ABSTRACTPediatric steroid-sensitive nephrotic syndrome (pSSNS) is the most common childhood glome... more ABSTRACTPediatric steroid-sensitive nephrotic syndrome (pSSNS) is the most common childhood glomerular disease. Previous genome-wide association studies (GWAS) identified a risk locus in the HLA Class II region and three additional signals. But the genetic architecture of pSSNS, and its genetically driven pathobiology, is largely unknown. We conducted a multi-population GWAS meta-analysis in 38,463 participants (2,440 cases) and population specific GWAS, discovering twelve significant associations (eight novel). Fine-mapping implicated specific amino acid haplotypes in HLA-DQA1 and HLA-DQB1 driving the HLA Class II risk signal. Non-HLA loci colocalized with eQTLs of monocytes and numerous T-cell subsets in independent datasets. Colocalization with kidney eQTLs was lacking, but overlap with kidney cell open chromatin suggests an uncharacterized disease mechanism in kidney cells. A polygenic risk score (PRS) associated with earlier disease onset in two independent cohorts. Altogether,...
Flowchart describing the workflow of analysis. The flowchart provides an overview of the analysis... more Flowchart describing the workflow of analysis. The flowchart provides an overview of the analysis performed for understanding regulation of miRNAs and their contribution to the disease phenotype. Figure S2 Interaction network accessed via IPA software for epistasis of miR-501. The graph depicts the interacting genes identified from epistasis scan of miRNA miR-501 in chromosome 1 and chromosome 2. The graph shows all known gene interactions between the two loci where genes colored in yellow are from locus on chromosome 2 and green are from locus on chromosome 1. The red line shows the possible pathway for the regulation of miR-501. Figure S3 qRT-PCR validation of miR-223 expression in EBA and normal murine skin. (ZIP 637 kb)
Manhattan plot of Neisseria_OTU1320 QTL mapping. Significant thresholds (p ≤ 0.05) are shown in a... more Manhattan plot of Neisseria_OTU1320 QTL mapping. Significant thresholds (p ≤ 0.05) are shown in a continuous line; suggestive thresholds (p ≤ 0.10) are shown in a discontinuous line. Chr: chromosome. (TIF 219 kb)
Apolipoprotein E ( APOE ) genotype regulates body weight and fatty acid utilization-Studies in ge... more Apolipoprotein E ( APOE ) genotype regulates body weight and fatty acid utilization-Studies in gene-targeted replacement mice
The G protein-coupled receptor 15 (GPR15) regulates homing of different T-cell populations into t... more The G protein-coupled receptor 15 (GPR15) regulates homing of different T-cell populations into the gut, thus, preserving tissue homeostasis. Its potential role in the preservation of homeostasis on other body interfaces, including the skin, is less well understood. We addressed the impact of GPR15 on cutaneous T-cell populations and the skin microbiome under steady-state conditions. Genetic deficiency in GPR15 substantially altered the composition of skin-resident T-cell populations. Precisely, dendritic epidermal T cells were almost absent in the epidermis of Gpr15-/mice. The niche of dendritic epidermal T cells in the epidermis was, instead, populated by αβ TCR + T cells. These changes were associated with shifts in the skin microbiota in Gpr15-/mice. Collectively, our results uncover a role of GPR15 in the regulation of the cutaneous immune system and, thus, highlight the receptor as important general regulator of tissue homeostasis of exterior body interfaces. Keywords: dendritic epidermal T cells (DETCs) r G protein-coupled receptor 15 (GPR15) r g/d T cells r skin microbiome r tissue homeostasis Additional supporting information may be found online in the Supporting Information section at the end of the article.
Autoimmune pancreatitis (AIP) is a rare but clinically relevant form of chronic pancreatitis (CP)... more Autoimmune pancreatitis (AIP) is a rare but clinically relevant form of chronic pancreatitis (CP). AIP patients may present with obstructive jaundice due to the formation of inflammatory pseudotumours, a finding that may evoke the differential diagnosis of pancreatic cancer and result in surgical treatment. Correct diagnosis is of particular importance since AIP patients, in contrast to patients with other forms of CP, usually respond well to steroid treatment. 1,2 The current concept of AIP pathophysiology differentiates two subtypes of AIP, AIP type 1 and type 2, which are distinguished by their pathogenesis: AIP type 1 is the pancreatic manifestation of IgG4-related disease and is characterized by dense infiltrates of IgG4-positive plasma cells. In AIP type 2,
Phenotypic variation of quantitative traits is orchestrated by a complex interplay between the en... more Phenotypic variation of quantitative traits is orchestrated by a complex interplay between the environment (e.g. diet) and genetics. However, the impact of gene-environment interactions on phenotypic traits mostly remains elusive. To address this, we feed 1154 mice of an autoimmunity-prone intercross line (AIL) three different diets. We find that diet substantially contributes to the variability of complex traits and unmasks additional genetic susceptibility quantitative trait loci (QTL). By performing whole-genome sequencing of the AIL founder strains, we resolve these QTLs to few or single candidate genes. To address whether diet can also modulate genetic predisposition towards a given trait, we set NZM2410/J mice on similar dietary regimens as AIL mice. Our data suggest that diet modifies genetic susceptibility to lupus and shifts intestinal bacterial and fungal community composition, which precedes clinical disease manifestation. Collectively, our study underlines the importance...
Because of the morbidity and limited therapeutic options of autoimmune diseases, there is a high,... more Because of the morbidity and limited therapeutic options of autoimmune diseases, there is a high, and thus far, unmet medical need for development of novel treatments. Pemphigoid diseases, such as epidermolysis bullosa acquisita (EBA), are prototypical autoimmune diseases that are caused by autoantibodies targeting structural proteins of the skin, leading to inflammation, mediated by myeloid cells. To identify novel treatment targets, we performed cutaneous genome-wide mRNA expression profiling in 190 outbred mice after EBA induction. Comparison of genome-wide mRNA expression profiles in diseased and healthy mice, and construction of a co-expression network identified(spleen tyrosine kinase, SYK) as a major hub gene. Aligned, pharmacological SYK inhibition protected mice from experimental EBA. Using lineage-specific SYK-deficient mice, we identified SYK expression on myeloid cells to be required to induce EBA. Within the predicted co-expression network, interactions ofwith several p...
Pemphigus and systemic lupus erythematosus (SLE) are severe potentially life-threatening autoimmu... more Pemphigus and systemic lupus erythematosus (SLE) are severe potentially life-threatening autoimmune diseases. They are classified as B-cell-mediated autoimmune diseases, both depending on autoreactive CD4T lymphocytes to modulate the autoimmune B-cell response. Despite the reported association of pemphigus and SLE, the molecular mechanisms underlying their comorbidity remain unknown. Weighted gene co-expression network analysis (WGCNA) of publicly available microarray datasets of CD4T cells was performed, to identify shared gene expression signatures and putative overlapping biological molecular mechanisms between pemphigus and SLE. Using WGCNA, we identified 3,280 genes co-expressed genes and 14 co-expressed gene clusters, from which one was significantly upregulated for both diseases. The pathways associated with this module include type-1 interferon gamma and defense response to viruses. Network-based meta-analysis identified RSAD2 to be the most highly ranked hub gene. By associ...
Polycystic liver diseases (PCLDs) are autosomal dominant disorders. To date, 3 genes are known to... more Polycystic liver diseases (PCLDs) are autosomal dominant disorders. To date, 3 genes are known to be associated with the disease, SEC63 and PRKCSH and LRP5. Here, we report that mice deficient in the mitochondrial uncoupling protein 2 gene (Ucp2-/-) spontaneously developed PCLDs when they were over 12months old. Macroscopical observation, blood chemistry as well as histopathological analysis demonstrated the PCLDs found in Ucp2-/- mice were very similar to the findings in human PCLDs. This is the first report describing the gene encoding mitochondrial protein is causative for PCLDs. UCP2 may be a biomarker of the PCLDs in humans.
SummaryMutations in mitochondrial DNA (mtDNA) lead to heteroplasmy, i.e. the intracellular coexis... more SummaryMutations in mitochondrial DNA (mtDNA) lead to heteroplasmy, i.e. the intracellular coexistence of wild-type and mutant mtDNA strands, which impact a wide spectrum of diseases but also physiological processes, including endurance exercise performance in athletes. However, the phenotypic consequences of limited levels of naturally-arising heteroplasmy have not been experimentally studied to date. We hence generated a conplastic mouse strain carrying the mitochondrial genome of a AKR/J mouse strain (B6-mtAKR) together with a C57BL/6J nuclear genomic background, leading to >20% heteroplasmy in the origin of light-strand DNA replication (OriL). These conplastic mice demonstrate a shorter lifespan as well as dysregulation of multiple metabolic pathways, culminating in impaired glucose metabolism, compared to wild-type C57BL/6J mice carrying lower levels of heteroplasmy. Our results indicate that physiologically relevant differences in mtDNA heteroplasmy levels at a single, func...
Recruitment of neutrophils and eosinophils into the skin is a hallmark of pemphigoid diseases. Th... more Recruitment of neutrophils and eosinophils into the skin is a hallmark of pemphigoid diseases. The molecular cues regulating granulocyte recruitment into the skin and the individual contributions of neutrophils and eosinophils to pemphigoid diseases are, however, poorly understood. The lipid mediator leukotriene B 4 (LTB 4) is a potent granulocyte chemoattractant and is abundant in the skin blister fluid of bullous pemphigoid (BP) patients, but its pathogenic significance is unknown. Using mouse models of BP-like epidermolysis bullosa acquisita and of BP, we show that LTB 4 and its receptor BLT1 act as critical drivers of neutrophil entry into the skin upon antibody deposition at the dermal-epidermal junction. Mice deficient in 5-lipoxygenase, a key enzyme in LTB 4 biosynthesis, or in BLT1 exhibited dramatic resistance to neutrophil recruitment and, consequently, skin inflammation. Accordingly, liquid chromatography-mass spectrometry, used to comprehensively profile lipid mediator generation in the first 48 hours after antibody deposition, showed a pronounced parallel increase in LTB 4 and in neutrophils in the skin. Subsequent mechanistic studies in BP-like epidermolysis bullosa acquisita uncovered that neutrophils are necessary for skin inflammation, whereas eosinophils are dispensable, thus identifying neutrophils as major culprits of blister formation. Our results highlight LTB 4 /BLT1 as absolutely critical drivers of murine pemphigoid disease-like skin inflammation.
MicroRNAs (miRNAs) are small endogenous non-coding RNAs that control genes at post-transcriptiona... more MicroRNAs (miRNAs) are small endogenous non-coding RNAs that control genes at post-transcriptional level. They are essential for development and tissue differentiation, and such altered miRNA expression patterns are linked to the pathogenesis of inflammation and cancer. There is evidence that miRNA expression is genetically controlled similar to the transcription of protein-coding genes and previous studies identified quantitative trait loci (QTL) for miRNA expression in the liver. So far, little attention has been paid to miRNA expression in the skin. Moreover, epistatic control of miRNA expression remains unknown. In this study, we characterize genetic regulation of cutaneous miRNA and their correlation with skin inflammation using a previously established murine autoimmune-prone advanced intercross line. We identified in silico 42 eQTL controlling the expression of 38 cutaneous miRNAs and furthermore found two chromosomal hot-spots on chromosomes 2 and 8 that control the expressi...
Journal of immunology (Baltimore, Md. : 1950), Jan 22, 2015
Animal models have enhanced our understanding of the pathogenesis of autoimmune diseases. For the... more Animal models have enhanced our understanding of the pathogenesis of autoimmune diseases. For these models, genetically identical, inbred mice have commonly been used. Different inbred mouse strains, however, show a high variability in disease manifestation. Identifying the factors that influence this disease variability could provide unrecognized insights into pathogenesis. We established a novel Ab transfer-induced model of epidermolysis bullosa acquisita (EBA), an autoimmune disease characterized by (muco)-cutaneous blistering caused by anti-type VII collagen (COL7) autoantibodies. Blistering after anti-COL7 IgG (directed against the von Willebrand factor A-like domain 2) transfer showed clear variability among inbred mouse strains, that is, severe cutaneous blistering and inflammation in C57BL/6J and absence of skin lesions in MRL/MpJ mice. The transfer of anti-COL7 IgG into irradiated, EBA-resistant MRL/MpJ mice, rescued by transplantation with bone marrow from EBA-susceptible ...
ABSTRACTPediatric steroid-sensitive nephrotic syndrome (pSSNS) is the most common childhood glome... more ABSTRACTPediatric steroid-sensitive nephrotic syndrome (pSSNS) is the most common childhood glomerular disease. Previous genome-wide association studies (GWAS) identified a risk locus in the HLA Class II region and three additional signals. But the genetic architecture of pSSNS, and its genetically driven pathobiology, is largely unknown. We conducted a multi-population GWAS meta-analysis in 38,463 participants (2,440 cases) and population specific GWAS, discovering twelve significant associations (eight novel). Fine-mapping implicated specific amino acid haplotypes in HLA-DQA1 and HLA-DQB1 driving the HLA Class II risk signal. Non-HLA loci colocalized with eQTLs of monocytes and numerous T-cell subsets in independent datasets. Colocalization with kidney eQTLs was lacking, but overlap with kidney cell open chromatin suggests an uncharacterized disease mechanism in kidney cells. A polygenic risk score (PRS) associated with earlier disease onset in two independent cohorts. Altogether,...
Flowchart describing the workflow of analysis. The flowchart provides an overview of the analysis... more Flowchart describing the workflow of analysis. The flowchart provides an overview of the analysis performed for understanding regulation of miRNAs and their contribution to the disease phenotype. Figure S2 Interaction network accessed via IPA software for epistasis of miR-501. The graph depicts the interacting genes identified from epistasis scan of miRNA miR-501 in chromosome 1 and chromosome 2. The graph shows all known gene interactions between the two loci where genes colored in yellow are from locus on chromosome 2 and green are from locus on chromosome 1. The red line shows the possible pathway for the regulation of miR-501. Figure S3 qRT-PCR validation of miR-223 expression in EBA and normal murine skin. (ZIP 637 kb)
Manhattan plot of Neisseria_OTU1320 QTL mapping. Significant thresholds (p ≤ 0.05) are shown in a... more Manhattan plot of Neisseria_OTU1320 QTL mapping. Significant thresholds (p ≤ 0.05) are shown in a continuous line; suggestive thresholds (p ≤ 0.10) are shown in a discontinuous line. Chr: chromosome. (TIF 219 kb)
Apolipoprotein E ( APOE ) genotype regulates body weight and fatty acid utilization-Studies in ge... more Apolipoprotein E ( APOE ) genotype regulates body weight and fatty acid utilization-Studies in gene-targeted replacement mice
The G protein-coupled receptor 15 (GPR15) regulates homing of different T-cell populations into t... more The G protein-coupled receptor 15 (GPR15) regulates homing of different T-cell populations into the gut, thus, preserving tissue homeostasis. Its potential role in the preservation of homeostasis on other body interfaces, including the skin, is less well understood. We addressed the impact of GPR15 on cutaneous T-cell populations and the skin microbiome under steady-state conditions. Genetic deficiency in GPR15 substantially altered the composition of skin-resident T-cell populations. Precisely, dendritic epidermal T cells were almost absent in the epidermis of Gpr15-/mice. The niche of dendritic epidermal T cells in the epidermis was, instead, populated by αβ TCR + T cells. These changes were associated with shifts in the skin microbiota in Gpr15-/mice. Collectively, our results uncover a role of GPR15 in the regulation of the cutaneous immune system and, thus, highlight the receptor as important general regulator of tissue homeostasis of exterior body interfaces. Keywords: dendritic epidermal T cells (DETCs) r G protein-coupled receptor 15 (GPR15) r g/d T cells r skin microbiome r tissue homeostasis Additional supporting information may be found online in the Supporting Information section at the end of the article.
Autoimmune pancreatitis (AIP) is a rare but clinically relevant form of chronic pancreatitis (CP)... more Autoimmune pancreatitis (AIP) is a rare but clinically relevant form of chronic pancreatitis (CP). AIP patients may present with obstructive jaundice due to the formation of inflammatory pseudotumours, a finding that may evoke the differential diagnosis of pancreatic cancer and result in surgical treatment. Correct diagnosis is of particular importance since AIP patients, in contrast to patients with other forms of CP, usually respond well to steroid treatment. 1,2 The current concept of AIP pathophysiology differentiates two subtypes of AIP, AIP type 1 and type 2, which are distinguished by their pathogenesis: AIP type 1 is the pancreatic manifestation of IgG4-related disease and is characterized by dense infiltrates of IgG4-positive plasma cells. In AIP type 2,
Phenotypic variation of quantitative traits is orchestrated by a complex interplay between the en... more Phenotypic variation of quantitative traits is orchestrated by a complex interplay between the environment (e.g. diet) and genetics. However, the impact of gene-environment interactions on phenotypic traits mostly remains elusive. To address this, we feed 1154 mice of an autoimmunity-prone intercross line (AIL) three different diets. We find that diet substantially contributes to the variability of complex traits and unmasks additional genetic susceptibility quantitative trait loci (QTL). By performing whole-genome sequencing of the AIL founder strains, we resolve these QTLs to few or single candidate genes. To address whether diet can also modulate genetic predisposition towards a given trait, we set NZM2410/J mice on similar dietary regimens as AIL mice. Our data suggest that diet modifies genetic susceptibility to lupus and shifts intestinal bacterial and fungal community composition, which precedes clinical disease manifestation. Collectively, our study underlines the importance...
Because of the morbidity and limited therapeutic options of autoimmune diseases, there is a high,... more Because of the morbidity and limited therapeutic options of autoimmune diseases, there is a high, and thus far, unmet medical need for development of novel treatments. Pemphigoid diseases, such as epidermolysis bullosa acquisita (EBA), are prototypical autoimmune diseases that are caused by autoantibodies targeting structural proteins of the skin, leading to inflammation, mediated by myeloid cells. To identify novel treatment targets, we performed cutaneous genome-wide mRNA expression profiling in 190 outbred mice after EBA induction. Comparison of genome-wide mRNA expression profiles in diseased and healthy mice, and construction of a co-expression network identified(spleen tyrosine kinase, SYK) as a major hub gene. Aligned, pharmacological SYK inhibition protected mice from experimental EBA. Using lineage-specific SYK-deficient mice, we identified SYK expression on myeloid cells to be required to induce EBA. Within the predicted co-expression network, interactions ofwith several p...
Pemphigus and systemic lupus erythematosus (SLE) are severe potentially life-threatening autoimmu... more Pemphigus and systemic lupus erythematosus (SLE) are severe potentially life-threatening autoimmune diseases. They are classified as B-cell-mediated autoimmune diseases, both depending on autoreactive CD4T lymphocytes to modulate the autoimmune B-cell response. Despite the reported association of pemphigus and SLE, the molecular mechanisms underlying their comorbidity remain unknown. Weighted gene co-expression network analysis (WGCNA) of publicly available microarray datasets of CD4T cells was performed, to identify shared gene expression signatures and putative overlapping biological molecular mechanisms between pemphigus and SLE. Using WGCNA, we identified 3,280 genes co-expressed genes and 14 co-expressed gene clusters, from which one was significantly upregulated for both diseases. The pathways associated with this module include type-1 interferon gamma and defense response to viruses. Network-based meta-analysis identified RSAD2 to be the most highly ranked hub gene. By associ...
Polycystic liver diseases (PCLDs) are autosomal dominant disorders. To date, 3 genes are known to... more Polycystic liver diseases (PCLDs) are autosomal dominant disorders. To date, 3 genes are known to be associated with the disease, SEC63 and PRKCSH and LRP5. Here, we report that mice deficient in the mitochondrial uncoupling protein 2 gene (Ucp2-/-) spontaneously developed PCLDs when they were over 12months old. Macroscopical observation, blood chemistry as well as histopathological analysis demonstrated the PCLDs found in Ucp2-/- mice were very similar to the findings in human PCLDs. This is the first report describing the gene encoding mitochondrial protein is causative for PCLDs. UCP2 may be a biomarker of the PCLDs in humans.
SummaryMutations in mitochondrial DNA (mtDNA) lead to heteroplasmy, i.e. the intracellular coexis... more SummaryMutations in mitochondrial DNA (mtDNA) lead to heteroplasmy, i.e. the intracellular coexistence of wild-type and mutant mtDNA strands, which impact a wide spectrum of diseases but also physiological processes, including endurance exercise performance in athletes. However, the phenotypic consequences of limited levels of naturally-arising heteroplasmy have not been experimentally studied to date. We hence generated a conplastic mouse strain carrying the mitochondrial genome of a AKR/J mouse strain (B6-mtAKR) together with a C57BL/6J nuclear genomic background, leading to >20% heteroplasmy in the origin of light-strand DNA replication (OriL). These conplastic mice demonstrate a shorter lifespan as well as dysregulation of multiple metabolic pathways, culminating in impaired glucose metabolism, compared to wild-type C57BL/6J mice carrying lower levels of heteroplasmy. Our results indicate that physiologically relevant differences in mtDNA heteroplasmy levels at a single, func...
Recruitment of neutrophils and eosinophils into the skin is a hallmark of pemphigoid diseases. Th... more Recruitment of neutrophils and eosinophils into the skin is a hallmark of pemphigoid diseases. The molecular cues regulating granulocyte recruitment into the skin and the individual contributions of neutrophils and eosinophils to pemphigoid diseases are, however, poorly understood. The lipid mediator leukotriene B 4 (LTB 4) is a potent granulocyte chemoattractant and is abundant in the skin blister fluid of bullous pemphigoid (BP) patients, but its pathogenic significance is unknown. Using mouse models of BP-like epidermolysis bullosa acquisita and of BP, we show that LTB 4 and its receptor BLT1 act as critical drivers of neutrophil entry into the skin upon antibody deposition at the dermal-epidermal junction. Mice deficient in 5-lipoxygenase, a key enzyme in LTB 4 biosynthesis, or in BLT1 exhibited dramatic resistance to neutrophil recruitment and, consequently, skin inflammation. Accordingly, liquid chromatography-mass spectrometry, used to comprehensively profile lipid mediator generation in the first 48 hours after antibody deposition, showed a pronounced parallel increase in LTB 4 and in neutrophils in the skin. Subsequent mechanistic studies in BP-like epidermolysis bullosa acquisita uncovered that neutrophils are necessary for skin inflammation, whereas eosinophils are dispensable, thus identifying neutrophils as major culprits of blister formation. Our results highlight LTB 4 /BLT1 as absolutely critical drivers of murine pemphigoid disease-like skin inflammation.
MicroRNAs (miRNAs) are small endogenous non-coding RNAs that control genes at post-transcriptiona... more MicroRNAs (miRNAs) are small endogenous non-coding RNAs that control genes at post-transcriptional level. They are essential for development and tissue differentiation, and such altered miRNA expression patterns are linked to the pathogenesis of inflammation and cancer. There is evidence that miRNA expression is genetically controlled similar to the transcription of protein-coding genes and previous studies identified quantitative trait loci (QTL) for miRNA expression in the liver. So far, little attention has been paid to miRNA expression in the skin. Moreover, epistatic control of miRNA expression remains unknown. In this study, we characterize genetic regulation of cutaneous miRNA and their correlation with skin inflammation using a previously established murine autoimmune-prone advanced intercross line. We identified in silico 42 eQTL controlling the expression of 38 cutaneous miRNAs and furthermore found two chromosomal hot-spots on chromosomes 2 and 8 that control the expressi...
Journal of immunology (Baltimore, Md. : 1950), Jan 22, 2015
Animal models have enhanced our understanding of the pathogenesis of autoimmune diseases. For the... more Animal models have enhanced our understanding of the pathogenesis of autoimmune diseases. For these models, genetically identical, inbred mice have commonly been used. Different inbred mouse strains, however, show a high variability in disease manifestation. Identifying the factors that influence this disease variability could provide unrecognized insights into pathogenesis. We established a novel Ab transfer-induced model of epidermolysis bullosa acquisita (EBA), an autoimmune disease characterized by (muco)-cutaneous blistering caused by anti-type VII collagen (COL7) autoantibodies. Blistering after anti-COL7 IgG (directed against the von Willebrand factor A-like domain 2) transfer showed clear variability among inbred mouse strains, that is, severe cutaneous blistering and inflammation in C57BL/6J and absence of skin lesions in MRL/MpJ mice. The transfer of anti-COL7 IgG into irradiated, EBA-resistant MRL/MpJ mice, rescued by transplantation with bone marrow from EBA-susceptible ...
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