Interleukin (IL)-36α, IL-36β, and IL-36γ are innate mediators of acute epithelial inflammation. We sought to demonstrate that these cytokines are also required for the pathogenesis of plaque ...psoriasis, a common and chronic skin disorder, caused by abnormal T helper 17 (T
17) cell activation. To investigate this possibility, we first defined the genes that are induced by IL-36 cytokines in primary human keratinocytes. This enabled us to demonstrate a significant IL-36 signature among the transcripts that are up-regulated in plaque psoriasis and the susceptibility loci associated with the disease in genome-wide studies. Next, we investigated the impact of in vivo and ex vivo IL-36 receptor blockade using a neutralizing antibody or a recombinant antagonist. Both inhibitors had marked anti-inflammatory effects on psoriatic skin, demonstrated by statistically significant reductions in IL-17 expression, keratinocyte activation, and leukocyte infiltration. Finally, we explored the potential safety profile associated with IL-36 blockade by phenotyping 12 individuals carrying knockout mutations of the IL-36 receptor gene. We found that normal immune function was broadly preserved in these individuals, suggesting that IL-36 signaling inhibition would not substantially compromise host defenses. These observations, which integrate the results of transcriptomics and model system analysis, pave the way for early-stage clinical trials of IL-36 antagonists.
Human skin immune homeostasis, and its regulation by specialized subsets of tissue-residing immune sentinels, is poorly understood. In this study, we identify an immunoregulatory tissue-resident ...dendritic cell (DC) in the dermis of human skin that is characterized by surface expression of CD141, CD14, and constitutive IL-10 secretion (CD141(+) DDCs). CD141(+) DDCs possess lymph node migratory capacity, induce T cell hyporesponsiveness, cross-present self-antigens to autoreactive T cells, and induce potent regulatory T cells that inhibit skin inflammation. Vitamin D(3) (VitD3) promotes certain phenotypic and functional properties of tissue-resident CD141(+) DDCs from human blood DCs. These CD141(+) DDC-like cells can be generated in vitro and, once transferred in vivo, have the capacity to inhibit xeno-graft versus host disease and tumor alloimmunity. These findings suggest that CD141(+) DDCs play an essential role in the maintenance of skin homeostasis and in the regulation of both systemic and tumor alloimmunity. Finally, VitD3-induced CD141(+) DDC-like cells have potential clinical use for their capacity to induce immune tolerance.
Abstract
The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that regulates skin homeostasis. AHR activation by physiological ligands (e.g. indoles originated from host or ...microbial metabolism of tryptophan) leads to the expression of cytochrome P450 (CYP) enzymes, such as CYP1A1 and has anti-inflammatory effects in animal models and clinical samples of psoriasis and atopic dermatitis. Manuka honey (MH) is a complex nutritional material with antimicrobial, antioxidant and anti-inflammatory activity. We have previously shown that MH downregulates interleukin (IL)-4-induced CCL26 expression in immortalized keratinocytes. Recent studies suggest that MH contains potential AhR ligands, including indoles. Thus, we hypothesize that its anti-inflammatory effect may be mediated, at least partially, via AhR activation. Here, we treated HaCaT keratinocytes, either stably transfected with an empty vector (EV-HaCaT) or in which AhR had been stably silenced (AhR-silenced HaCaT), or primary normal human epithelial keratinocytes (NHEK) with 2% MH for 24 h and measured gene expression by reverse transcriptase quantitative polymerase chain reaction. We detected a 15.4-fold upregulation of CYP1A1 in EV-HaCaTs, which was significantly reduced (P ≤ 0.001) in AhR-silenced cells. Pretreatment with the AhR antagonist CH223191 significantly (P ≤ 0.0001) abrogated this effect. Similar findings were observed in NHEKs. In vivo epicutaneous treatment of Cyp1a1Cre × R26ReYFP reporter mice with pure MH significantly (P < 0.0001) induced CYP1A1 expression compared with Vaseline. Treatment of HaCaT with 2% MH significantly decreased (P < 0.0001) baseline CYP1A1 enzymatic activity at 3 and 6 h but increased it after 12 h, suggesting that MH may activate the AhR through both direct and indirect means. Importantly, MH downregulation of IL-4-induced CCL26 mRNA and protein was abrogated in AhR-silenced HaCaTs and by pretreatment with CH223191. Finally, MH significantly (P < 0.0001) upregulated filaggrin (FLG) expression in NHEKs in an AhR-dependent manner. In conclusion, MH activates AhR, both in vitro and in vivo, thereby providing a mechanism of its IL-4-induced CCL26 downregulation and upregulation of FLG expression. Taken together, our data show that the AhR-activating properties of MH could have broader clinical implications in a range of skin inflammatory conditions.
γδ T cells mediate rapid tissue responses in murine skin and participate in cutaneous immune regulation including protection against cancer. The role of human γδ cells in cutaneous homeostasis and ...pathology is characterized poorly. In this study, we show in vivo evidence that human blood contains a distinct subset of proinflammatory cutaneous lymphocyte Ag and CCR6-positive Vγ9Vδ2 T cells, which is rapidly recruited into perturbed human skin. Vγ9Vδ2 T cells produced an array of proinflammatory mediators including IL-17A and activated keratinocytes in a TNF-α- and IFN-γ-dependent manner. Examination of the common inflammatory skin disease psoriasis revealed a striking reduction of circulating Vγ9Vδ2 T cells in psoriasis patients compared with healthy controls and atopic dermatitis patients. Decreased numbers of circulating Vγ9Vδ2 T cells normalized after successful treatment with psoriasis-targeted therapy. Taken together with the increased presence of Vγ9Vδ2 T cells in psoriatic skin, these data indicate redistribution of Vγ9Vδ2 T cells from the blood to the skin compartment in psoriasis. In summary, we report a novel human proinflammatory γδ T cell involved in skin immune surveillance with immediate response characteristics and with potential clinical relevance in inflammatory skin disease.
Studies in humans and animal models suggest that interleukin-18 (IL-18) plays a crucial role in vascular pathologies. IL-18 is a predictor of cardiovascular death in angina and is involved in ...atherotic plaque destabilization. Higher IL-18 plasma levels also are associated with restenosis after coronary artery angioplasty performed in patients with acute myocardial infarction. We investigated the effective role of IL-18 in neointimal formation in a balloon-induced rat model of vascular injury.
Endothelial denudation of the left carotid artery was performed by use of a balloon embolectomy catheter. Increased expression of IL-18 and IL-18Ralpha/beta mRNA was detectable in carotid arteries from days 2 to 14 after angioplasty. The active form of IL-18 was highly expressed in injured arteries. Strong immunoreactivity for IL-18 was detected in the medial smooth muscle cells at days 2 and 7 after balloon injury and in proliferating/migrating smooth muscle cells in neointima at day 14. Moreover, serum concentrations of IL-18 were significantly higher among rats subjected to vascular injury. Treatment with neutralizing rabbit anti-rat IL-18 immunoglobulin G significantly reduced neointimal formation (by 27%; P < 0.01), reduced the number of proliferating cells, and inhibited interferon-gamma, IL-6, and IL-8 mRNA expression and nuclear factor-kappaB activation in injured arteries. In addition, in vitro data show that IL-18 affects smooth muscle cell proliferation.
These results identify a critical role for IL-18 in neointimal formation in a rat model of vascular injury and suggest a potential role for IL-18 neutralization in the reduction of neointimal development.
The glucocorticoid receptor (GR) and peroxisome proliferator-activated receptors (PPARs) play important roles in both physiological and pathological conditions such as cell differentiation, ...lipolysis, control of glucose metabolism, immunity, and inflammation. In fact, recent studies suggest that the thiazolidinedione (TZD) class of PPAR-gamma ligands, like glucocorticoids, may also be clinically beneficial in several inflammatory diseases, even if the molecular mechanisms responsible for these activities have not yet been clarified. In this study, by using a murine model of inflammation, the carrageenin-induced paw edema in mouse, we show that the anti-inflammatory activity exhibited by the PPAR-gamma agonists rosiglitazone and ciglitazone is reversed by the GR antagonist RU486 (17 beta-hydroxy-11 beta-4-dimethylamino phenyl-17 alpha-1-propynylestra-4,9-dien-3-one). Moreover, by using a conditional GR null cell line, we demonstrate, for the first time to our knowledge, that one of the possible mechanisms explaining the anti-inflammatory activity of TZDs is their ability to activate GR nuclear translocation. In addition, by using J774 cell line lacking PPAR-gamma, we demonstrate that PPAR-gamma expression could not be essential for TZD-mediated GR nuclear translocation, thus explaining, at least in part, the molecular mechanism underlying their anti-inflammatory activity.
The AHR is an environmental sensor and transcription factor activated by a variety of man-made and natural ligands, which has recently emerged as a critical regulator of homeostasis at barrier organs ...such as the skin. Activation of the AHR pathway downmodulates skin inflammatory responses in animal models and psoriasis clinical samples. In this study, we identify CYP1A1 enzymatic activity as a critical regulator of beneficial AHR signaling in the context of skin inflammation. Mice constitutively expressing Cyp1a1 displayed increased CYP1A1 enzymatic activity in the skin, which resulted in exacerbated immune cell activation and skin pathology, mirroring that observed in Ahr-deficient mice. Inhibition of CYP1A1 enzymatic activity ameliorated the skin immunopathology by restoring beneficial AHR signaling. Importantly, patients with psoriasis displayed reduced activation of the AHR pathway and increased CYP1A1 enzymatic activity compared with healthy donors, suggesting that dysregulation of the AHR/CYP1A1 axis may play a role in inflammatory skin disease. Thus, modulation of CYP1A1 activity may represent a promising alternative strategy to harness the anti-inflammatory effect exerted by activation of the AHR pathway in the skin.
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Epigenetic modifications such as DNA methylation play a key role in gene regulation and disease susceptibility. However, little is known about the genome-wide frequency, localization, and function of ...methylation variation and how it is regulated by genetic and environmental factors. We utilized the Multiple Tissue Human Expression Resource (MuTHER) and generated Illumina 450K adipose methylome data from 648 twins. We found that individual CpGs had low variance and that variability was suppressed in promoters. We noted that DNA methylation variation was highly heritable (h2median = 0.34) and that shared environmental effects correlated with metabolic phenotype-associated CpGs. Analysis of methylation quantitative-trait loci (metQTL) revealed that 28% of CpGs were associated with nearby SNPs, and when overlapping them with adipose expression quantitative-trait loci (eQTL) from the same individuals, we found that 6% of the loci played a role in regulating both gene expression and DNA methylation. These associations were bidirectional, but there were pronounced negative associations for promoter CpGs. Integration of metQTL with adipose reference epigenomes and disease associations revealed significant enrichment of metQTL overlapping metabolic-trait or disease loci in enhancers (the strongest effects were for high-density lipoprotein cholesterol and body mass index BMI). We followed up with the BMI SNP rs713586, a cg01884057 metQTL that overlaps an enhancer upstream of ADCY3, and used bisulphite sequencing to refine this region. Our results showed widespread population invariability yet sequence dependence on adipose DNA methylation but that incorporating maps of regulatory elements aid in linking CpG variation to gene regulation and disease risk in a tissue-dependent manner.
Human translational research is hampered by limited amounts of samples, intrinsic human variability and practical issues involving multi-centre sample collection and analysis. ... human ...immuno-monitoring studies need to be standardized.
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