Our researchers presented their work at the Society of Investigative Dermatology Meeting Annual Meeting that took place in Chicago, IL May 8-11, 2019. Please see below for awards, presentations, and abstracts.
Chelsea Handfield received 2019 Albert M Kligman/SID Travel Fellowship Award.
Chelsea Handfield presented “Activation of Trpv1 nociceptive fibers stimulates an innate antiviral immune response following skin injury” at Plenary Session
Dan Hlavaty presented “The role of Ndel1 and Nde1 in regulating keratin assembly at the desmosome” at a Minisymposium.
Wenxiu Ning presented “Mechanical regulation of stem cell proliferation and fate decisions by their differentiated daughters” at a Minisymposium.
Jutamas Suwanpradid presented “IL-27 in macrophages mediates T cell survival and dermal cluster formation in allergic contact hypersensitivity” at a Minisymposium.
Y Kim, D Hlavaty, J Maycock, T Lechler. “The role of Ndel1 and Nde1 in regulating keratin assembly at the desmosome.”
Keratin filaments are essential for the mechanical strength of the epidermis. This function requires the attachment to cell-cell adhesion structures called desmosomes. While we know the molecular basis of this attachment, whether and how keratin filament dynamics are controlled by the desmosome remains unknown. We identified the paralogs Ndel1 and Nde1 as desmosome-associated proteins whose loss in the epidermis resulted in disruption of keratin organization and desmosomal defects. We demonstrated that Ndel1 and Nde1 bind directly to keratin subunits through a conserved intermediate filament consensus motif but do not bind to assembled filaments. Targeting Ndel1 to the mitochondria resulted in reorganization of the keratin network around these organelles. Finally, we demonstrated that the binding partner of Ndel1/Nde1, Lis1, is necessary for their proper localization to the desmosomes but not for keratin filament assembly. Our result demonstrate that Ndel1 and Nde1 promote the local assembly of keratin at the desmosomes and thus are important for cell adhesion and epidermal function.
W Ning, A Muroyama, T Lechler. “Mechanical regulation of stem cell proliferation and fate decisions by their differentiated daughters.”
Differentiated cells of the epidermis have an essential role in generating the epidermal barrier, but their communication with and regulation of their stem cell progenitors is poorly understood. Here we use novel mouse transgenic lines to define roles for differentiated suprabasal cells in epidermal morphogenesis. Two mouse models demonstrate that increasing contractility of differentiated cells results in profound changes to basal cell proliferation and fate decisions. Increased proliferation resulted in hyperthickening of the epidermis and premature barrier formation. In addition, hair cell fates were inhibited. Pharmacological inhibition of the contractility pathway rescued both basal cell proliferation and hair follicle morphogenesis. These date suggest that intra-tissue tension regulates stem cell proliferation and fate decisions, similar to the known roles of extracellular matrix rigidity. Importantly, this demonstrates that differentiated cells are part of the stem cell niche that regulates development and homeostasis of the skin.
J Suwanpradid, P Hoang, J Kwock, L Floyd, J Smith, R Kedl, A Atwater, S Rajagopal, D Corcoran, AS MacLeod. “IL-27 in macrophages mediates T cell survival and dermal cluster formation in allergic contact hypersensitivity.”
Re-exposure to a relevant skin allergen initiates the efferent phase and clinical expression of allergic contact dermatitis (ACD) characterized by local DC-mediated T cell activation and recruitment of Ag-specific T cells to the sites of allergen challenge. We here make the novel observation that, in contrast to non-lesional skin of ACD patients, skin of (+) patch-tested individuals (effector phase of ACD) showed high expression of IL-15 in epidermal keratinocytes and in dermal DC-macropahge-T-cell-clusters. Interestingly, within these clusters, T cells with increased anti-apoptotic BCL2 were found directly adjacent to IL-27 producing CD14+CD86+CD172a+ cells and IL-15+ cells. Moreover, in the allergic contact hypersensitivity (CHS) model using IL-27p28eGFP mice, we found increased IL-27 production in a direct macrophage subset expressing CD172a. Functionally, IL-27 conditional knockout mice (driven by LysM-cre and CD11c-cre promoters) demonstrated suppressed DNFB-induced ear thickening (p<0.05) supporting a role for IL-27 in CHS. Mechanistically, IL-27 stimulated IL15 in human epidermal keratinocytes in a STAT-1 dependent manner as evidenced by rapid p-STAT1 nuclear translocation and abrogation of this response by silencing STAT1 (p<0.05). Given the relevance of the recently identified allergen-induced dermal T cell clusters for CHS responses, we tested the functional relevance of IL-27-induced IL-15 signaling in this context. In the CHS mouse model, administration of the IL-27-neutralizing antibody (i.d.) resulted in decreased IL-15 expression associated with dowregulation of BCL2 in T cells, the number of total cutaneous CD8+ T cells and T cell clusters (p<0.05). Similarly, when treated with recombinant IL-15, human skin T cells increased BCL2 expression (p<0.05). Overall our finding implicate IL-27 as a potential therapeutic agent in regulating cutaneous T cell immunity.
S Kirchner, X Ling, M Coates, J Shannon, P Rosa Coutinho Goulart Mariottoni, AS MacLeod, D Corcoran. “Cutaneous antiviral protein expression is regulated by the circadian clock.”
Antiviral proteins (AVPs) such as the oligoadenylate-syntase (OAS) family are a first line of innate immunity in the skin against intruding viruses. While inflammatory signaling pathways such as interferon on IL-27 can induce AVP expression, the temporal pattern of constitutive expression and induction of AVPs is unknown. We found the AVP gene expression in human keratinocytes oscillates over 24 hours. Our lab also noted decreased AVP expression in aged and young mice. It has been described that young and old individuals have disturbed circadian rhythms. The circadian genetic apparatus has a known role in cellular immunity, but it’s role in AVP expression in unknown. We hypothesized that the circadian clock plays a key role in innate antiviral immunity. Here, we show by qPCR in human keratinocytes that the expression of AVPs, including OAS2 and MX1, cycles along with circadian factors BMAL1, CLOCK, and PER2 over 24 hours. AVP peak expression correlated to peak expression of the positive circadian regulator BMAL1 and AVP expression decreased with low BMAL1 expression. This data suggests that AVP expression may be regulated by BMAL1. We examined this relationship further using Bmal1-/- mice. Biocomputational and qPCR analysis revealed that skin from Bmal1-/- mice expressed lower levels of OAS2 relative to control wild type mice. In contrast, Protein Kinase R (PKR), another AVP, was not significantly altered in Bmal1-/- mice. We conclude from these studies that the circadian clock influences expression of distinct AVPs in vitro and in vivo. Future directions for this project include determining the mechanism by which the circadian clock influences AVP expression and the functional role of this phenotype in infection. A better knowledge of this mechanism is relevant to understanding how our skin responds to viral challenges, and may help us understand differential responses to cutaneous viral infection in relation to daily timing and across ages.
P Rosa Coutinho Goulart Mariottoni, J Shannon, AS MacLeod. “Staphylococcus aureus components increase Th2 response in cutaneous lymphoma.”
Mycosis fungoides and Sezary Syndrome are the most common types of cutaneous T-cell Lymphomas (CTCL) and are often characterized by a Th2-dominant phenotype. This Th2 microenvironment is advantageous for tumor cells but may cause higher susceptibility to infections. Indeed, it has been shown that Stapylococcus aureus (S. aureus) frequently colonizes the skin and nares of CTCL patients which can result in chronic and recurrent skin and systemic infections. Furthermore, patients colonized by S. aureus who were treated with antibiotics frequently show clinical improvement of their skin lesion, suggesting a possible functional role of S. aureus in the pathogenesis and/or course of CTCL. We demonstrate that cell membrane components from S. aureus can increase Th2 cytokines production by tumor cells. Moreover, previous studies have revealed that S. aureus-derived microvesicles (MVs) carry a complex arsenal or virulence factors and toxins that allow better infection and serve host immunomodulatory functions. Furthermore, bacteria cell wall components such as peptidoglycans and lipoteichoic acid are part of the MV complex. We demonstrate that, when exposed to various S. aureus cell wall and MV components, human CTCL cell line (HUT78) increased expression of Th2 cytokines (IL-4 and IL-13) up to 20 and 80 fold, respectively (p≤0.001) and thymic stromal lymphopoeitin receptor (TSLPR) and IL7Ra by approximately five to ten fold (p≤0.05). TSLPR and IL7Ra form the heterodimeric receptor for TSLP, a cytokine able to promote Th2 differentiation in naïve T cells. These results suggest that S. aureus MV and cell wall components provide signals that may render CTCL cells more susceptible to TSLP effects. Together, our data shows that S. aureus components help CTCL to maintain a tumor-beneficial Th2 microenvironment. Understanding the role of S. aureus colonization in CTCL may contribute to better targets for future therapies.
C Handfield, J Kwock, P Hoang, AS MacLeod. “Activation of Trpv1 nociceptive fibers stimulates an innate antiviral immune response following skin injury.”
Multiple innate immune signaling pathways become activated upon skin injury in order to reestablish the antimicrobial barrier, prevent infection and excessive tissue injury. Damage to the skin barrier often elicits pain and leads to activation of cutaneous nociceptive fibers. We hypothesized that nociceptors in the skin are activated upon breach of the skin barrier and contribute to the innate antiviral response to skin injury. Here, we show that pharmacological or genetic ablation of TRPV1-mediated nociception impairs the innate antiviral response to skin injury. Using resiniferatoxin-treated mice or Trvp1-/- mice in murine skin excisional wound assays, we identified that preferential ablation of Trvp1 channels reduces the transcription of IFNb (p<0.01), IL27 (p<0.05), and multiple antiviral proteins including Oas2 and Oasl2 (p<0.05) in the skin. The neuromediator N-acetyl-galactosamine (GalNAc) is present in sensory nerves and binds with high specificity to the CD301 lectin, a marker for a subset of dendritic cells that releases IL-27 in the process of skin wound repair. We here show that human THP1-derived immature dendritic cells and Langerhans cells isolated from fresh human skin express IL27 when stimulated with GalNAc (p<0.01 and p<0.001, respectively). Given the knowledge from previous work that cathelicidin LL-37 is produced within the skin following injury and complexes with nucleic acids to enhance production to type I IFNs, we tested the possibility that LL-37 could also alter GalNAc signaling. We demonstrate that LL-37 with GalNAc significantly enhances IL27 (p<00.1), IFNa4 (p<0.01) and IFNb1 (p<0.01) production. Together, our data suggest that nociception promotes skin antiviral competence through activation of antiviral signaling pathways upon wounding. Better understanding or the role of pain and itch in stimulating antimicrobial peptide and protein expression and induction following injury is relevant for improving skin repair.
MJ Whitley, C Lai, J Suwanpradid, RR Rudolph, D Zelac, W Havran, J Cook, D Erdmann, H Levinson, EP Healy, AS MacLeod. “UV-induced CD39 expression on immunosuppressive memory T cells in human cutaneous squamous cell carcinoma.”
UV radiation and immunosuppression are two major risk factors for cutaneous squamous cell carcinoma (cSCC). Previous studies suggest a role for regulatory T cells in the pathogenesis of cSCC. We have shown that purinergic signaling plays a role in DNA damage repair (DDR). To investigate whether immunosuppressive T cells function to suppress DDR via purinergic signaling, we queried the expression of CD39 with human cSCC. CD39 catalyzes the conversion of extracellular ATP to ADP, leaning to elevated extracellular AMP and adenosine (ADO) levels. We found that CD39 expression is increased on Tregs with human cSCCs when compared to T cells isolated from blood or non-lesional skin (p<0.05). Accordingly, the concentrations of extracellular ADP, AMP, and ADO are increased in tumors compared to normal skin (p<0.05). We find that increased ADO downregulates the expression of nucleosome assembly protein, an important component of DDR. We then examined the role of IL27, a proposed mediator of CD39 expression. Using a murine IL27 receptor KO model, we show that UV-induced CD39 expression is IL27-dependent and IL27 expression blocks UV-induced DDR in an in vitro keratinocyte model. In a mouse model of UV-induced cSCCs. Inhibition of CD39 with POM1 limits early tumor growth. Interestingly, CD39 expression is significantly higher in human cSCCs that metastasize compared to those that are non-metastatic (p<0.01). Together, these data suggest a role for IL27-mediated CD39 upregulation on Tregs to mitigate effective DDR, promoting carcinogenesis and metastasis. This serves to further elucidate the mechanisms by which the immune system regulates carcinogenesis and provides several potential targets for therapeutic advancements.
M Coates, D Corcoran, P Rosa Coutinho Goulart Mariottoni, T Jaleel, D Brown, J Murray, M Morasso, AS MacLeod. “The skin transcriptome in hidradenitis Suppurativa uncovers an antimicrobial and sweat gland gene signature that has distinct overlap with wounded skin.”
Hidradenitis Suppurativa (HS) is characterized by recurrent abscesses and sinus tract formation keading to chronic non-healing wounds. We analyzed microarray gene expression of HS lesional and non-lesional skin samples. Differentially expressed genes (DEGs) in HS lesional vs. non-lesional skin were also compared to DEGs in punch-biopsy wounded vs. normal skin. A number of antimicrobial peptides and proteins (AMP), including members of the S100 family (log2FC=4.85, p<0.001), defensins (log2FC=3.75, p<0.001), and interferon stimulated antiviral genes, such as oligoadenylate synthetase 2 (log2FC=1.88, p<0.001), were significantly upregulated in both HS lesional skin and wounded skin. In contrast, the AMP dermcidin (DCD) was among the most down-regulated genes in HS lesional skin (log2FC=-4.90, p<0.001), but was up-regulated in wounded skin. Findings were confirmed via qPCR (log2FC=-3.57, p<0.05) and immnunofluorescence, which showed decreased expression of DCD in eccrine sweat glands of HS lesional skin, compared to HS non-lesional and normal skin. Other sweat gland-associated genes, such as secretoglobins (log2FC=-4.64, p<0.001) and aquaporin 5 (log2FC=-1.69, p<0.01), also had decreased expression in HS lesional skin and increased expression in wounded skin. The increased expression of many AMPs in HS lesional skin and wounded skin suggests a similar inflammatory environment in both conditions. Conversely, our discovery that sweat-gland associated proteins, such as DCD, were decreased in HS lesional skin but not in wounded skin suggests that impaired sweat gland function may be a unique pathological feature of HS.