Short-form thymic stromal lymphopoietin (sfTSLP) restricts herpes simplex virus infection of human primary keratinocytes.

Eczema herpeticum (EH) is a disseminated severe herpes simplex virus type 1 (HSV-1) infection that mainly occurs in a subset of patients suffering from atopic dermatitis (AD). EH is complex and multifaceted, involving immunological changes, environmental influences, and genetic aberrations. Certain genetic variants of the thymic stromal lymphopoietin (TSLP) may predispose to develop severe HSV-1-induced eczema. Therefore, we investigated the impact of TSLP on HSV-1 infection. TSLP encodes for two distinct forms: a long-form (lfTSLP), primarily associated with type 2 immunity, and a short-form (sfTSLP) with anti-inflammatory and antimicrobial properties. While sfTSLP reduced HSV-1 infectibility in human primary keratinocytes (HPK), lfTSLP did not. In HPK treated with sfTSLP, HSV-1 gene expression, and replication decreased, while virion binding to cells and targeting of incoming capsids to the nucleus were not diminished compared to untreated cells. sfTSLP caused only minor changes in the expression of innate immunity cytokines, and its inhibition of HSV-1 infection did not require de novo protein synthesis. Time window experiments indicated a different antiviral mechanism than LL-37. sfTSLP showed the strongest antiviral effect when administered to HPK before or after inoculation with HSV-1, and outperformed the inhibitory potential of LL-37 under these conditions. Our data show that sfTSLP has antiviral functions and promotes repression of the HSV-1 infection in HPK.

Exogenous factors in the pathogenesis of atopic dermatitis: Irritants and cutaneous infections.

Atopic dermatitis (AD) is a chronic relapsing inflammatory cutaneous disease that is often associated with other atopic symptoms, such as food allergy, allergic rhinitis and asthma, leading to significant morbidity and healthcare costs. The pathogenesis of AD is complicated and multifactorial. Although the aetiology of AD remains incompletely understood, recent studies have provided further insight into AD pathophysiology, demonstrating that the interaction among genetic predisposition, immune dysfunction and environmental provocation factors contributes to its development. However, the increasing prevalence of AD suggests that environmental factors such as irritation and cutaneous infection play a crucial role in triggering and/or aggravating the disease. Of note, AD skin is susceptible to bacterial, fungal and viral infections, and microorganisms may colonize the skin and aggravate AD symptoms. Overall, understanding the mechanisms by which these risk factors affect the cutaneous immunity of patients with AD is of great importance for developing a precision medicine approach for treatment. This review summarizes recent developments in exogenous factors involved in the pathogenesis of AD, with special emphasis on irritants and microbial infections.

IL-1R Type 1-Deficient Mice Demonstrate an Impaired Host Immune Response against Cutaneous Vaccinia Virus Infection.

The IL-1 superfamily of cytokines and receptors has been studied extensively. However, the specific roles of IL-1 elements in host immunity to cutaneous viral infection remain elusive. In this study, we applied vaccinia virus (VACV) by scarification to IL-1R1 knockout mice (IL-1R1-/-) and found that these mice developed markedly larger lesions with higher viral genome copies in skin than did wild-type mice. The phenotype of infected IL-1R1-/- mice was similar to eczema vaccinatum, a severe side effect of VACV vaccination that may develop in humans with atopic dermatitis. Interestingly, the impaired cutaneous response of IL-1R1-/- mice did not reflect a systemic immune deficiency, because immunized IL-1R1-/- mice survived subsequent lethal VACV intranasal challenge, or defects of T cell activation or T cell homing to the site of inoculation. Histologic evaluation revealed that VACV infection and replication after scarification were limited to the epidermal layer of wild-type mice, whereas lack of IL-1R1 permitted extension of VACV infection into dermal layers of the skin. We explored the etiology of this discrepancy and determined that IL-1R1-/- mice contained significantly more macrophages and monocyte-derived dendritic cells in the dermis after VACV scarification. These cells were vulnerable to VACV infection and may augment the transmission of virus to adjacent skin, thus leading to larger skin lesions and satellite lesions in IL-1R1-/- mice. These results suggest new therapeutic strategies for treatment of eczema vaccinatum and inform assessment of risks in patients receiving IL-1 blocking Abs for treatment of chronic inflammatory disorders.

Ankyrin repeat domain 1 regulates innate immune responses against herpes simplex virus 1: A potential role in eczema herpeticum.

BACKGROUND: Atopic dermatitis (AD) is a common inflammatory skin disease. A subset of patients with AD are susceptible to disseminated herpes simplex virus (HSV) infection, a complication termed eczema herpeticum (ADEH+). The immune mechanisms causing ADEH+ remain elusive. Using RNA sequencing, we recently found that ankyrin repeat domain 1 (ANKRD1) was significantly induced in human PBMCs upon HSV-1 stimulation, and its induction in patients with ADEH+ was significantly reduced compared with that seen in AD patients without a history of eczema herpeticum (ADEH-). OBJECTIVE: We sought to validate ANKRD1 gene expression in nonatopic (NA) subjects, patients with ADEH-, and patients with ADEH+ and to delineate the biological function of ANKRD1 and the signaling pathway or pathways involved. METHODS: Purification of human PBMCs, monocytes, B cells, dendritic cells, T cells, and natural killer cells; RNA extraction and quantitative RT-PCR; small interfering RNA technique; co-immunoprecipitation; and Western blot assays were used. RESULTS: ANKRD1 expression was significantly reduced in PBMCs from patients with ADEH+ after HSV-1 stimulation compared with PBMCs from patients with ADEH-. We found that the induction of ANKRD1 by HSV-1 and multiple pattern recognition receptor agonists are mediated by inflammatory cytokines. Silencing ANKRD1 gene expression in antigen-presenting cells led to increased viral load and reduced IFNB1 and IL29 production. Using co-immunoprecipitation methods, we demonstrated that ANKRD1 formed protein complexes with interferon regulatory factor (IRF) 3 and IRF7, which are important transcription factors regulating signaling transduction of pattern recognition receptors. Overexpression of ANKRD1 enhanced the IRF3-mediated signaling pathways. CONCLUSION: ANKRD1 is involved in IRF3-mediated antiviral innate immune signaling pathways. Its reduced expression in patients with ADEH+ might contribute to the pathogenesis of ADEH+.

Atypical Varicella-Zoster Kaposi Varicelliform Eruption in Sézary Syndrome.

Patients with mycosis fungoides experience considerable morbidity and mortality from secondary bacterial and viral infections. Staphylococcus aureus, ß-hemolytic streptococci, herpes simplex virus, and herpes zoster virus remain the most common infectious pathogens in this group of patients. With depressed cellular immunity and diminished skin barrier as the main precipitating risk factors, immunocompromised patients can often present with an atypical presentation of a common dermatologic condition. The case herein discusses a clinically atypical nonvesicular Kaposi varicelliform eruption secondary to a varicella-zoster virus in a patient with Sézary syndrome. Concurrent polypharmacy in these patients is also a risk factor for development of drug hypersensitivity reactions. However, given their immunocompromised status, first and foremost, a careful inspection should be made of every atypical skin eruption in search of an infectious etiology, and afterward, an appropriate treatment should be promptly initiated.

Defective natural killer cell activity in a mouse model of eczema herpeticum.

BACKGROUND: Patients with atopic dermatitis (AD) are susceptible to several viruses, including herpes simplex virus (HSV). Some patients experience 1 or more episodes of a severe skin infection caused by HSV termed eczema herpeticum (EH). There are numerous mouse models of AD, but no established model exists for EH. OBJECTIVE: We sought to establish and characterize a mouse model of EH. METHODS: We infected AD-like skin lesions with HSV1 to induce severe skin lesions in a dermatitis-prone mouse strain of NC/Nga. Gene expression was investigated by using a microarray and quantitative PCR; antibody titers were measured by means of ELISA; and natural killer (NK) cell, cytotoxic T-cell, regulatory T-cell, and follicular helper T-cell populations were evaluated by using flow cytometry. The role of NK cells in HSV1-induced development of severe skin lesions was examined by means of depletion and adoptive transfer. RESULTS: Inoculation of HSV1 induced severe erosive skin lesions in eczematous mice, which had an impaired skin barrier, but milder lesions in small numbers of normal mice. Eczematous mice exhibited lower NK cell activity but similar cytotoxic T-cell activity and humoral immune responses compared with normal mice. The role of NK cells in controlling HSV1-induced skin lesions was demonstrated by experiments depleting or transferring NK cells. CONCLUSION: A murine model of EH with an impaired skin barrier was established in this study. We demonstrated a critical role of defective NK activities in the development of HSV1-induced severe skin lesions in eczematous mice.

Pathological role of regulatory T cells in the initiation and maintenance of eczema herpeticum lesions.

It remains unknown why the occurrence of eczema herpeticum (EH) caused by an extensive disseminated cutaneous infection with HSV-1 or HSV-2 is associated with the exacerbation of atopic dermatitis lesions after withdrawal of treatment. Although regulatory T cells (Tregs) limit the magnitude of HSV-specific T cell responses in mice, their role in the induction and resolution of EH has not been defined. We initially investigated the frequencies, phenotype, and function of Tregs in the peripheral blood of atopic dermatitis with EH (ADEH) patients at onset and after clinical resolution, atopic dermatitis patients without EH, and healthy controls. Tregs with the skin-homing phenotype and the activated/induced phenotype were expanded at onset and contracted upon resolution. Treg-suppressive capacity was retained in ADEH patients and, the expanded Tregs suppressed IFN-γ production from HSV-1-specific CD8(+) and CD4(+) T cells. The increased frequency of CD14(dim)CD16(+) proinflammatory monocytes (pMOs) was also observed in the blood and EH skin lesions. Thus, pMOs detected in ADEH patients at onset were characterized by an increased ability to produce IL-10 and a decreased ability to produce proinflammatory cytokines, unlike their normal counterparts. Our coculture study using Tregs and pMOs showed that the pMOs can promote the expansion of inducible Tregs. Tregs were detected frequently in the vicinity of HSV-expressing and varicella zoster virus-expressing CD16(+) monocytes in the EH lesions. Expansions of functional Tregs, together with pMOs, initially required for ameliorating excessive inflammation occurring after withdrawal of topical corticosteroids could, in turn, contribute to the initiation and progression of HSV reactivation, resulting in the onset of EH.

Filaggrin deficiency promotes the dissemination of cutaneously inoculated vaccinia virus.

BACKGROUND: Eczema vaccinatum is a life-threatening complication of smallpox vaccination in patients with atopic dermatitis (AD) characterized by dissemination of vaccinia virus (VV) in the skin and internal organs. Mutations in the filaggrin (FLG) gene, the most common genetic risk factor for AD, confer a greater risk for eczema herpeticum in patients with AD, suggesting that it impairs the response to cutaneous viral infections. OBJECTIVE: We sought to determine the effects of FLG deficiency on the response of mice to cutaneous VV inoculation. METHODS: VV was inoculated by means of scarification of unsensitized skin or skin topically sensitized with ovalbumin in FLG-deficient flaky tail (ft/ft) mice or wild-type (WT) control mice. The sizes of primary and satellite skin lesions were measured, and hematoxylin and eosin staining was performed. VV genome copy numbers and cytokine mRNA levels were measured by using quantitative PCR. RESULTS: VV inoculation in unsensitized skin of ft/ft mice, independent of the matted hair mutation, resulted in larger primary lesions, more abundant satellite lesions, heavier viral loads in internal organs, greater epidermal thickness, dermal cellular infiltration, and higher local Il17a, Il4, Il13, and Ifng mRNA levels than in WT control mice. VV inoculation at sites of topical ovalbumin application amplified all of these features in ft/ft mice but had no detectable effect in WT control mice. The number of satellite lesions and the viral loads in internal organs after cutaneous VV inoculation were significantly reduced in both unsensitized and topically sensitized ft/ftxIl17a(-/-) mice. CONCLUSION: FLG deficiency predisposes to eczema vaccinatum. This is mediated primarily through production of IL-17A.

IL-10 suppresses IL-17-mediated dermal inflammation and reduces the systemic burden of Vaccinia virus in a mouse model of eczema vaccinatum.

Individuals with atopic dermatitis (AD) are susceptible to a severe, potentially fatal, systemic infection and inflammatory response following exposure to Vaccinia virus (VV). IL-10 acts both as an inducer of Th2 responses and as a regulator of T cell activation. It has been shown to limit skin inflammation elicited by contact sensitizers. AD exacerbations have been associated with decreased IL-10 function. We used IL-10(-/-) mice to test the role of the cytokine in VV immunity. They exhibited larger primary lesions and increased cutaneous neutrophil infiltration compared to wild-type (WT) counterparts. This was associated with enhanced production of IL-17A, IL-17F and CXCL2. Paradoxically, despite intact adaptive immune responses, tissue viral burdens were increased in IL-10(-/-) mice. These findings suggest that IL-10 is important in limiting skin inflammation induced by VV and that abnormal IL-17-driven neutrophil recruitment at the primary infection site in the skin results in increased systemic viral dissemination.

Atopic dermatitis complicated by eczema herpeticum is associated with HLA B7 and reduced interferon-γ-producing CD8+ T cells.

BACKGROUND: The increased susceptibility of patients with atopic dermatitis (AD) to disseminated viral skin infections such as eczema herpeticum (ADEH+) is poorly understood. OBJECTIVES: The primary goal of the current study was to determine whether ADEH+ subjects have identifiable defects in cell-mediated immunity that reduce their ability to control viral infections. MATERIALS AND METHODS: In this study, we evaluated cytokine expression by various subsets of peripheral blood mononuclear cells from ADEH+ (n = 24) compared with AD without a history of viral infections (ADEH-) (n = 20) before and after treatment with herpes simplex virus (HSV). RESULTS: We found that interferon (IFN)-γ expression after HSV treatment was lower in the CD8+ T cells and monocytes from patients with ADEH+ compared with patients who are ADEH- or nonatopic. Given the induction of CD8+ T cells as the result of antigen presentation by human leucocyte antigen (HLA) class I, consistent with the findings described above we also found that the HLA B7 allele was significantly associated with risk of the ADEH+ phenotype (odds ratio = 1·91, P = 0·02, 125 ADEH+ and 161 ADEH- subjects). CONCLUSIONS: These data suggest that defects in viral-induced IFN-γ from CD8+ T cells contribute to the ADEH+ phenotype.

Susceptibility to vaccinia virus infection and spread in mice is determined by age at infection, allergen sensitization and mast cell status.

BACKGROUND: Patients, especially young children, with atopic dermatitis are at an increased risk of developing eczema vaccinatum, a severe reaction to the smallpox vaccine, either through direct vaccination or indirect contact with a person recently vaccinated. METHODS: Using a mouse model of infection, the severity of vaccinia-induced lesions was assessed from their appearance and viral DNA content. The response to vaccinia inoculation was assessed in young and adult mice, allergen-sensitized mice, and in mast cell-deficient mice. RESULTS: Young age, sensitization to an allergen prior to infection, and a mast cell deficit, accomplished by using mast cell-deficient mice, resulted in more severe viral lesions at the site of inoculation, according to lesion appearance and viral DNA content. All three factors combined demonstrated maximal susceptibility, characterized by the severity of primary lesions and the development of secondary (satellite) lesions, as occurs in eczema vaccinatum in humans. Resistance to the appearance of satellite lesions could be restored by adoptive transfer of bone marrow-derived mast cells from either wild-type or cathelicidin-related antimicrobial peptide-deficient mice. Primary lesions were more severe following the latter transfer, indicating that cathelicidin-related antimicrobial peptide does contribute to the protective activity of mast cells against infection. CONCLUSIONS: The combination of young age, allergen sensitization and a mast cell deficit resulted in the most severe lesions, including satellite lesions. Understanding the factors determining the relative resistance/sensitivity to vaccinia virus will aid in the development of strategies for preventing and treating adverse reactions which can occur after smallpox vaccination.

Skin inflammation arising from cutaneous regulatory T cell deficiency leads to impaired viral immune responses.

Individuals with atopic dermatitis immunized with the small pox vaccine, vaccinia virus (VV), are susceptible to eczema vaccinatum (EV), a potentially fatal disseminated infection. Dysfunction of Forkhead box P3 (FoxP3)-positive regulatory T cells (Treg) has been implicated in the pathogenesis of atopic dermatitis. To test whether Treg deficiency predisposes to EV, we percutaneously VV infected FoxP3-deficient (FoxP3(KO)) mice, which completely lack FoxP3(+) Treg. These animals generated both fewer VV-specific CD8(+) effector T cells and IFN-gamma-producing CD8(+) T cells than controls, had higher viral loads, and exhibited abnormal Th2-polarized responses to the virus. To focus on the consequences of Treg deficiency confined to the skin, we generated mixed CCR4(KO) FoxP3(KO) bone marrow (CCR4/FoxP3) chimeras in which skin, but not other tissues or central lymphoid organs, lack Treg. Like FoxP3(KO) mice, the chimeras had impaired VV-specific effector T cell responses and higher viral loads. Skin cytokine expression was significantly altered in infected chimeras compared with controls. Levels of the antiviral cytokines, type I and II IFNs and IL-12, were reduced, whereas expression of the proinflammatory cytokines, IL-6, IL-10, TGF-beta, and IL-23, was increased. Importantly, infection of CCR4/FoxP3 chimeras by a noncutaneous route (i.p.) induced immune responses comparable to controls. Our findings implicate allergic skin inflammation resulting from local Treg deficiency in the pathogenesis of EV.

Vaccinia virus inoculation in sites of allergic skin inflammation elicits a vigorous cutaneous IL-17 response.

Eczema vaccinatum (EV) is a complication of smallpox vaccination occurring in patients with atopic dermatitis. In affected individuals, vaccinia virus (VV) spreads through the skin, resulting in large primary lesions and satellite lesions, and infects internal organs. BALB/c mice inoculated with VV at sites of Th2-biased allergic skin inflammation elicited by epicutaneous ovalbumin (OVA) sensitization exhibited larger primary lesions that were erosive, more satellite lesions, and higher viral loads in skin and internal organs than mice inoculated in saline-exposed skin, unsensitized skin, or skin sites with Th1-dominant inflammation. VV inoculation in OVA-sensitized skin induced marked local expression of IL-17 transcripts and massive neutrophil infiltration compared to VV inoculation in saline-exposed skin. Treatment with anti-IL-17 decreased the size of primary lesions, numbers of satellite lesions, and viral loads. Addition of IL-17 promoted VV replication in skin explants. These results suggest that IL-17 may be a potential therapeutic target in EV.

Human atopic dermatitis complicated by eczema herpeticum is associated with abnormalities in IFN-γ response.

BACKGROUND: The basis for increased susceptibility of patients with atopic dermatitis (AD) to develop disseminated viral skin infections such as eczema herpeticum (AD with a history of eczema herpeticum, ADEH(+)) is poorly understood. OBJECTIVE: We sought to determine whether subjects with AD prone to disseminated viral skin infections have defects in their IFN responses. METHODS: GeneChip profiling was used to identify differences in gene expression of PBMCs from patients with ADEH(+) compared with patients with AD without a history of eczema herpeticum (ADEH(-)) and nonatopic controls. Key differences in protein expression were verified by enzyme-linked immunosorbent spot assay and/or ELISA. Clinical relevance was further demonstrated by a mouse model of disseminated viral skin infection and genetic association analysis for genetic variants in IFNG and IFNGR1 and ADEH among 435 cases and controls. RESULTS: We demonstrate by global gene expression analysis selective transcriptomic changes within the IFN superfamily of PBMCs from subjects with ADEH(+) reflecting low IFN-γ and IFN-γ receptor gene expression. IFN-γ protein production was also significantly lower in patients with ADEH(+) (n = 24) compared with patients with ADEH(-) (n = 20) and nonatopic controls (n = 20). IFN-γ receptor knockout mice developed disseminated viral skin infection after epicutaneous challenge with vaccinia virus. Genetic variants in IFNG and IFNGR1 single nucleotide polymorphisms (SNPs) were significantly associated with ADEH (112 cases, 166 controls) and IFN-γ production: a 2-SNP (A-G) IFNGR1 haplotype (rs10457655 and rs7749390) showed the strongest association with a reduced risk of ADEH+ (13.2% ADEH(+) vs 25.5% ADEH(-); P = .00057). CONCLUSION: Patients with ADEH(+) have reduced IFN-γ production, and IFNG and IFNGR1 SNPs are significantly associated with ADEH(+) and may contribute to an impaired immune response to herpes simplex virus.

Specificity protein 1 is pivotal in the skin's antiviral response.

BACKGROUND: Previous studies have found specificity protein (Sp) 1 transcription factor in the viral replication machinery and postulated that Sp1 was required for viral replication in host cells. OBJECTIVES: We investigated the role of Sp1 in the skin's antiviral responses from the perspective of host defense and its biological relevance in patients with atopic dermatitis and a history of eczema herpeticum (ADEH(+)). METHODS: Small interfering RNA duplexes were used to knock down Sp1 in keratinocytes. The expression of vaccinia virus (VV), herpes simplex virus 1, and other genes were evaluated by real-time PCR, or combined with Western blot and immunohistofluorescence staining. A total of 106 human subjects participated in this study. RESULTS: Both VV and herpes simplex virus 1 replication were enhanced in Sp1 knocked-down keratinocytes. Sp1 gene expression was significantly decreased in ADEH(+) subjects compared with patients with atopic dermatitis without a history of eczema herpeticum and nonatopic subjects (P < .0001) and inversely correlated with VV DNA copy number in human skin explants incubated with VV in vitro (partial correlation r = -0.256; P = .009). Gene profiling revealed that the antiviral genes, double-stranded RNA-dependent protein kinase (PKR) and 2'5'-oligoadenylate synthetase 2 (OAS2), were significantly downregulated in Sp1-silenced keratinocytes. Gene expression of PKR and OAS2 was also significantly decreased in skin biopsies from ADEH(+) subjects compared with patients with atopic dermatitis without a history of eczema herpeticum and nonatopic subjects. IFN-γ augmented the antiviral capacity of Sp1-silenced keratinocytes. CONCLUSION: Specificity protein 1 knockdown enhances viral replication in keratinocytes by downregulating gene expression of PKR and OAS2. Sp1 deficiency in ADEH(+) patients may contribute to their increased propensity to disseminated skin viral infections. IFN-γ augmentation may be a potential treatment for ADEH(+) patients.

Vaccinia virus-specific molecular signature in atopic dermatitis skin.

BACKGROUND: Eczema vaccinatum (EV), a disseminated viral skin infection, is a life-threatening complication of vaccinia virus (VV) inoculation in patients with atopic dermatitis (AD) and is thought to be associated with a defective innate immune response. However, the precise mechanism or mechanisms and key factor or factors of EV are unknown. OBJECTIVE: Given that patients with psoriasis, another inflammatory skin disorder, are not susceptible to EV, we compared the global transcriptional response of skin to VV in healthy subjects, patients with psoriasis, and patients with AD, focusing on AD-specific genes. We hypothesized that differences in the transcriptional response to VV between patients with AD and patients with psoriasis or healthy subjects would identify a defective pathway or pathways that might be associated with the development of EV. METHODS: Gene expression profiling of sham-treated and VV-treated unaffected skin explants from patients with AD (n = 12), patients with psoriasis (n = 12), or healthy subjects (n = 13) were generated with U133_Plus2 (54,613 probe sets) GeneChips and analyzed with the GCOS_1.4/SAM_2.1/MAPPFinder_2.0 pipeline. RESULTS: Sixty-seven genes were significantly affected by VV in AD skin but not in psoriatic and healthy skin. Genes associated with defense response, response to wounding, and immune response were the most affected by VV in AD skin. All genes in these ontologies were downregulated, including the innate immunity genes leukotriene B(4) receptor (LTB4R), orosomucoid 1 (ORM1), coagulation factor II (thrombin) receptor (F2R), complement component 9 (C9), and LPS-binding protein (LBP). These findings were confirmed by means of real-time PCR and validated by means of PubMatrix analysis. ORM1, Toll-like receptor 4 (TLR4), and NLR family pyrin domain containing 1 (NLRP1) genes were also linked to AD severity. CONCLUSION: This study identified groups of innate immunity genes that are associated with the aberrant response of AD skin to VV and represent potential targets for EV pathogenesis.

Recurrent herpes simplex infection in multiple locations in an otherwise healthy boy.

An 8-year-old boy developed vesicular lesions on the skin in different parts of the body, occurring four to six times a year. He had a history of eczema herpeticum as a young child. We confirmed a diagnosis of multifocal herpes simplex infection, and hypothesize that this was a result of his previous eczema herpeticum, an unusual complication, in an otherwise healthy and immunocompetent child.