT6A mutations, and a strong association of natal teeth and cysts in carriers of a KRT17 mutation. Most keratin subgroups expressed a mixed constellation of findings historically reported as PC-1 and PC-2., Limitations: Data were obtained through questionnaires, not by direct examination. Patients were self- or physician-referred., Conclusions: We propose a new classification for PC based on the specific keratin gene affected to help clinicians improve their diagnostic and prognostic accuracy, correct spurious associations, and improve therapeutic development., (Copyright © 2011 American Academy of Dermatology, Inc. Published by Mosby, Inc. All rights reserved.)

Background: Mutations in the KRT6A or KRT16 gene cause pachyonychia congenita type 1 (PC-1), while mutations in KRT16 or KRT6C underlie focal palmoplantar keratoderma (FPPK). A new classification system of PC has been adopted based on the mutated gene. PC rarely presents the symptoms of diffuse plantar keratoderma. Mutation in the tail domain of keratins is rarely reported. PC combined with fissured tongue has never been described., Objectives: To investigate the genotype-phenotype correlations between clinical features and gene mutational sites in two unrelated southern Chinese PC pedigrees (one family presented with specific fissured tongue, the other with diffuse plantar keratoderma)., Materials & Methods: The whole coding regions of the KRT6A/KRT16/KRT17/KRT6B genes were amplified and directly sequenced to detect the mutation. To confirm the effect of the IVS8-2A>C mutation in KRT6A at the mRNA level, total RNA from the plantar lesion of a patient was extracted and reverse-transcribed to cDNA for sequence analysis., Results: Two novel de novo mutations, a splice acceptor site variant IVS8-2A>C (p.S487FfsX72) in KRT6A and a heterozygous substitution c.AA373&#95;374GG (p.N125G) within exon 1 of KRT16, were found separately in the two PC families., Conclusion: Genotype-phenotype correlations among PC patients with codon-125 mutation in KRT16 were established, while the phenotypes caused by the IVS8-2A>C mutation in KRT6A need further studies to confirm the rare feature of fissured tongue.

Immunohistochemical loss of keratin (K)13 is one of the most valuable diagnostic criteria for discriminating carcinoma in situ (CIS) from non-malignancies in the oral mucosa while K13 is stably immunolocalized in the prickle cells of normal oral epithelium. To elucidate the molecular mechanism for the loss of K13, we compared the immunohistochemical profiles for K13 and K16 which is not expressed in normal epithelia, but instead enhanced in CIS, with their mRNA levels by in-situ hybridization in formalin-fixed paraffin sections prepared from 23 CIS cases of the tongue, which were surgically removed. Reverse transcriptase-PCR was also performed using RNA samples extracted from laser-microdissected epithelial fragments of the serial paraffin sections in seven of the cases. Although more enhanced expression levels for K16 were confirmed at both the protein and gene levels in CIS in these seven cases, the loss of K13 was associated with repressed mRNA levels in four cases, but not in the other three cases. The results suggest that the loss of K13 is partly due to its gene repression, but may also be due to some unknown post-translational events.

Palmoplantar keratoderma is a hallmark of pachyonychia congenita (PC) and focal non-epidermolytic palmoplantar keratoderma (FNEPPK). By generating keratin 16 (Krt16)-deficient mice, Lessard and Coulombe, as described in this issue, have generated a mouse model to replicate these palmoplantar lesions. Studies using this model may provide novel insights into the molecular mechanisms responsible for the formation of palmoplantar lesions in PC and FNEPPK patients.

Keratin 16 (KRT16 in human, Krt16 in mouse), a type I intermediate filament protein, is constitutively expressed in epithelial appendages and is induced in the epidermis upon wounding and other stressors. Mutations altering the coding sequence of KRT16 cause pachyonychia congenita (PC), a rare autosomal dominant disorder characterized by hypertrophic nail dystrophy, oral leukokeratosis, and palmoplantar keratoderma (PPK). PPK associated with PC is extremely painful and compromises patient mobility, making it the most debilitating PC symptom. In this study, we show that, although inherited in a recessive manner, the inactivation of Krt16 in mice consistently causes oral lesions as well as PPK-like hyperkeratotic calluses on Krt16(-/-) front and hind paws, which severely compromise the animals' ability to walk. Our findings call into question the view that PC-related PPK arises exclusively as a gain-of-function on account of dominantly acting mutated keratins, and highlight the key role of modifiers in the clinical heterogeneity of PC symptoms.

Background: Pachyonychia congenita (PC) is an autosomal dominant, very rare keratin disorder caused by mutations in any of at least four genes (KRT6A, KRT6B, KRT16 or KRT17), which can lead to hypertrophic nail dystrophy and palmoplantar keratoderma, among other manifestations. Classically, patients with mutations in KRT6A and KRT16 have been grouped to the PC-1 subtype (Jadassohn-Lewandowsky type) and KRT6B and KRT17 to PC-2 (Jackson-Lawler type)., Objectives: To describe clinical heterogeneity among patients with PC who have genetic mutations in KRT6A and KRT16., Methods: In 2004, the Pachyonychia Congenita Project established the International PC Research Registry (IPCRR) for patients with PC. All patients reporting here underwent genetic testing and responded to a standardized, validated survey about their PC symptoms. We report results from 89 patients with KRT6A mutations and 68 patients with KRT16 mutations., Results: Patients with PC who have KRT6A and KRT16 mutations display distinct phenotypic differences. Patients with PC-K6a experience earlier onset, more extensive nail disease and more substantial disease outside palms and soles, as they reported a higher prevalence of oral leucokeratosis (P < 0·001), cysts (P < 0·001) and follicular hyperkeratosis (P < 0·001) compared with their PC-K16 counterparts., Conclusion: Phenotypic differences between patients with KRT6A and KRT16 mutations support adoption of a new classification system based on the mutant gene (PC-6a, PC-16) rather than the PC-1 nomenclature., (© 2011 The Authors. BJD © 2011 British Association of Dermatologists.)

The mechanism of action of pimecrolimus (PIM) on atopic lesions is still under consideration. Thus far, we have evidence of its anti-inflammatory and immunomodulatory activity, and recent papers focus on its effect on epidermal barrier function. This study analysed changes in the expression of genes associated with skin barrier dysfunction in atopic dermatitis (AD) skin lesions after 2 weeks of exposure to PIM 1% cream. A real-time quantitative PCR analysis of selected epidermal differentiation complex genes and three alternative pathway keratins was performed in skin biopsies from 11 individuals with AD before and after PIM exposure. The real-time quantitative PCR analysis was compared to non-lesional skin in the same patients. Involucrin, a small proline-rich region (SPRR) 2C gene, and alternative pathway keratin 16 showed significant over-expression in lesional skin followed by significant decrease after PIM therapy. The SPRR1A gene, S100A9, and keratin 6A were also increased; however, the decrease after PIM treatment was not significant. The changes in S100 A2, A7 and A8 followed a similar course with borderline significance. SPRR4 had a significant decrease in expression in lesional versus non-lesional skin, which persisted after PIM treatment. No significant changes were detected in mRNA expression levels of filaggrin and loricrin. Our results suggest that PIM can be effective in restoring the epidermal barrier in patients with AD at least in part by its impact on expression of genes, which are important for the normal barrier function of skin., (© 2011 John Wiley & Sons A/S.)

Purpose: Circulating tumor cells (CTC) might function as early markers for breast cancer metastasis or monitoring therapy efficacy. Enrichment and identification of CTCs are based on epithelial markers that might be modulated during epithelial-mesenchymal transition. Little is known about the expression of keratins in CTCs and whether all CTCs can be detected with antibodies directed against a limited panel of keratins., Experimental Design: Protein expression of keratin 2, 4-10, 13-16, 18, and 19 were assessed by a cocktail of antibodies (C11, AE1, AE3, and K7) and keratin antibodies C11 and A45-B/B3 alone in 11 breast cancer cell lines and 50 primary breast carcinomas and their lymph node metastases. Furthermore, CTCs were assessed in blood of 70 metastatic breast cancer patients., Results: Claudin-low cell lines did not show expression of normal breast epithelial keratins but were positive for K14 and K16, detected by the cocktail only. Primary breast carcinomas showed changes in keratin expression during metastatic progression to the lymph nodes. In 35 of 70 patients CTCs were identified, of which 83%, 40%, and 57% were identified by the cocktail, C11 and A45-B/B3, respectively. Identification of CTCs by the cocktail was associated with shorter survival (P < 0.01). In silico analyses revealed association between KRT16 expression and shorter relapse-free survival in metastatic breast cancer., Conclusion: Breast cancer cells show a complex pattern of keratin expression with potential biologic relevance. Individual keratin antibodies recognizing only a limited set of keratins inherit the risk to miss biologically relevant CTCs in cancer patients, and antibody cocktails including these keratins are therefore recommended., (©2012 AACR.)

Background: Pachyonychia congenita (PC) is a rare keratin disorder that typically presents with nail dystrophy and focal plantar keratoderma. We present seven cases of PC with transgrediens involvement of the dorsal feet., Objectives: To document the extension of their disease to the dorsum of the feet in patients with mutation-confirmed PC, to report the natural history of PC with such transgrediens involvement, to generate hypotheses regarding aetiology, and to suggest prevention and treatment modalities., Methods: Genetically confirmed cases of PC with transgrediens foot involvement were verified through the International Pachyonychia Congenita Research Registry (IPCRR) and characterized via telephone survey and photography., Results: Seven patients with PC in the IPCRR were confirmed to have transgrediens lesions on the dorsal feet (six KRT6A mutations; one KRT16 mutation). Six cases had pre-existing nontransgrediens keratoderma and all cases reported standing, wearing shoes, foot moisture, and/or infection as exacerbating or predisposing factors. Improvement, reported in six cases, was attributed to use of antibiotics or gentian violet, or improved footwear., Conclusions: Transgrediens involvement of the dorsal feet is a rare manifestation of mutation-confirmed PC and may be more common in patients who carry a KRT6A mutation. Trauma, friction, infection and wound healing may exacerbate or predispose toward transgrediens lesions. It remains to be proven whether transgrediens-associated infection is causal or represents a primary or secondary process. Patients with PC who develop transgrediens lesions may benefit from fungal and bacterial cultures, followed by appropriate antimicrobial treatments. Efforts to decrease skin friction and moisture may also improve and/or prevent transgrediens spread., (© 2011 The Authors. BJD © 2011 British Association of Dermatologists.)

Pachyonychia congenita (PC) is a rare, autosomal dominant keratin disorder caused by mutations in four genes (KRT6A, KRT6B, KRT16, or KRT17). The International PC Research Registry is a database with information on patients' symptoms as well as genotypes. We sought to describe the heterogeneity of clinical symptoms and to investigate possible genotype-phenotype correlations in patients with two types of K16 mutations, p.Asn125 and p.Arg127, causing the PC-16 subtype of PC. We found that clinical symptoms depended on the type of amino-acid substitution. Patients with p.Asn125Asp and p.Arg127Pro mutations exhibited more severe disease than patients carrying p.Asn125Ser and p.Arg127Cys mutations in terms of age of onset of symptoms, extent of nail involvement, and impact on daily quality of life. We speculate that amino-acid substitutions causing larger, more disruptive changes to the K16 protein structure, such as a change in amino-acid charge in the p.Asn125Asp mutation or a bulky proline substitution in the p.Arg127Pro mutation, may also lead to more severe disease phenotypes. The variation in phenotypes seen with different substitutions at the same mutation site suggests a genotype-phenotype correlation. Knowledge of the exact gene defect is likely to assist in predicting disease prognosis and clinical management.

Pachyonychia congenita (PC) is a rare autosomal dominant skin disorder characterized predominantly by nail dystrophy and painful palmoplantar keratoderma. Additional clinical features include oral leukokeratosis, follicular keratosis, and cysts (steatocysts and pilosebaceous cysts). PC is due to heterozygous mutations in one of four keratin genes, namely, KRT6A, KRT6B, KRT16, or KRT17. Here, we report genetic analysis of 90 new families with PC in which we identified mutations in KRT6A, KRT6B, KRT16, or KRT17, thereby confirming their clinical diagnosis. A total of 21 previously unreported and 22 known mutations were found. Approximately half of the kindreds had mutations in KRT6A (52%), 28% had mutations in KRT16, 17% in KRT17, and 3% of families had mutations in KRT6B. Most of the mutations were heterozygous missense or small in-frame insertion/deletion mutations occurring within one of the helix boundary motif regions of the keratin polypeptide. More unusual mutations included heterozygous splice site mutations, nonsense mutations, and a 1-bp insertion mutation, leading to a frameshift and premature termination codon. This study, together with previously reported mutations, identifies mutation hotspot codons that may be useful in the development of personalized medicine for PC.

Treatment with the natural chemical sulforaphane (SF) ameliorates skin blistering in keratin 14 (K14)-deficient mice, correlating with the induction of K16 and K17 in the basal layer of epidermis (Kerns et al., PNAS 104:14460, 2007). Here we address the basis for the SF-mediated K16 and K17 induction in mouse epidermis in vivo. As expected, induction of K16 partly depends on the transcription factor Nrf2, which is activated by SF exposure. Strikingly, K17 induction occurs independently of Nrf2 activity and parallels the decrease in glutathione occurring shortly after epidermal exposure to SF. Pharmacological manipulation of glutathione levels in mouse epidermis in vivo alters K17 and K16 expression in the expected manner. We present findings suggesting that select MAP kinases participate in mediating the Nrf2- and glutathione-dependent alterations in K16 and K17 levels in SF-treated epidermis. These findings advance our understanding of the effect of SF on gene expression in epidermis, point to a role for glutathione in mediating some of these effects, and establish that SF induces the expression of two contiguous and highly related genes, K16 and K17, via distinct mechanisms.

Conspicuous facial pores are one type of serious esthetic defects for many women. We previously reported that the severity of impairment of skin architecture around facial pores correlates well with the appearance of facial pores in several ethnic groups. In our last report, we showed that serum levels of insulin-like growth factor-1 (IGF-1) correlate well with facial pore size and with the severity of impairment of epidermal architecture around facial pores. However, our results could not fully explain the implication between facial pores and IGF signaling. In this study, we conducted a histological analysis of facial skin to determine whether potential changes in IGF-1 availability occur in the skin with or without conspicuous pores. Immunohistochemical observations showed that expression of insulin-like growth factor binding protein-3 (IGFBP-3) is limited to the suprapapillary epidermis around facial pores and to basal cells of rete pegs without tips in epidermis with conspicuous pores. In contrast, in basal cells of skin without conspicuous pores, IGFBP-3 expression is very low. Ki-67 and IGF-1 receptor-positive cells are abundant in basal cells in the tips of the rete pegs in skin with typical epidermal architecture around facial pores. No obvious differences were observed in the expression of filaggrin, involucrin, K1, K6 or K17 in skin with or without conspicuous pores. However, increased expression of K16 was observed in skin with conspicuous pores suggesting hyperproliferation. These results suggest that the IGF-1/IGFBP-3 signaling pathway is involved in the formation of conspicuous facial pores due to the epidermal architecture around facial pores.