Increased Keratinocyte Proliferation Research Articles

Sensory neurons increase keratinocyte proliferation through CGRP release in a tissue engineered in vitro model of innervation in psoriasis

Skin denervation has been shown to cause remission of psoriatic lesions in patients, which can reappear if reinnervation occurs. This effect can be induced by the activation of dendritic cells through sensory innervation. However, a direct effect of nerves on the proliferation of keratinocytes involved in the formation of psoriatic plaques has not been investigated. We developed, by tissue engineering, a model of psoriatic skin made of patient skin cells that showed increased keratinocyte proliferation and epidermal thickness compared to healthy controls. When this model was treated with CGRP, a neuropeptide released by sensory neurons, an increased keratinocyte proliferation was observed in the psoriatic skin model, but not in the control. When a sensory nerve network was incorporated in the psoriatic model and treated with capsaicin to induce neuropeptide release, an increase of keratinocyte proliferation was confirmed, which was blocked by a CGRP antagonist while no difference was noticed in the innervated healthy control. We showed that sensory neurons can participate directly to keratinocyte hyperproliferation in the formation of psoriatic lesions through the release of CGRP, independently of the immune system. Our unique tissue-engineered innervated psoriatic skin model could be a valuable tool to better understand the mechanism by which nerves may modulate psoriatic lesion formation in humans. Statement of significanceThis study shows that keratinocytes extracted from patients’ psoriatic skin retain, at least in part, the disease phenotype. Indeed, when combined in a 3D model of tissue-engineered psoriatic skin, keratinocytes exhibited a higher proliferation rate, and produced a thicker epidermis than a healthy skin control. In addition, their hyperproliferation was aggravated by a treatment with CGRP, a neuropeptide released by sensory nerves. In a innervated model of tissue-engineered psoriatic skin, an increase in keratinocyte hyperproliferation was also observed after inducing neurons to release neuropeptides. This effect was prevented by concomitant treatment with an antagonist to CGRP. Thus, this study shows that sensory nerves can directly participate to affect keratinocyte hyperproliferation in psoriasis through CGRP release.

Retinoids in the treatment of photoageing: A histological study of topical retinoid efficacy in black skin.

Photoageing describes complex cutaneous changes that occur due to chronic exposure to solar ultraviolet radiation (UVR). The 'gold standard' for the treatment of photoaged white skin is all-trans retinoic acid (ATRA); however, cosmetic retinol (ROL) has also proven efficacious. Recent work has identified that black skin is susceptible to photoageing, characterized by disintegration of fibrillin-rich microfibrils (FRMs) at the dermal-epidermal junction (DEJ). However, the impact of topical retinoids for repair of black skin has not been well investigated. To determine the potential of retinoids to repair photoaged black skin. An exploratory intervention study was performed using an invivo, short-term patch test protocol. Healthy but photoaged black volunteers (>45 years) were recruited to the study, and participant extensor forearms were occluded with either 0.025% ATRA (n = 6; 4-day application due to irritancy) or ROL (12-day treatment protocol for a cosmetic) at concentrations of 0.3% (n = 6) or 1% (n = 6). Punch biopsies from occluded but untreated control sites and retinoid-treated sites were processed for histological analyses of epidermal characteristics, melanin distribution and dermal remodelling. Treatment with ATRA and ROL induced significant acanthosis (all p < 0.001) accompanied by a significant increase in keratinocyte proliferation (Ki67; all p < 0.01), dispersal of epidermal melanin and restoration of the FRMs at the DEJ (all p < 0.01), compared to untreated control. This study confirms that topical ATRA has utility for the repair of photoaged black skin and that ROL induces comparable effects on epidermal and dermal remodelling, albeit over a longer timeframe. The effects of topical retinoids on black photoaged skin are similar to those reported for white photoaged skin and suggest conserved biology in relation to repair of UVR-induced damage. Further investigation of topical retinoid efficacy in daily use is warranted for black skin.

Lysophosphatidic acid down-regulates human RIPK4 mRNA in keratinocyte- derived cell lines.

The tight control of proliferating keratinocytes is vital to the successful function of the skin. Differentiation of dividing cells is necessary to form a skin barrier. The same dividing cells are necessary to heal wounds and when malignant form tumors. RIPK4, a serine-threonine kinase, plays critical roles in these processes. Its loss of function was associated with pathological keratinocyte proliferation and development of squamous cell carcinoma (SCC) in humans and mice. The current study extends previous findings in the importance of RIPK4 in keratinocyte proliferation. A serum-derived phospholipid, lysophosphatidic acid (LPA), was identified as an important biologic inhibitor of RIPK4. LPA functions by inhibiting the transcription of RIPK4 mRNA. LPA treatment led to increased keratinocyte proliferation, and this was compromised in cells with reduced RIPK4 expression. The current study may help to explain the mechanism by which RIPK4 was downregulated during SCC progression and provide insights on RIPK4 functions. It may also allow for targeting of RIPK4 through a natural component of serum.

Effects of an Adipose Mesenchymal Stem Cell-Derived Conditioned medium and TGF-β1 on Human Keratinocytes In Vitro.

Human keratinocytes play a crucial role during skin wound healing and in skin replacement therapies. The secretome of adipose-derived stem cells (ASCs) has been shown to secrete pro-healing factors, among which include TGF-β1, which is essential for keratinocyte migration and the re-epithelialization of cutaneous wounds during skin wound healing. The benefits of an ASC conditioned medium (ASC-CM) are primarily orchestrated by trophic factors that mediate autocrine and paracrine effects in keratinocytes. Here, we evaluated the composition and the innate characteristics of the ASC secretome and its biological effects on keratinocyte maturation and wound healing in vitro. In particular, we detected high levels of different growth factors, such as HGF, FGFb, and VEGF, and other factors, such as TIMP1 and 4, IL8, PAI-1, uPA, and IGFBP-3, in the ASC-CM. Further, we investigated, using immunofluorescence and flow cytometry, the distinct effects of a human ASC-CM and/or synthetic TGF-β1 on human keratinocyte proliferation, migration, and cell apoptosis suppression. We demonstrated that the ASC-CM increased keratinocyte proliferation as compared to TGF-β1 treatment. Further, we found that the ASC-CM exerted cell cycle progression in keratinocytes via regulating the phases G1, S, and G2/M. In particular, cells subjected to the ASC-CM demonstrated increased DNA synthesis (S phase) compared to the TGF-β1-treated KCs, which showed a pronounced G0/G1 phase. Furthermore, both the ASC-CM and TGF-β1 conditions resulted in a decreased expression of the late differentiation marker CK10 in human keratinocytes in vitro, whereas both treatments enhanced transglutaminase 3 and loricrin expression. Interestingly, the ASC-CM promoted significantly increased numbers of keratinocytes expressing epidermal basal keratinocyte markers, such DLL1 and Jagged2 Notch ligands, whereas those ligands were significantly decreased in TGF-β1-treated keratinocytes. In conclusion, our findings suggest that the ASC-CM is a potent stimulator of human keratinocyte proliferation in vitro, particularly supporting basal keratinocytes, which are crucial for a successful skin coverage after transplantation. In contrast, TGF-β1 treatment decreased keratinocyte proliferation and specifically increased the expression of differentiation markers in vitro.

Current insight flavonoid quercetin in epithelial maturation vagina for menopause treatment: a narrative review

Background: Menopause affects all parts of the women's body system, especially in the genitourinary area. The impact on the genitourinary system causes post-menopausal women to complain of menopause-related genitourinary syndrome due to vaginal atrophy, sexual dysfunction, and pelvic organ prolapse in the long term. Various research has been conducted to find alternative therapy, especially flavonoid quercetin, due to its phytoestrogen characteristic. Methods: This review article assembles much literature to highlight the menopause effect on the vagina and the phytoestrogen effect of flavonoid quercetin as a potential alternative therapy. This study aims to develop future experimental studies of quercetin's effect on the vagina. Results: Hypoestrogenic condition affects all vaginal layers. This is due to estrogen receptors distributed in the epithelial lining, lamina propria, and muscular layer of the vagina. In vitro studies showed quercetin induces cell proliferation by activating signal pathway b-catenin/c-Myc/cyclin A2, mediated by estrogen receptor. The signal pathway increases the cell cycle transition to the G1 phase. It also increases keratinocyte proliferation and migration, proliferating cell nuclear antigen, and positive a6 and b1 integrin cells in the basal layer of the epidermis, which causes thickening of the epidermis. Conclusion: Flavonoid quercetin is a promising alternative for menopause therapy in the vagina. Further, in vivo studies are needed to confirm the result of in vitro study and as basic data for comparison with other potent phytoestrogen.

Dasatinib Ointment Promotes Healing of Murine Excisional Skin Wound.

Dasatinib, a tyrosine kinase inhibitor, has been shown to produce anti-inflammatory activity and impair vascular integrity in vivo, including during skin wound healing, potentially promoting the repair process. Given that dasatinib is a lipophilic small molecule capable of penetrating skin, topical dasatinib might provide benefits in wound healing. In the present study, we investigated the impact of dasatinib ointments in skin wound healing in mice. A full thickness excisional skin wound (4 mm diameter) was generated on the shaved dorsum of eight-week-old C57BL/6 mice. Dasatinib ointment (0.1 or 0.2% w/w) or ointment base was applied twice daily (every 12 h) for 10 days. Elizabethan collars were used to prevent animal licking. The wound size was monitored daily for 14 days. The results showed that dasatinib ointments, particularly 0.1% dasatinib, promoted a 16-23% reduction in wound size (p < 0.05) during day 2 to day 6 postinjury compared to controls. Immunohistochemistry analyses demonstrated a reduction in wound neutrophils (38% reduction, p = 0.04), macrophages (47% reduction, p = 0.005), and tumor necrosis factor-α levels (73% reduction, p < 0.01), together with an induction of vascular leakage-mediated fibrin(ogen) accumulation (2.5-fold increase, p < 0.01) in the wound during day 3 postinjury (an early phase of repair) in 0.1% dasatinib-treated mice relative to control mice. The anti-inflammatory and vascular hyperpermeability activities of dasatinib were associated with an enhanced healing process, including increased keratinocyte proliferation (1.8-fold increase in Ki67+ cells, p < 0.05) and augmented angiogenesis (1.7-fold increase in CD31+ area, p < 0.05), compared to the ointment base-treated group. Following treatment with 0.2% dasatinib ointment, minor wound bleeding and scab reformation were observed during the late phase, which contributed to delayed healing. In conclusion, our data suggest that dasatinib ointment, mainly at 0.1%, promotes the repair process by reducing inflammation and producing a local and temporal vascular leakage, leading to an increase in fibrin(ogen) deposition, re-epithelialization, and angiogenesis. Therefore, topical dasatinib might be a potential novel candidate to facilitate skin wound healing.

Interleukin 17 Receptor C, but not Polo-like Kinase 4, Plays a Critical Role in the Development of Skin Papilloma and Psoriasis in Mouse Models

Abstract Introduction/Objective Hyperproliferation of epidermis is a histopathological hallmark of skin cancers and psoriasis. As a master regulator of centriole replication, overexpression of Polo-like kinase 4 (PLK4) has been identified in skin cancers. The aberrant proliferation of epidermal keratinocytes provoked by interleukin 17 (IL17) leads to psoriasis. Thus, targeting centriole replication and IL17 signaling simultaneously has been speculated as a potential therapeutic strategy. We hypothesized that inhibition of centriole duplication enhances the blockade of epidermal proliferation through Il17rc knockout. Methods/Case Report To test our hypothesis, 37 mice were used to imbed a two-stage model of skin carcinogenesis using wild-type (WT), IL17 receptor A (T779A) knock-in (Il17ra(T779A)-KI), and IL17 receptor C knock-out (Il17rc-KO) C57BL/6J mouse strains. Furthermore, an imiquimod-induced psoriasis model was established using 69 mice with the same strains. Results (if a Case Study enter NA) In our two-stage skin carcinogenesis model, Il17ra(T779A)-KI mice showed significantly decreased tumor incidence, tumor multiplicity, and tumor volume compared to the WT mice. Il-17rc-KO mice didn’t develop any skin papilloma. The skin papilloma formed never progressed into squamous carcinoma. However, centrinone, a selective inhibitor of PLK4, didn’t affect skin papilloma formation or epidermal thickening. In our psoriasis model, the epidermis thickness of Il-17rc-KO mice was dramatically decreased compared to WT and Il17ra(T779A)-KI mice. There was no significant difference between WT and Il17ra(T779A)-KI mice in terms of skin lesion and thickening of the epidermis. Centrinone didn’t stall the thickening of the epidermis in the psoriasis model. IHC staining showed significantly increased Ki67+ basal keratinocytes in the untreated skin of Il17ra(T779A)-KI male mice compared to WT mice. In imiquimod-treated skin, the percentage of Ki67+ basal keratinocytes significantly decreased in the order from WT, Il17ra(T779A)-KI to Il17rc-KO mice. Conclusion Our data suggest that the proliferation of keratinocytes is not stalled by centrinone but is inhibited by Il17rc-KO. Il17ra(T779A)-KI significantly inhibits skin papilloma formation, but slightly decreases epidermal thickening in the psoriasis model. However, Il17ra(T779A)-KI increases keratinocyte proliferation based on Ki67 staining in the untreated normal skin.

Abstract 1313: IL-17RC, but not PLK4 inhibitor centrinone, plays a critical role in the development of skin papilloma and psoriasis in mouse models

Abstract Introduction: Hyperproliferation of epidermis is a histopathological hallmark of skin disorders, such as skin cancers and psoriasis. Polo-like kinase 4 (PLK4) is a master regulator of centriole replication and its overexpression has been identified in non-melanoma skin cancers. The aberrant proliferation of epidermal keratinocytes provoked by interleukin-17 (IL-17) leads to psoriasis. Thus, targeting centriole replication and IL-17 signaling simultaneously has been speculated as a potential therapeutic strategy. We hypothesized that inhibition of centriole duplication might enhance the blockade of epidermal proliferation through Il-17rc knockout. Methods: To investigate cooperation between IL-17 signaling and centriole duplication in the proliferation of epidermis, we used 37 mice to establish a two-stage model of skin carcinogenesis and 69 mice to establish imiquimod-induced psoriasis model in wild-type (WT), IL-17 receptor A (T779A) knock-in (Il-17ra(T779A)-KI) and IL-17 receptor C knock-out (Il-17rc-KO) C57BL/6J mouse strains. Results: During our 13-week monitor of the two-stage skin carcinogenesis model, we found that Il-17ra(T779A)-KI mice showed significantly decreased tumor incidence, tumor multiplicity, and tumor volume, compared to the wild-type mice. Il-17rc-KO mice didn’t develop any skin papilloma. The skin papilloma formed never progressed into squamous carcinoma based on our immunohistochemical (IHC) staining of laminin and cytokeratin 8. However, centrinone, a selective inhibitor of PLK4, didn’t affect skin papilloma formation or epidermal thickening. In our psoriasis model, we found that the thickness of the epidermis of Il-17rc-KO mice was dramatically decreased, compared to WT and Il-17ra(T779A)-KI mice. There was no significant difference between WT and Il-17ra(T779A)-KI mice, in terms of psoriasis formation and the thickness of the epidermis. Centrinone didn’t stall the thickening of the epidermis in the psoriasis model. IHC staining showed significantly increased Ki-67+ basal keratinocytes in the untreated skin of Il-17ra(T779A)-KI male mice, but not in female mice, compared to WT mice. In imiquimod-treated skin, the percentage of Ki-67+ basal keratinocytes significantly decreased in Il-17ra(T779A)-KI mice, compared to WT mice. The percentage of Ki-67+ basal keratinocytes was also dramatically decreased in Il-17rc-KO mice, compared to WT and Il-17ra(T779A)-KI mice. Conclusion: Our findings suggest that the proliferation of keratinocytes is not stalled by PLK4 inhibitor centrinone but is inhibited by Il-17rc-KO. Il-17ra(T779A)-KI significantly inhibits skin papilloma formation, but only slightly decreases epidermal thickening in the psoriasis model. However, in the untreated normal skin, Il-17ra(T779A)-KI increases keratinocyte proliferation based on Ki-67 staining. Citation Format: Ben Jin, Yongfeng Zhang, Haiyan D. Miller, Ling He, Zongbing You. IL-17RC, but not PLK4 inhibitor centrinone, plays a critical role in the development of skin papilloma and psoriasis in mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1313.

Tapping basement membrane motifs: Oral junctional epithelium for surface-mediated soft tissue attachment to prevent failure of percutaneous devices

Teeth, long-lasting percutaneous organs, feature soft tissue attachment through adhesive structures, hemidesmosomes, in the junctional epithelium basement membrane adjacent to teeth. This soft tissue attachment prevents bacterial infection of the tooth despite the rich – and harsh – microbial composition of the oral cavity. Conversely, millions of percutaneous devices (catheters, dental, and orthopedic implants) fail from infection yearly. Standard of care antibiotic usage fuels antimicrobial resistance and is frequently ineffective. Infection prevention strategies, like for dental implants, have failed in generating durable soft tissue adhesion – like that seen with the tooth – to prevent bacterial colonization at the tissue-device interface. Here, inspired by the impervious natural attachment of the junctional epithelium to teeth, we synthesized four cell adhesion peptide (CAPs) nanocoatings, derived from basement membranes, to promote percutaneous device soft tissue attachment. The two leading nanocoatings upregulated integrin-mediated hemidesmosomes, selectively increased keratinocyte proliferation compared to fibroblasts, which cannot form hemidesmosomes, and expression of junctional epithelium adhesive markers. CAP nanocoatings displayed marked durability under simulated clinical conditions and the top performer CAP nanocoating was validated in a percutaneous implant murine model. Basement membrane CAP nanocoatings, inspired by the tooth and junctional epithelium, may provide an alternative anti-infective strategy for percutaneous devices to mitigate the worldwide threat of antimicrobial resistance. Statement of significancePrevention and management of medical device infection is a significant healthcare challenge. Overzealous antibiotic use has motivated alternative material innovations to prevent infection. Here, we report implant cell adhesion peptide nanocoatings that mimic a long-lasting, natural “medical device,” the tooth, through formation of cell adhesive structures called hemidesmosomes. Such nanocoatings sidestep the use of antimicrobial or antibiotic elements to form a soft-tissue seal around implants. The top performing nanocoatings prompted expression of hemidesmosomes and defensive factors to mimic the tooth and was validated in an animal model. Application of cell adhesion peptide nanocoatings may provide an alternative to preventing, rather that necessarily treating, medical device infection across a range of device indications, like dental implants.

α4/α9 Integrins Coordinate Epithelial Cell Migration Through Local Suppression of MAP Kinase Signaling Pathways.

Adhesion of basal keratinocytes to the underlying extracellular matrix (ECM) plays a key role in the control of skin homeostasis and response to injury. Integrin receptors indirectly link the ECM to the cell cytoskeleton through large protein complexes called focal adhesions (FA). FA also function as intracellular biochemical signaling platforms to enable cells to respond to changing extracellular cues. The α4β1 and α9β1 integrins are both expressed in basal keratinocytes, share some common ECM ligands, and have been shown to promote wound healing in vitro and in vivo. However, their roles in maintaining epidermal homeostasis and relative contributions to pathological processes in the skin remain unclear. We found that α4β1 and α9β1 occupied distinct regions in monolayers of a basal keratinocyte cell line (NEB-1). During collective cell migration (CCM), α4 and α9 integrins co-localized along the leading edge. Pharmacological inhibition of α4β1 and α9β1 integrins increased keratinocyte proliferation and induced a dramatic change in cytoskeletal remodeling and FA rearrangement, detrimentally affecting CCM. Further analysis revealed that α4β1/α9β1 integrins suppress extracellular signal-regulated kinase (ERK1/2) activity to control migration through the regulation of downstream kinases including Mitogen and Stress Activated Kinase 1 (MSK1). This work demonstrates the roles of α4β1 and α9β1 in regulating migration in response to damage cues.

Phospholipase A2 enzymes represent a shared pathogenic pathway in psoriasis and pityriasis rubra pilaris.

Altered epidermal differentiation along with increased keratinocyte proliferation is a characteristic feature of psoriasis and pityriasis rubra pilaris (PRP). However, despite this large degree of overlapping clinical and histologic features, the molecular signatures these skin disorders share are unknown. Using global transcriptomic profiling, we demonstrate that plaque psoriasis and PRP skin lesions have high overlap, with all differentially expressed genes in PRP relative to normal skin having complete overlap with those in psoriasis. The major common pathway shared between psoriasis and PRP involves the phospholipases PLA2G2F, PLA2G4D, and PLA2G4E, which were found to be primarily expressed in the epidermis. Gene silencing each of the 3 PLA2s led to reduction in immune responses and epidermal thickness both in vitro and in vivo in a mouse model of psoriasis, establishing their proinflammatory roles. Lipidomic analyses demonstrated that PLA2s affect mobilization of a phospholipid-eicosanoid pool, which is altered in psoriatic lesions and functions to promote immune responses in keratinocytes. Taken together, our results highlight the important role of PLA2s as regulators of epidermal barrier homeostasis and inflammation, identify PLA2s as a shared pathogenic mechanism between PRP and psoriasis, and as potential therapeutic targets for both diseases.

082 Bath-PUVA therapy ameliorates pathogenetic chemokines for psoriasis

Psoriasis is a T-cell mediated autoimmune disease characterized by increased keratinocyte proliferation. In psoriasis, chemokines are secreted by a wide variety of cells (e.g. keratinocytes, dendritic cells, monocytes), which migrate immune cells (Th17 cells, Th1 cells, neutrophils, etc.) involved in the pathogenesis. Bath-psoralen plus ultraviolet A (PUVA) treatment is an established phototherapy for psoriasis, but its effects on chemokine levels remain unknown. We measured the levels of 22 serum chemokines by using a flow cytometry bead-based immunoassay in 20 psoriasis patients first treated with bath-PUVA therapy between 2007 and 2011 in our hospital.

139 Local dermatitis upon tapestripping and vaseline treatment of healthy human skin is attenuated by coal tar: insights into AHR-targeting therapies for inflammatory skin diseases

Coal tar therapy, an old dermatological therapy, exerts its therapeutic effect through activation of the aryl hydrocarbon receptor (AHR) and AHR-targeting molecules show therapeutic efficacy in phase 2/3 trials for chronic inflammatory skin diseases. The anti-inflammatory mechanisms of AHR activation in the skin are partially understood, but mostly studied in animal models. Here we aimed to investigate the therapeutic effects of AHR activation by coal tar application on epidermal responses after skin barrier disruption and inflammatory cues in human skin. Healthy volunteers (n=9) were treated with pix lithanthracis (5% in vaseline lanette) or vehicle (vaseline lanette) after complete removal of the stratum corneum by tape stripping. Non-invasive biophysical measurements (TEWL, hydration, erythema), stratum corneum tapes (NMF and protein profiling) and biopies were taken during 4 days of treatment. Application of vaseline on tapestripped skin led to an acute inflammatory response characterized by increased keratinocyte proliferation and subsequent acanthosis, a dense dermal CD45+ and MPO+ immune cell infiltrate indicative of (neutrophilic) granulocytes and increased levels of human beta defensin-2 (hBD2). Most strikingly, coal tar treated sites clearly showed less immune cell infiltrate, less pronounced acanthosis and hBD2 levels in the stratum corneum were significantly lower than upon vaseline treatment. No differential effects were found between vaseline and coal tar treatment for the other parameters we investigated. This in vivo human acute dermatitis model indicates that activation of AHR signaling reduces the influx of inflammatory cells potentially resulting in less epidermal activation. The lack in positive effects on skin barrier parameters by coal tar may be due to the relative short study period. Further investigations on the inflammatory signaling molecules driving the acute inflammation can provide new insights in the coal tar and AHR-mediated therapeutic effects.

Interleukin-17A Drives IL-19 and IL-24 Expression in Skin Stromal Cells Regulating Keratinocyte Proliferation.

IL-17A has been shown to be up-regulated in psoriasis lesions and is central to psoriasis pathogenesis. IL-19, along with other IL-20 subfamily cytokines such as IL-20 and IL-24, is induced by IL-17A and contributes especially to epidermal hyperplasia in psoriasis. However, the regulation, cellular sources of IL-19 and whether targeting of IL-17A by biologics influence IL-19 expression is not completely understood. To investigate the regulation of IL-19 by IL-17A in psoriasis, the imiquimod-induced psoriasis mouse (IMQ) model was used. Enhanced expression of IL-17A in the IMQ model was achieved by anti-IL-10 antibody treatment. Assessments of skin inflammation macroscopically, by histology and flow cytometry, all confirmed increased psoriatic symptoms. Interestingly, depletion of IL-10 markedly upregulated IL-23/IL-17 pathway related cytokines followed by a significant increase in IL-19 and IL-24. The up-regulation of IL-19 and IL-24, but not IL-17A, coincided with increased keratinocyte proliferation. To investigate the cellular source and effects of biologics on IL-19, human skin fibroblasts from healthy controls and psoriasis patients were cultured alone or co-cultured with activated memory CD4+ T cells. Besides IL-1β, IL-17A induced direct expression of IL-19 and IL-24 in skin fibroblasts and keratinocytes. Importantly, intrinsic higher expression of IL-19 in psoriatic skin fibroblasts was observed in comparison to healthy skin fibroblasts. Neutralization of IL-17A in the human skin fibroblast-T cell co-culture system significantly suppressed IL-19 and IL-24 expression. Together, our data show that IL-17A-induced IL-19 and IL-24 expression in skin stromal cells contribute to keratinocyte proliferation.

Major approaches of molecular and physiopathological mechanisms for hormonal treatment of melasma: a systematic review

Introduction: Epidemiological data showed that melasma occurs in 14.5% -56% of pregnant women and in 11.3% -46% of individuals who take oral contraceptives in different countries. In this respect, by inducing the synthesis of melanogenic enzymes, such as tyrosinase and proteins related to tyrosinase 1 and 2, estrogens stimulate melanogenesis in human melanocytes. The increase in progesterone levels that occurs during pregnancy and the increase in estrogen production that occurs from the eighth to the thirtieth week of pregnancy reflects the progression of hyperpigmentation. Objective: The main predictors, pathophysiological mechanisms, and experimental and clinical results of melasma were addressed through a systematic literature review, discussing the main outcomes of clinical studies. Methods: The present study followed a systematic review model. After literary search criteria using MeSH Terms, a total of 98 clinical studies were compared and submitted to eligibility analysis and, after that, 70 studies were selected, following the rules of PRISMA. The search strategy was carried out in the databases PubMed, Embase, Ovid and Cochrane Library, Web Of Science, ScienceDirect Journals (Elsevier), Scopus (Elsevier), OneFile. Results: Sex hormones, such as estrogens, especially 17-β-estradiol (E2) and progesterone, are factors involved in the regulation of pigmentation. Levels of estradiol, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were found to be higher in the serum of women with melasma. Estrogen and progesterone receptors are expressed differently in melasma skin than in perilesional or non-lesional skin. Few authors have analyzed the effects of progesterone on pigmentation, and no data has been reported with concentrations of progesterone found in non-pregnant men and women. An increase in keratinocyte proliferation was observed with baseline progesterone concentrations. It is well known that keratinocytes secrete exosomes and factors that regulate melanogenesis, differentiation, and proliferation of melanocytes. Progesterone is involved in the pathogenesis of melasma, stimulating melanogenesis in epidermal melanocytes. Conclusion: The prevention of melasma can occur by progesterone components in oral contraceptives since progesterone can reduce melanocyte proliferation without significant effects on tyrosine activity. Estrogen can increase melanogenesis in the melanocyte monolayer and induce the production of melanogenic factors by keratinocytes. Estrogen can support hyperpigmentation, increasing the number of blood vessels and, thus, stimulating the secretion of endothelin-1.

Interleukin-22 and keratinocytes; pathogenic implications in skin inflammation

Interleukin (IL)-22 is produced from immune cells such as T helper (Th)22 cells, Th17/22 cells, and group 3 innate lymphoid cells. IL-22 signals via the IL-22 receptor 1(IL-22R1) and the IL-10 receptor 2 (IL-10R2). As the IL-22R1/IL-10R2 heterodimer is preferentially expressed on border tissue between the host and the environment, IL-22 is believed to be involved in border defense. Epidermal keratinocytes are the first-line skin barrier and express IL-22R1/IL-10R2. IL-22 increases keratinocyte proliferation but inhibits differentiation. Aryl hydrocarbon receptor (AHR) is a chemical sensor and an essential transcription factor for IL-22 production. In addition, AHR also upregulates the production of barrier-related proteins such as filaggrin in keratinocytes, suggesting a pivotal role for the AHR-IL-22 axis in regulating the physiological skin barrier. Although IL-22 signatures are elevated in atopic dermatitis and psoriasis, their pathogenic and/or protective implications are not fully understood.

Functional cooperation between ASK1 and p21Waf1/Cip1 in the balance of cell-cycle arrest, cell death and tumorigenesis of stressed keratinocytes

Both CDKN1A (p21 Waf1/Cip1) and Apoptosis signal-regulating kinase 1 (ASK1) play important roles in tumorigenesis. The role of p21 Waf1/Cip1 in attenuating ASK1-induced apoptosis by various stress conditions is well established. However, how ASK1 and p21 Waf1/Cip1 functionally interact during tumorigenesis is still unclear. To address this aspect, we crossed ASK1 knockout (ASK1KO) mice with p21Waf1/Cip1 knockout (p21KO) mice to compare single and double-mutant mice. We observed that deletion of p21Waf1/Cip1 leads to increased keratinocyte proliferation but also increased cell death. This is mechanistically linked to the ASK1 axis-induced apoptosis, including p38 and PARP. Indeed, deletion of ASK1 does not alter the proliferation but decreases the apoptosis of p21KO keratinocytes. To analyze as this interaction might affect skin carcinogenesis, we investigated the response of ASK1KO and p21KO mice to DMBA/TPA-induced tumorigenesis. Here we show that while endogenous ASK1 is dispensable for skin homeostasis, ASK1KO mice are resistant to DMBA/TPA-induced tumorigenesis. However, we found that epidermis lacking both p21 and ASK1 reacquires increased sensitivity to DMBA/TPA-induced tumorigenesis. We demonstrate that apoptosis and cell-cycle progression in p21KO keratinocytes are uncoupled in the absence of ASK1. These data support the model that a critical event ensuring the balance between cell death, cell-cycle arrest, and successful divisions in keratinocytes during stress conditions is the p21-dependent ASK1 inactivation.

Evaluation of Real Time Cell Proliferation, Anti-Inflammatory and Wound Healing Potential of Helenalin on HaCaT Keratinocytes Treated with Lipopolysaccharide Stimulated Monocytes

Arnica montana, of the family Asteraceae, is used for its anti-inflammatory and wound healing properties, especially for bruises, tissue injuries and other traumas. However, its cellular and molecular mechanisms are not yet fully known. Its main active ingredient is helenalin, a sesquiterpene lactone, known as one of the main active ingredients of Asteraceae species. The aim of this study was to investigate cell proliferation, wound healing and anti-inflammatory activity of helenalin on human keratinocyte cells treated with lipopolysaccharide stimulated human monocytic cell line. Human keratinocyte cells were treated with different helenalin concentrations (0.095-50 μM) to determine toxic and nontoxic concentrations. Then, the effects of nontoxic (0.02 and 0.2 μM) and toxic (2 μM) helenalin concentrations on human keratinocyte cell proliferation and migration were determined using the real time cell analysis system. The scratch assay was performed to determine wound healing activity and supernatants were collected to determine the effects of helenalin on cytokine levels of human keratinocyte cells under inflammatory and non-inflammatory conditions. Under inflammatory conditions, 0.02 and 0.2 μM helenalin increased keratinocyte proliferation and induced the most wound healing and anti-inflammatory activity in human keratinocyte cells. Cell proliferation and migration, wound healing and anti-inflammatory effects were higher in cells exposed to 0.02 μM helenalin than 0.2 μM. Helenalin has been found to decrease the production of inflammatory cytokines, especially under inflammatory conditions and increase wound closure by increasing cell proliferation and migration.

Deletion of Mediator 1 suppresses TGFβ signaling leading to changes in epidermal lineages and regeneration.

Epidermal lineages and injury induced regeneration are controlled by transcriptional programs coordinating cellular signaling and epigenetic regulators, but the mechanism remains unclear. Previous studies showed that conditional deletion of the transcriptional coactivator Mediator 1 (Med1) changes epidermal lineages and accelerates wound re-epithelialization. Here, we studied a molecular mechanism by which Med1 facilitates these processes, in particular, by focusing on TGFβ signaling through genome wide transcriptome analysis. The expression of the TGF ligands (Tgfβ1/β2) and their downstream target genes is decreased in both normal and wounded Med1 null skin. Med1 silencing in cultured keratinocytes likewise reduces the expression of the ligands (TGFβ1/β2) and diminishes activity of TGFβ signaling as shown by decreased p-Smad2/3. Silencing Med1 increases keratinocyte proliferation and migration in vitro. Epigenetic studies using chromatin immuno-precipitation and next generation DNA sequencing reveals that Med1 regulates transcription of TGFβ components by forming large clusters of enhancers called super-enhancers at the regulatory regions of the TGFβ ligand and SMAD3 genes. These results demonstrate that Med1 is required for the maintenance of the TGFβ signaling pathway. Finally, we show that pharmacological inhibition of TGFβ signaling enhances epidermal lineages and accelerates wound re-epithelialization in skin similar to that seen in the Med1 null mice, providing new insights into epidermal regeneration.

Psoriatic Dermal-derived Mesenchymal Stem Cells Reduce Keratinocyte Junctions, and Increase Glycolysis

Although it is known that psoriatic dermal-derived mesenchymal stem cells (DMSCs) dysregulate keratinocyte proliferation, the biological activity profile of keratinocytes influenced by psoriatic DMSCs remain unknown. In the present study, we assessed the impact of psoriatic DMSCs on keratinocyte proliferation, differentiation, and glucose metabolism in normal human epidermal keratinocytes co-cultured with or without psoriatic DMSCs. Co-culture of normal human epidermal keratinocytes with psoriatic DMSCs down-regulated expression levels of proteins associated with cell junction assembly (alpha-actinin-1, catenin beta-1, poliovirus receptor-related protein 4 and procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2), while upregulating proteins associated with keratinocyte proliferation and differentiation (involucrin, isoform 2 of Histone-binding protein, isoform 3 of Telomeric repeat-binding factor 2 and keratin 13). Moreover, coculture of normal human epidermal keratinocytes with psoriatic DMSCs stimulated keratinocyte proliferation and glycolysis, but reduced keratinocyte junctions. Taken together, these results demonstrate that psoriatic DMSCs increase keratinocyte proliferation and glycolysis, and reduce cell junctions, suggesting a pathogenic role of psoriatic DMSCs in epidermal hyperplasia, aberrant differentiation, and reduction in turnover time of keratinocytes in psoriasis.