Ele Prans

Genome-wide association studies of autoimmune vitiligo identify 23 new risk loci and highlight key pathways and regulatory variants

Vitiligo is an autoimmune disease in which depigmented skin results from the destruction of melanocytes, with epidemiological association with other autoimmune diseases. In previous linkage and genome-wide association studies (GWAS1 and GWAS2), we identified 27 vitiligo susceptibility loci in patients of European ancestry. We carried out a third GWAS (GWAS3) in European-ancestry subjects, with augmented GWAS1 and GWAS2 controls, genome-wide imputation, and meta-analysis of all three GWAS, followed by an independent replication. The combined analyses, with 4,680 cases and 39,586 controls, identified 23 new significantly associated loci and 7 suggestive loci. Most encode immune and apoptotic regulators, with some also associated with other autoimmune diseases, as well as several melanocyte regulators. Bioinformatic analyses indicate a predominance of causal regulatory variation, some of which corresponds to expression quantitative trait loci (eQTLs) at these loci. Together, the identified genes provide a framework for the genetic architecture and pathobiology of vitiligo, highlight relationships with other autoimmune diseases and melanoma, and offer potential targets for treatment.

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.

Copy number variations in IL22 gene are associated with Psoriasis vulgaris.

Psoriasis vulgaris (PsV) is a frequent, chronically relapsing, immune-mediated systemic disease with characteristic skin changes. IL22 is a cytokine of IL10 family, with significant proliferative effect on different cell lines. Copy number variations (CNV) have been discovered to have phenotypic consequences and are associated with various types of diseases. In the work presented here we analyzed the copy number variations in IL22 gene of exon1 and exon5. Our results showed that the IL22 gene exon1 was significantly associated with psoriasis severity (P<0.0001). However, the association between IL22 gene exon5 copy numbers and psoriasis was not detected.

CTLA-4 promoter polymorphisms are associated with latent autoimmune diabetes in adults

The cytotoxic T lymphocyte antigen-4 (CTLA-4) molecule is an important regulator of T-cell activation and a susceptibility candidate for autoimmune diseases. To evaluate the impact of CTLA-4 promoter allelic variants of the CTLA-4 gene in latent autoimmune diabetes in adults (LADA), the MH30 (rs231806), -1147 (rs16840252), and -318 (rs5742909) single nucleotide polymorphisms (SNPs) were studied in a population of Estonian origin, including 61 LADA patients and 230 controls. The MH30 GG genotype (p = 0.0051) and the G allele (p = 0.0023) were significantly associated with LADA. The frequency distribution of alleles and genotypes of rs16840252 and rs5742909 SNPs were not significantly different between the patient and control groups. The frequency of the CTLA-4 GCC (p = 0.000073) haplotype was significantly higher in LADA patients, whereas the frequency of the CTLA-4 CCC (p = 0.0019) was significantly lower in LADA patients in comparison with the control group. The current study confirms the involvement of CTLA-4 gene promoter polymorphisms in the susceptibility of LADA and extends our previous findings of associations with other CTLA-4 polymorphisms.

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.

Transcriptome analysis of osteosarcoma identifies suppression of wnt pathway and up-regulation of adiponectin as potential biomarker

Osteosarcoma (OS) is primary malignant bone tumour with complicated early diagnosis. There are no specific markers currently available for predicting the prognosis and chemosensitivity of OS. In present study we performed transcriptome profiling of single patient tumour tissue with RNA-seq technology. We analysed surgically removed sarcoma sample from single 16 years old male patient. Transciptome analysis was done with RNA-seq technology, bioinformatics with Lifescope and R Bioconductor. Validation experiments were done with quantitative real-time PCR (QRTPCR). After quality and coverage filtering, RNA-seq experiment resulted 29,311,899 mapped reads for sarcoma and 22,099,159 mapped reads for normal bone tissue. 65 genes were differentially expressed with FDR corrected statistical significance below 0.05. Seven genes were down-regulated and 58 genes were up-regulated in sarcoma. The most highly up-regulated gene in sarcoma was adiponectin, ADIPOQ (with adjusted p-value 5.5E-07, log2 fold change was 7.9). Many of the genes we found are related to the adipose tissue metabolism (ADIPOQ, PLIN1, FABP4) and to the Wnt signalling suppression (WIF1, SOST). We also found novel fusion transcript between the genes LMTK2 and ZSWIM5. LMTK2 is lemur tyrosine kinase 2, and it has been shown to be involved in NGF-TrkA signalling. Interestingly, studies support the involvement of LMTK2 in development of prostate cancer. ZSWIM5 is zinc finger SWIM domain protein 5 and its function is not known. Immunohistochemical analysis confirmed positive staining for adiponectin in osteosarcoma. This paper is a good illustration how transcriptome analysis can find new biomarkers and targets for complex diseases.

Cytotoxic T-lymphocyte antigen 4 gene polymorphisms are associated with latent autoimmune diabetes in adults.

BACKGROUND Cytotoxic T lymphocyte antigen-4 (CTLA-4) molecule is an important regulator of T cell activation involved in the down-regulation of immune response. Polymorphisms within the CTLA-4 gene have been suggested to confer susceptibility to autoimmune endocrine disorders. METHODS In order to evaluate the impact of allelic variants of the CTLA-4 gene in latent autoimmune diabetes in adults (LADA), the CT60 A/G SNP and the CTBC217_1 C/T SNP were studied in a population of Estonian origin, including 61 LADA patients and 230 controls. RESULTS It was found that the CT60 GG genotype (p=0.004) and the CTBC217_1 TT genotype (p=0.007) were significant associated with LADA. CONCLUSIONS Our investigation revealed that not only type 1 diabetes but also LADA is associated with CTLA-4 gene polymorphisms. The role of CTLA-4 gene in the pathogenesis of LADA is open and needs further investigations.

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.