Recommendations for rosacea diagnosis, classification and management: update from the global ROSacea COnsensus 2019 panel.

BACKGROUND: A transition from a subtyping to a phenotyping approach in rosacea is underway, allowing individual patient management according to presenting features instead of categorization by predefined subtypes. The ROSacea COnsensus (ROSCO) 2017 recommendations further support this transition and align with guidance from other working groups. OBJECTIVES: To update and extend previous global ROSCO recommendations in line with the latest research and continue supporting uptake of the phenotype approach in rosacea through clinical tool development. METHODS: Nineteen dermatologists and two ophthalmologists used a modified Delphi approach to reach consensus on statements pertaining to critical aspects of rosacea diagnosis, classification and management. Voting was electronic and blinded. RESULTS: Delphi statements on which the panel achieved consensus of ≥ 75% voting 'Agree' or 'Strongly agree' are presented. The panel recommends discussing disease burden with patients during consultations, using four questions to assist conversations. The primary treatment objective should be achievement of complete clearance, owing to previously established clinical benefits for patients. Cutaneous and ocular features are defined. Treatments have been reassessed in line with recent evidence and the prior treatment algorithm updated. Combination therapy is recommended to benefit patients with multiple features. Ongoing monitoring and dialogue should take place between physician and patients, covering defined factors to maximize outcomes. A prototype clinical tool (Rosacea Tracker) and patient case studies have been developed from consensus statements. CONCLUSIONS: The current survey updates previous recommendations as a basis for local guideline development and provides clinical tools to facilitate a phenotype approach in practice and improve rosacea patient management. What's already known about this topic? A transition to a phenotype approach in rosacea is underway and is being recommended by multiple working groups. New research has become available since the previous ROSCO consensus, necessitating an update and extension of recommendations. What does this study add? We offer updated global recommendations for clinical practice that account for recent research, to continue supporting the transition to a phenotype approach in rosacea. We present prototype clinical tools to facilitate use of the phenotype approach in practice and improve management of patients with rosacea.

Calcitonin gene-related peptide: key regulator of cutaneous immunity.

Calcitonin gene-related peptide (CGRP) has been viewed as a neuropeptide and vasodilator. However, CGRP is more appropriately thought of as a pleiotropic signalling molecule. Indeed, CGRP has key regulatory functions on immune and inflammatory processes within the skin. CGRP-containing nerves are intimately associated with epidermal Langerhans cells (LCs), and CGRP has profound regulatory effects on Langerhans cell antigen-presenting capability. When LCs are exposed to CGRP in vitro, their ability to present antigen for in vivo priming of naïve mice or elicitation of delayed-type hypersensitivity is inhibited in at least some situations. Administration of CGRP intradermally inhibits acquisition of immunity to Th1-dominant haptens applied to the injected site while augmenting immunity to Th2-dominant haptens, although the cellular targets of activity in these experiments remain unclear. Although CGRP can be a pro-inflammatory agent, several studies have demonstrated that administration of CGRP can inhibit the elicitation of inflammation by inflammatory stimuli in vivo. In this regard, CGRP inhibits the release of certain chemokines by stimulated endothelial cells. This is likely to be physiologically relevant as cutaneous blood vessels are innervated by sensory nerves. Exciting new studies suggest a significant role for CGRP in the pathogenesis of psoriasis and, most strikingly, that CGRP inhibits the ability of LCs to transmit the human immunodeficiency virus 1 to T lymphocytes. A more complete understanding of the role of CGRP in the skin immune system may lead to new and novel approaches for the therapy of immune-mediated skin disorders.

What is the physiological function of mast cells?

Under physiological conditions, skin mast cells preferentially localize around nerves, blood vessels and hair follicles. This observation, which dates back to Paul Ehrlich, intuitively suggests that these enigmatic, multifacetted protagonists of natural immunity are functionally relevant to many more aspects of tissue physiology than just to the generation of inflammatory and vasodilatory responses to IgE-dependent environmental antigens. And yet, for decades, mainstream-mast cell research has been dominated by a focus on the -undisputedly prominent and important - mast cell functions in type I immune responses and in the pathogenesis and management of allergic diseases. Certainly, it is hard to believe that the very large and rather selectively distributed number of mast cells in normal, uninflamed, non-infected, non-traumatized mammalian skin or mucosal tissue simply hanging around there lazily day and night, just wait for the odd allergen or parasite-associated antigen to come by so the mast cell can finally swing into action. Indeed, the past decade has witnessed a renaissance of mast cell research 'beyond allergy', along with a more systematic exploration of the surprisingly wide range of physiological functions that mast cells may be involved in. The current debate sketches many exciting horizons that have recently come into our vision during this intriguing, ongoing search.

Langerhans cell expression of neuropeptide Y and peptide YY.

Neuropeptide Y (NPY) and peptide YY (PYY) are structurally related peptides with a variety of known functions. The role of these peptides in the skin is largely unknown, although NPY-like immunoreactivity has been reported in the epidermis. The recent report that these peptides have antimicrobial properties suggests that NPY and PYY may contribute to the skin's defense mechanisms against invading microorganisms. We have demonstrated that Langerhans cells (LC) and a certain BALB/c epidermis-derived dendritic cell line contain mRNA for NPY and PYY using RT-PCR. Furthermore, this dendritic cell line as well as an epidermis-derived dendritic cell line from A/J mice were found to produce NPY and PYY and LC produced PYY, as assessed by radioimmunoassay. These data suggest that the protective function of LC include not only antigen presentation, but also production of antimicrobial peptides.

IL-12 prevents the inhibitory effects of cis-urocanic acid on tumor antigen presentation by Langerhans cells: implications for photocarcinogenesis.

UV radiation induces skin cancer primarily by its DNA-damaging properties, but also by its capacity to suppress the immune system. The photoisomer of urocanic acid (UCA), cis-UCA, is an important mediator of UV-induced immunosuppression and is involved in the inhibition of tumor immunity. The immunomodulatory cytokine IL-12 is known to counteract many of the immunosuppressive effects of UV radiation, including UV-induced immune tolerance. In this study, we addressed whether IL-12 also reverts the immunosuppressive activities of cis-UCA. Cis-UCA inhibits the ability of Langerhans cells to present tumor Ags for primary and secondary tumor immune responses. IL-12 treatment completely prevented the suppression by cis-UCA. IL-12 also protected mice from cis-UCA-induced suppression of contact hypersensitivity responses. To study the effects of cis-UCA on Ag-processing and Ag-presenting function in vitro, Langerhans cells were treated with UCA isomers and incubated with OVA or OVA peptide(323-339) before exposure to OVA-specific transgenic T cells. Cis-, but not trans-UCA suppressed Ag presentation, which was completely reversed upon addition of IL-12. Since these findings suggest that cis-UCA may play an important role in photocarcinogenesis by inhibiting a tumor immune response, mice were chronically UVB irradiated to induce skin cancer. Whereas all mice in the control groups developed tumors, mice treated with a mAb with specificity for cis-UCA showed a significantly reduced tumor incidence. These data strongly indicate the importance of cis-UCA during photocarcinogenesis and support the concept of counteracting cis-UCA as an alternative strategy to prevent UV-induced skin cancer, possibly via the application of IL-12.

Stress-induced changes in skin barrier function in healthy women.

Despite clear exacerbation of several skin disorders by stress, the effect of psychologic or exertional stress on human skin has not been well studied. We investigated the effect of three different stressors, psychologic interview stress, sleep deprivation, and exercise, on several dermatologic measures: transepidermal water loss, recovery of skin barrier function after tape stripping, and stratum corneum water content (skin conductance). We simultaneously measured the effects of stress on plasma levels of several stress-response hormones and cytokines, natural killer cell activity, and absolute numbers of peripheral blood leukocytes. Twenty-five women participated in a laboratory psychologic interview stress, 11 women participated in one night of sleep deprivation, and 10 women participated in a 3 d exercise protocol. The interview stress caused a delay in the recovery of skin barrier function, as well as increases in plasma cortisol, norepinephrine, interleukin-1beta and interleukin-10, tumor necrosis factor-alpha, and an increase in circulating natural killer cell activity and natural killer cell number. Sleep deprivation also decreased skin barrier function recovery and increased plasma interleukin-1beta, tumor necrosis factor-alpha, and natural killer cell activity. The exercise stress did not affect skin barrier function recovery, but caused an increase in natural killer cell activity and circulating numbers of both cytolytic T lymphocytes and helper T cells. In addition, cytokine responses to the interview stress were inversely correlated with changes in barrier function recovery. These results suggest that acute psychosocial and sleep deprivation stress disrupts skin barrier function homeostasis in women, and that this disruption may be related to stress-induced changes in cytokine secretion.

RNA as a tumor vaccine: a review of the literature.

Many approaches have been attempted to harness the host immune system to act against malignant tumors. These have included animal and clinical trials with agents to non-specifically boost immunity, factors to augment specific immunity, transfer of lymphokine-activated killer cells and transfer of expanded populations of tumor-infiltrating lymphocytes. Therapeutic vaccination strategies have been employed using tumor extracts, purified tumor antigens, recombinant peptide tumor antigens and specific DNA sequences coding for a tumor antigen (genetic vaccination) both through direct administration to the host and by administration of antigen presenting cells exposed to these materials ex vivo. Recently, the use of RNA has been proposed for use in tumor vaccination protocols. The use of RNA has several potential advantages. Since total cellular RNA or mRNA can be utilized, it is not necessary to know the molecular nature of the putative tumor antigen(s). RNA can be effectively amplified; thus, unlike tumor-extract vaccines, only a small amount of tumor is needed to prepare the material for vaccination. Also, unlike DNA-based vaccines, there is little danger of incorporation of RNA sequences into the host genome. The possible utility of RNA-based vaccines for tumor immunotherapy should be further explored to determine whether such approaches are clinically useful.

Tumor antigen presentation by dermal antigen-presenting cells.

Several phenotypes of antigen-presenting cells are present in the dermis, where they presumably function to present encountered antigens for immune responses. This study examined the ability of dermal antigen-presenting cells to present tumor-associated antigens for the induction of in vivo antitumor immunity. Total murine dermal cells were exposed either to medium alone or to medium containing tumor-associated antigens from S1509a tumor cells. Subsequently, dermal cells were injected subcutaneously at weekly intervals into naïve mice for a total of three immunizations. One week following the final immunization, mice were challenged with living tumor cells. In these experiments, dermal cells pulsed with tumor-associated antigens induced protective immunity to tumor growth. Dermal cells exposed to tumor-associated antigens were also able to elicit delayed-type hypersensitivity after footpad injection into mice previously immunized against S1509a tumor cells. The ability to present tumor-associated antigens for both induction of antitumor immunity and elicitation of delayed-type hypersensitivity was dependent on I-A+ cells and was genetically restricted. Finally, dermal cells tended towards eliciting a greater antitumor delayed-type hypersensitivity response than epidermal cells. These results show that the murine dermis contains antigen-presenting cells capable of processing S1509a tumor antigens for the generation of protective antitumor immunity in vivo.

Induction of anti-tumor immunity with epidermal cells pulsed with tumor-derived RNA or intradermal administration of RNA.

The skin is well-suited to serve as a substrate for vaccination strategies. In this regard, epidermal cells exposed to granulocyte-macrophage colony-stimulating factor can, upon subcutaneous injection into naïve mice, present a soluble extract of tumor as a source of tumor-associated antigens for the induction of in vivo anti-tumor immunity. Use of RNA for immunization has a potential advantage over this technique. Because RNA can be amplified, only a small amount of tumor is needed for antigen preparation and, as with a soluble extract, it is not necessary to know the molecular nature of the antigen(s) relevant to immunity. To test the hypothesis that RNA-pulsed epidermal cells can induce anti-tumor immunity, total cellular RNA was isolated from the S1509a spindle cell tumor and used to pulse CAF1 epidermal cells enriched for Langerhans cell content and pre-exposed to granulocyte-macrophage colony-stimulating factor. These cells were then injected subcutaneously into naïve CAF1 mice three times at weekly intervals followed by challenge with living S1509a cells. Tumor growth was significantly less than in control animals immunized in an identical fashion but with irrelevant RNA. Digestion of S1509a RNA with RNase prior to pulsing of epidermal cells prevented the development of immunity. In separate experiments, intradermal injection of S1509a RNA into naïve mice three times at weekly intervals also induced immunity to challenge with the tumor. Digestion of S1509a RNA with RNase also prevented development of immunity in this system. Effective anti-tumor immunity can be induced in mice utilizing RNA-pulsed epidermal cells for in vivo immunization or by injecting RNA intradermally into naïve mice.

Granulocyte-macrophage colony-stimulating factor gene transfer to dendritic cells or epidermal cells augments their antigen-presenting function including induction of anti-tumor immunity.

Dendritic antigen-presenting cells derived from epidermis (Langerhans cells), bone marrow, and peripheral blood can present a wide variety of antigens, including tumor-associated antigens, for various immune responses. The development and function of dendritic cells is dependent upon a number of cytokines including granulocyte-macrophage-colony-stimulating factor. For example, Langerhans cells can present tumor-associated antigens for the induction of substantial in vivo anti-tumor immunity but only after activation in vitro by granulocyte-macrophage-colony-stimulating factor. Thus, we reasoned that insertion of a cDNA for granulocyte-macrophage-colony-stimulating factor into dendritic antigen-presenting cells may allow for autocrine stimulation and increased antigen-presenting capability. To test this possibility, we utilized an adenovirus vector to insert a cDNA for murine granulocyte-macrophage-colony-stimulating factor into the dendritic cell lines XS52-4D and XS106 (derived from neonatal mouse epidermis), bone marrow-derived dendritic cells, and epidermal cells that contain Langerhans cells. Infection of each of these cell types resulted in release of abundant quantities of granulocyte-macrophage-colony-stimulating factor. XS52-4D and XS106 cells infected with adenovirus granulocyte-macrophage-colony-stimulating factor exhibited prolonged dendrites and greater expression of major histocompatibility complex class II molecules and CD86 compared with cells infected with a null vector. Granulocyte-macrophage-colony-stimulating factor cDNA-containing XS cells, bone marrow-derived dendritic cells, and epidermal cells had more potent alloantigen presenting capability than cells infected with a null vector. Most importantly, granulocyte-macrophage-colony-stimulating factor gene-transferred epidermal cells were able to present tumor-associated antigens for in vivo anti-tumor immunity against challenge with the S1509a spindle-cell tumor whereas null vector-infected cells were unable to prime for immunity. These results suggest that introduction of a cDNA for granulocyte-macrophage-colony-stimulating factor into dendritic cells may be an effective means to augment their antigen-presenting capability and that granulocyte-macrophage-colony-stimulating factor gene-transfer- red epidermal cells may be useful in tumor vaccination strategies.

Transfection of immature murine bone marrow-derived dendritic cells with the granulocyte-macrophage colony-stimulating factor gene potently enhances their in vivo antigen-presenting capacity.

Ag presentation by dendritic cells (DC) is crucial for induction of primary T cell-mediated immune responses in vivo. Because DC culture from blood or bone marrow-derived progenitors is now clinically applicable, this study investigated the effectiveness of in vitro-generated murine bone marrow-derived DC (Bm-DC) for in vivo immunization protocols. Previous studies demonstrated that GM-CSF is an essential growth and differentiation factor for DC in culture and that in vivo administration of GM-CSF augments primary immune responses, which renders GM-CSF an attractive candidate to further enhance the effectiveness of DC-based immunotherapy protocols. Therefore, immature Bm-DC were transiently transfected with the GM-CSF gene and tested for differentiation, migration, and Ag-presenting capacity in vitro and in vivo. In vitro, GM-CSF gene-transfected Bm-DC were largely unaltered with regard to MHC and costimulatory molecule expression as well as alloantigen or peptide Ag-presenting capacity. When used for in vivo immunizations, however, the Ag-presenting capacity of GM-CSF gene-transfected Bm-DC was greatly enhanced compared with mock-transfected or untransfected cells, as determined by their effectiveness to induce primary immune reactions against hapten, protein Ag, and tumor Ag, respectively. Increased effectiveness in vivo correlated with the better migratory capacity of GM-CSF gene-transfected Bm-DC. These results show that GM-CSF gene transfection significantly enhances the capacity of DC to induce primary immune responses in vivo, which might also improve DC-based vaccines currently under clinical investigation.

beta-Endorphin binding and regulation of cytokine expression in Langerhans cells.

Neuropeptides and neurohormones have been shown to be able to regulate cutaneous immune reactions. Binding of beta-endorphin (beta-end) on epidermal Langerhans cells (LC) and effects of beta-end on cytokine expression were examined. Biotinylated beta-end bound to the mouse LC-like cell line, XS52, and the binding was replaced with intact beta-end but not with substance P. beta-End augmented secretion of IL-1 beta and IL-10 from XS52 cells were induced by a combination of LPS and GM-CSF. Induction of TNF alpha was suppressed by beta-end. The regulation of cytokine expression was confirmed in fresh LC by RT-PCR. These results suggest that beta-end is a regulator of skin immune function.

Modification of LC phenotype and suppression of contact hypersensitivity response by stress.

BACKGROUND: Stress is thought to exacerbate a number of diseases, some of which are skin disorders. Epidermal Langerhans' cells play a major role in cutaneous immune reactions. OBJECTIVE: The effects of two types of stress on the cutaneous immune system were to be assessed in mice. METHODS: Mice received stress by immobilization or housing at various population densities. Epidermal sheets were stained for I-A molecules (a member of class II major histocompatibility complex) and analyzed with a confocal-laser- scanning microscope. Contact hypersensitivity reaction to 2,4,6-trinitrochlorobenzene was elicited in mouse ears. RESULTS: The cell density, intensity of I-A expression, and number of dendrites were decreased as the population density increased. Elicitation of contact hypersensitivity was suppressed in mice that received either population or immobilization stress. Increased I-A expression and number of dendrites were observed in adrenalectomized compared to sham-operated mice. The population-dependent suppression of contact hypersensitivity reaction was not observed in adrenalectomized mice. After incubation with serum from mice that received either immobilization stress or population stress, the expression of I-A molecules on a XS52 Langerhans' cell-like cell line was reduced. CONCLUSION: Stress affected the cutaneous immune system. There were indications that adrenergic hormones played a role in the regulation of the system.

Differential regulation of epidermal cell tumor-antigen presentation by IL-1alpha and IL-1beta.

IL-1 exists in two forms, termed IL-1alpha and IL-1beta, which exert similar effects in a number of biologic models. Recently, there have been reports of some differences in the activities of these two species in some systems. To address this issue with regard to Langerhans cells, Langerhans cell-enriched preparations of epidermal cells were treated with either IL-1alpha or IL-1beta before pulsing with S1509a tumor-associated antigens and subsequent use for immunization of naive mice to S1509a. While epidermal cells treated with 100 U IL-1beta per ml were able to induce protective tumor immunity (as indicated by the rejection of a subsequent tumor challenge with viable S1509a tumor cells), epidermal cells treated with 100 U IL-1alpha per ml failed to confer protective immunity. At 1000 U per ml, IL-1beta also inhibited the ability of epidermal cells to induce tumor immunity. To investigate the effects of the two IL-1 forms on elicitation of tumor immunity, naive mice were immunized against the S1509a tumor by s.c. injection of dead S1509a cells. Epidermal cells enriched for Langerhans cells were treated with either 100 U IL-1alpha or IL-1beta per ml before tumor-associated antigens-pulsing. Epidermal cells were then washed and injected into a hind footpad of tumor immune mice and 24 h footpad swelling was assessed as a measure of delayed-type hypersensitivity. Exposure to IL-1alpha led to suppressed elicitation of delayed-type hypersensitivity, whereas IL-1beta treated epidermal cells elicited a normal (100 U per ml) or enhanced (1000 U per ml) level of delayed-type hypersensitivity. Previous experiments indicated that the suppressive effects of IL-1alpha on induction of immunity may be mediated by TNF alpha. Therefore, the ability of IL-1alpha or IL-1beta to induce epidermal cell production of TNF alpha was assessed. IL-1alpha induced epidermal cells to secrete significantly higher amounts of TNF alpha protein compared with stimulation with IL-1beta. IL-1alpha and IL-1beta appear to differentially regulate epidermal cell antigen presenting capability.

Delayed wound healing and disorganized neovascularization in transgenic mice expressing the IP-10 chemokine.

IP-10 is a member of the alpha or cysteine-X amino acid-cysteine (CXC) chemokine family of chemotactic cytokines. High levels of IP-10 expression have been detected in a number of chronic human inflammatory conditions, including psoriasis, a common inflammatory disease of the skin. IP-10 has been shown to chemoattract activated T cells, inhibit the proliferation of endothelial cells, and inhibit the growth of tumors in vivo. To determine the capacity of IP-10 to modulate the inflammatory response in vivo, we have created transgenic mice that constitutively express IP-10 from keratinocytes. These mice developed normally and, in general, did not spontaneously recruit leukocytes into the skin or other organs that expressed the transgene. In addition, the transgenic mice had a normal cutaneous contact hypersensitivity cellular immune response. However, IP-10 transgenic mice had an abnormal wound healing response characterized by a more intense inflammatory phase and a prolonged and disorganized granulation phase with impaired blood vessel formation. These results have demonstrated that IP-10 can inhibit the neovascularization associated with a physiological response in vivo and have revealed a novel biologic activity of IP-10 as an inhibitor of wound healing.

Neuropeptides and Langerhans cells.

The immune system and nervous system are intimately related. In addition to neuroendocrine mechanisms, neuropeptides have a variety of effects on immune cells and are responsible at least in part for neurogenic inflammation. The presence of neuropeptides in the skin has been well documented. The influence of neuropeptides on Langerhans cells is the focus of this paper. The physical presence and effects of calcitonin gene-related peptide on Langerhans cells is emphasized. Discussion also includes the putative inflammatory and immunologic roles of vasoactive intestinal peptide, substance P, neurotensin, neuropeptide Y, and somatostatin in the skin.