Kristi Abram

Mechanisms of IFN-γ–induced apoptosis of human skin keratinocytes in patients with atopic dermatitis

BACKGROUND Enhanced apoptosis of keratinocytes is the main cause of eczema and spongiosis in patients with the common inflammatory skin disease atopic dermatitis (AD). OBJECTIVE The aim of the study was to investigate molecular mechanisms of AD-related apoptosis of keratinocytes. METHODS Primary keratinocytes isolated from patients with AD and healthy donors were used to study apoptosis by using annexin V/7-aminoactinomycin D staining. Illumina mRNA Expression BeadChips, quantitative RT-PCR, and immunofluorescence were used to study gene expression. In silico analysis of candidate genes was performed on genome-wide single nucleotide polymorphism data. RESULTS We demonstrate that keratinocytes of patients with AD exhibit increased IFN-γ-induced apoptosis compared with keratinocytes from healthy subjects. Further mRNA expression analyses revealed differential expression of apoptosis-related genes in AD keratinocytes and skin and the upregulation of immune system-related genes in skin biopsy specimens of chronic AD lesions. Three apoptosis-related genes (NOD2, DUSP1, and ADM) and 8 genes overexpressed in AD skin lesions (CCDC109B, CCL5, CCL8, IFI35, LYN, RAB31, IFITM1, and IFITM2) were induced by IFN-γ in primary keratinocytes. The protein expression of IFITM1, CCL5, and CCL8 was verified in AD skin. In line with the functional studies and AD-related mRNA expression changes, in silico analysis of genome-wide single nucleotide polymorphism data revealed evidence of an association between AD and genetic markers close to or within the IFITM cluster or RAB31, DUSP1, and ADM genes. CONCLUSION Our results demonstrate increased IFN-γ responses in skin of patients with AD and suggest involvement of multiple new apoptosis- and inflammation-related factors in the development of AD.

Prevalence of Rosacea in an Estonian Working Population Using a Standard Classification

Data about the prevalence of rosacea are scarce. The aim of this study was to evaluate the prevalence rate of rosacea according to the American National Rosacea Society Expert Committee (NRSEC) classification. A cross-sectional study of 348 subjects randomly selected from a working population >or= 30 years of age was performed. All subjects completed a questionnaire. Skin status was examined according to NRSEC criteria. Of the 348 subjects, 78 (22%) had one or more primary features of rosacea. The most common features were erythema (21%) and telangiectasia (18%). Of the subjects with rosacea, 78% had erythematotelangiectatic rosacea and 22% had papulopustular rosacea. Fifteen percent of the study subjects had experienced frequent episodes of flushing without permanent features of rosacea. No significant gender-related differences were found between study groups. In conclusion, according to the NRSEC, rosacea is a more common skin condition over the age of 30 years than previously thought.

Transcriptional landscape of psoriasis identifies the involvement of IL36 and IL36RN.

BackgroundIn present study we performed whole transcriptome analysis in plaque psoriasis patients and compared lesional skin with non-lesional skin and with the skin from healthy controls. We sequenced total RNA from 12 lesional (LP), 12 non-lesional (NLP) and from 12 normal (C) skin biopsies.ResultsCompared with previous gene expression profiling studies we had three groups under analysis - LP, NLP and C. Using NLP samples allows to see the transcriptome of visually normal skin from psoriasis patient. In LP skin S100A12, S100A7A, LCE3E, DEFB4A, IL19 were found up regulated. In addition to already these well-described genes, we also found several other genes related to psoriasis. Namely, KLK9, OAS2, OAS3, PLA2G, IL36G, IL36RN were found to be significantly and consistently related to the psoriatic lesions and this finding is supported also by previous studies. The genes up-regulated in the LP samples were related to the innate immunity, IL17 and IL10 networks. In NLP samples innate immunity and IL17 network were activated, but activation of IL10 network was not evident. The transcriptional changes characteristic in the NLP samples can be considered as a molecular signature of “dormant psoriasis”.ConclusionsTaken together, our study described the transcriptome profile characteristic for LP and NLP psoriatic skin. RNA profile of the NLP skin is in between the lesional and healthy skin, with its own specific pattern. We found that both LP and NLP have up-regulated IL17 network, whereas LP skin has up regulated IL10 related cytokines (IL19, IL20, IL24). Moreover, IL36G and IL36RN were identified as strong regulators of skin pathology in both LP and NLP skin samples, with stronger influence in LP samples.

Subjective disease perception and symptoms of depression in relation to healthcare-seeking behaviour in patients with rosacea.

Many patients with rosacea do not seek medical care. The aim of this study was to find predictors for healthcare-seeking behaviour among patients with rosacea. The study subjects were 70 consecutive patients attending a dermatologist (seekers) and 56 subjects with rosacea symptoms selected randomly from among the working population (non-seekers). All subjects completed an Estonian Mood Scale questionnaire, a screening instrument for depressive symptoms, and evaluated their subjective disease perception on a visual analogue scale (VAS). Multivariate analysis showed that the independent predictors for healthcare-seeking behaviour were VAS scores >5 and the presence of advanced forms of rosacea. Higher mean VAS scores were not related to severity of rosacea, but were associated with the presence of depressive symptoms among seekers. In conclusion, healthcare-seeking behaviour is associated with higher subjective disease perception. The presence of depressive symptoms is not related to severity of the disease, but to the subjective disease perception of rosacea patients.

MicroRNA-155 is Dysregulated in the Skin of Patients with Vitiligo and Inhibits Melanogenesis-associated Genes in Melanocytes and Keratinocytes.

Little is known about the functions of microRNAs (miRNAs) in skin pigmentation disorders. The aim of this study was to investigate the expression and potential role of miRNAs in vitiligo. Of 12 studied miRNAs with proven functions in cell proliferation, differentiation, immune responses and melanogenesis, miR-99b, miR-125b, miR-155 and miR-199a-3p were found to be increased and miR-145 was found to be decreased in the skin of patients with vitiligo. Combined pathway and target analysis revealed melanogenesis-associated targets for miR-99b, miR-125b, miR-155 and miR-199a-3p. In situ hybridization analysis demonstrated increased expression of miR-155 in the epidermis of patients with vitiligo. Correspondingly, miR-155 was induced by vitiligo-associated cytokines in human primary melanocytes and keratinocytes. When overexpressed, miR-155 inhibited the expression of melanogenesis-associated genes and altered interferon-regulated genes in melanocytes and keratinocytes. In conclusion, this study demonstrates that the expression of miRNAs is dysregulated in the skin of patients with vitiligo and suggests that miR-155 contributes to the pathogenesis of vitiligo.

Expression Profile of Genes Associated with the Dopamine Pathway in Vitiligo Skin Biopsies and Blood Sera

Background: Dopamine has been proven to be toxic for melanocytes. In vitiligo patients the level of dopamine is increased and the functioning of several enzymes participating in the dopamine pathway is changed. Methods: With the use of quantitative real-time polymerase chain reaction and ELISA the expression of genes connected to the dopamine pathway (PAH, PCD, TH, DDC, DBH, PNMT, GPX1, MAOA, MAOB, COMT, DRD1–DRD5, VMAT1 and VMAT2) was observed in vitiligo patients’ and control subjects’ skin and blood. Results: The mRNA expression of GPX1, DDC, MAOA, DRD1 and DRD5 differs in vitiligo skin and the protein level of DDC, MAOA, MAOB, DRD1 and DRD5 is changed in vitiligo patients’ skin and/or blood sera. Conclusions: The dopamine pathway probably influences melanogenesis directly or through the melanocortin pathway. We provide new data about changes of expression profile of the dopamine-synthesizing enzyme DDC, the dopamine-degrading enzymes MAOA and MAOB and the D1-like family dopamine receptors in vitiligo skin and blood sera.

Psoriasis-Specific RNA Isoforms Identified by RNA-Seq Analysis of 173,446 Transcripts

Background Several studies have been published that investigated potential links between transcriptome changes and psoriasis using microarrays and RNA-seq technologies, but no previous study has analyzed expression profile of alternatively spliced transcripts in psoriasis. Objectives Identification of potential alternatively spliced RNA isoforms with disease-specific expression profile. Methods Using our published RNA sequencing data from lesional psoriatic (LP), non-lesional psoriatic (NLP), and normal control skin (C), we analyzed the differential expression of RNA splicing variants. LP sample was compared with NLP, as was LP with C and NLP with C. Results Transcript-based annotation analyzed 173,446 transcripts (RNA isoforms), and around 9,000 transcripts were identified as differentially expressed between study groups. Several previously undescribed RNA variants were found. For instance, transcript ETV3_3 (ENST00000326786) was significantly downregulated in LP and NLP skin. ETV3 is a transcriptional repressor that contributes to the downstream anti-inflammatory effects of IL-10. We also identified diseases-specific transcripts (S100A7A, IL36RN_4, and IL36G_3) of genes already recognized to be involved in inflammation and immune response. Conclusion Psoriasis is characterized by significant differences in the expression of RNA alternative isoforms. Description of these new isoforms improves our knowledge about this complex disease.

miR-146b Probably Assists miRNA-146a in the Suppression of Keratinocyte Proliferation and Inflammatory Responses in Psoriasis

miR-146a inhibits inflammatory responses in human keratinocytes and in different mouse models of skin inflammation. Little is known about the role of miR-146b in the skin. In this study, we confirmed the increased expression of miR-146a and miR-146b (miR-146a/b) in the lesional skin of patients with psoriasis. The expression of miR-146a was approximately twofold higher than that of miR-146b in healthy human skin, and it was more strongly induced by stimulation of proinflammatory cytokines in keratinocytes and fibroblasts. miR-146a/b target genes regulating inflammatory responses or proliferation were altered in the skin of patients with psoriasis, among which FERMT1 was verified as a direct target of miR-146a. In silico analysis of genome-wide data from >4,000 psoriasis cases and >8,000 controls confirmed a moderate association between psoriasis and genetic variants in the miR-146a encoding gene. Transfection of miR-146a/b suppressed and inhibition enhanced keratinocyte proliferation and the expression of psoriasis-related target genes. Enhanced expression of miR-146a/b-influenced genes was detected in cultured keratinocytes from miR-146a-/- and skin fibroblasts from miR-146a-/- and miR-146b-/- mice stimulated with psoriasis-associated cytokines as compared with wild-type mice. Our results indicate that besides miR-146a, miR-146b is expressed and might be capable of modulation of inflammatory responses and keratinocyte proliferation in psoriatic skin.

Signs of innate immune activation and premature immunosenescence in psoriasis patients

Psoriasis is a chronic inflammatory disease that affects skin and is associated with systemic inflammation and many serious comorbidities ranging from metabolic syndrome to cancer. Important discoveries about psoriasis pathogenesis have enabled the development of effective biological treatments blocking the T helper 17 pathway. However, it has not been settled whether psoriasis is a T cell-mediated autoimmune disease or an autoinflammatory disorder that is driven by exaggerated innate immune signalling. Our comparative gene expression and hierarchical cluster analysis reveal important gene circuits involving innate receptors. Innate immune activation is indicated by increased absent in melanoma 2 (AIM2) inflammasome gene expression and active caspase 1 staining in psoriatic lesional skin. Increased eomesodermin (EOMES) expression in lesional and non-lesional skin is suggestive of innate-like virtual memory CD8+ T cell infiltration. We found that signs of systemic inflammation were present in most of the patients, correlated with the severity of the disease, and pointed to IL-6 involvement in the pathogenesis of psoriatic arthritis. Among the circulating T cell subpopulations, we identified a higher proportion of terminally differentiated or senescent CD8+ T cells, especially in patients with long disease duration, suggesting premature immunosenescence and its possible implications for psoriasis co-morbidities.

Transcriptional landscape of human endogenous retroviruses (HERVs) and other repetitive elements in psoriatic skin

Human endogenous retrovirus (HERV) sequences make up at least 8% of the human genome. Transcripts originating from these loci as well as proteins encoded by them have been detected in various tissues. HERVs are believed to be implicated in autoimmune diseases, however the extent to which, has remained unclear. Differential expression studies have so far been limited to certain HERV subfamilies with conserved sequences. No studies have been published describing the genome-wide expression pattern of HERVs and repetitive elements in the context of psoriasis. In the present study, we analysed total RNA sequencing data from skin samples of 12 psoriasis patients and 12 healthy controls, which enabled us to describe the entire transcriptional landscape of repetitive elements. We report high levels of repetitive element expression in the skin of psoriasis patients as well as healthy controls. The majority of differentially expressed elements were downregulated in lesional and non-lesional skin, suggesting active HERV suppression in the pro-inflammatory environment of psoriatic skin. However, we also report upregulation of a small subset of HERVs previously described in the context of autoimmune diseases, such as members of the HERV-K and W families, with the potential to affect the immunopathogenesis of psoriasis.