R. Dey-Rao

Genome-wide expression analysis suggests unique disease-promoting and disease-preventing signatures in Pemphigus vulgaris.

To evaluate pathogenetic mechanisms underlying disease development and progression in the autoimmune skin disease Pemphigus vulgaris (PV), we examined global peripheral blood gene expression in patients and healthy controls. Our goals were to: (1) assign blood gene expression signatures to patients and controls; (2) identify differentially expressed genes (DEGs) and investigate functional pathways associated with these signatures; and (3) evaluate the distribution of DEGs across the genome to identify transcriptional ‘hot spots’. Unbiased hierarchical clustering clearly separated patients from human leukocyte antigen (HLA)-matched controls (MCRs; ‘disease’ signature), and active from remittent patients (‘activity’ signature). DEGs associated with these signatures are involved in immune response, cytoskeletal reorganization, mitogen-activated protein kinase (MAPK) signaling, oxidation-reduction and apoptosis. We further found that MCRs carrying the PV-associated HLA risk alleles cluster distinctly from unmatched controls (UMCR) revealing an HLA-associated ‘control’ signature. A subset of DEGs within the ‘control’ signature overlap with the ‘disease’ signature, but are inversely regulated in MCR when compared with either PV patients or UMCR, suggesting the existence of a ‘protection’ signature in healthy individuals carrying the PV HLA genetic risk elements. Finally, we identified 19 transcriptional ‘hot spots’ across the signatures, which may guide future studies aimed at pinpointing disease risk genes.

Increased oxidative stress in pemphigus vulgaris is related to disease activity and HLA-association

Abstract Pemphigus vulgaris (PV) is a rare blistering skin disorder characterized by the disadhesion of keratinocytes due to autoantibody attack against epidermal targets including desmoglein (Dsg) 3, Dsg 1 and possibly other adhesion and non-adhesion molecules. The mechanisms leading to immune-mediated pathology in PV are multifactorial and not fully understood. Recently, oxidative stress (antioxidant/oxidant disequilibrium) has been proposed as a contributory mechanism of autoimmune skin diseases, including PV. In this study, we directly assessed oxidative stress via measurement of total antioxidant capacity (TAC) using ELISA in 47 PV patients, 25 healthy controls and 18 bullous pemphigoid (BP) patients. We also performed microarray gene expression analysis on a separate set of 21 PV patients and 10 healthy controls to evaluate transcriptional dysregulation in oxidative stress-related pathways. Our data indicate that there is a significant reduction in TAC levels in PV patients compared with healthy controls, as well as BP patients. Furthermore, PV patients with active disease have significantly lower TAC levels than PV patients in remission. We also find that HLA allele status has a significant influence on oxidative stress. These findings are corroborated by microarray analysis showing differentially expressed genes involved in oxidative stress between the aforementioned groups. Collectively, our findings provide support for a role of oxidative stress in PV. Whether increased oxidative stress leads to disease manifestation and/or activity, or if disease activity leads to increased oxidative stress remains unknown. Future longitudinal studies may help to further elucidate the relationship between PV and oxidative stress.

Vitiligo blood transcriptomics provides new insights into disease mechanisms and identifies potential novel therapeutic targets

BackgroundSignificant gaps remain regarding the pathomechanisms underlying the autoimmune response in vitiligo (VL), where the loss of self-tolerance leads to the targeted killing of melanocytes. Specifically, there is incomplete information regarding alterations in the systemic environment that are relevant to the disease state.MethodsWe undertook a genome-wide profiling approach to examine gene expression in the peripheral blood of VL patients and healthy controls in the context of our previously published VL-skin gene expression profile. We used several in silico bioinformatics-based analyses to provide new insights into disease mechanisms and suggest novel targets for future therapy.ResultsUnsupervised clustering methods of the VL-blood dataset demonstrate a “disease-state”-specific set of co-expressed genes. Ontology enrichment analysis of 99 differentially expressed genes (DEGs) uncovers a down-regulated immune/inflammatory response, B-Cell antigen receptor (BCR) pathways, apoptosis and catabolic processes in VL-blood. There is evidence for both type I and II interferon (IFN) playing a role in VL pathogenesis. We used interactome analysis to identify several key blood associated transcriptional factors (TFs) from within (STAT1, STAT6 and NF-kB), as well as “hidden” (CREB1, MYC, IRF4, IRF1, and TP53) from the dataset that potentially affect disease pathogenesis. The TFs overlap with our reported lesional-skin transcriptional circuitry, underscoring their potential importance to the disease. We also identify a shared VL-blood and -skin transcriptional “hot spot” that maps to chromosome 6, and includes three VL-blood dysregulated genes (PSMB8, PSMB9 and TAP1) described as potential VL-associated genetic susceptibility loci. Finally, we provide bioinformatics-based support for prioritizing dysregulated genes in VL-blood or skin as potential therapeutic targets.ConclusionsWe examined the VL-blood transcriptome in context with our (previously published) VL-skin transcriptional profile to address a major gap in knowledge regarding the systemic changes underlying skin-specific manifestation of vitiligo. Several transcriptional “hot spots” observed in both environments offer prioritized targets for identifying disease risk genes. Finally, within the transcriptional framework of VL, we identify five novel molecules (STAT1, PRKCD, PTPN6, MYC and FGFR2) that lend themselves to being targeted by drugs for future potential VL-therapy.

Front-end genomics: using an alternative approach for the recovery of high-quality DNA from core needle biopsies

Aims Determine whether a simple prewash step will provide adequate amounts of high-quality DNA from core needle biopsies for molecular sequencing studies. Methods The quantitative and qualitative metrics of DNA recovered from core needle biopsies processed either by 1) formalin fixation and paraffin embedding (FFPE), 2) cells recovered after the core needle biopsy was washed, and 3) frozen sections of the core needle biopsy tissue were evaluated and compared to one another. Results Fairly equivalent amounts of DNA can be obtained from cells recovered from a prewash step relative to the FFPE and frozen section samples. The number of amplifiable DNA in the wash sample was greater than that from the FFPE samples. The average molecular size of DNA in the wash sample was greater than that of both the FFPE and frozen samples. Conclusions Although more starting material in terms of the number of cells was present in both the FFPE and frozen section samples than the wash samples, equivalent to better results were obtained from the latter with regard to quality. This approach may be a means to better aliquot the diminutive amounts of tissue associated with core needle biopsies, allowing dissociated cells to be dedicated for molecular studies while keeping the tissue intact for morphological studies.

Genome-wide transcriptional profiling data from chronic cutaneous lupus erythematosus (CCLE) peripheral blood.

The disease Lupus erythematosus (LE), exhibits a variety of clinical manifestations with potentially wide-ranging multi-organ damage to joints, tendons, kidney, lung, heart, blood vessels, central nervous system and skin [1,2] Systemic changes are likely to trigger organ specific manifestation of the disease. Here, we provide the data examined to address the gap in knowledge regarding causes and mechanisms contributing to the autoimmune attack on skin in chronic cutaneous lupus erythematosus (CCLE). The raw gene expression data files (CEL files) are provided with this article [3].

Genome-wide transcriptional profiling data from skin of chronic cutaneous lupus erythematosus (CCLE) patients.

Cutaneous features manifest as a wide range of clinically significant, and in many cases disfiguring and debilitating components of lupus erythematosus (LE). While the definitive etiology is in question, multifactorial and polygenic causes are likely to be involved in the production of the characteristic anti-nuclear autoantibody titers and immune cell infiltrates observed in chronic cutaneous LE (CCLE) [1–3]. There is significant overlap of patients with systemic and cutaneous manifestations of LE, which suggests shared pathways and genetic background between the two. We have employed genome-wide microarray technology along with pathway-based analyses to investigate transcriptional differences between lesional and non-lesional skin from CCLE patients to address existing gaps in knowledge regarding disease mechanisms in lupus [4].

Genome-wide gene expression dataset used to identify potential therapeutic targets in androgenetic alopecia

The microarray dataset attached to this report is related to the research article with the title: “A genomic approach to susceptibility and pathogenesis leads to identifying potential novel therapeutic targets in androgenetic alopecia” (Dey-Rao and Sinha, 2017) [1]. Male-pattern hair loss that is induced by androgens (testosterone) in genetically predisposed individuals is known as androgenetic alopecia (AGA). The raw dataset is being made publicly available to enable critical and/or extended analyses. Our related research paper utilizes the attached raw dataset, for genome-wide gene-expression associated investigations. Combined with several in silico bioinformatics-based analyses we were able to delineate five strategic molecular elements as potential novel targets towards future AGA-therapy.