Elevated monocyte-specific type I interferon signalling correlates positively with cardiac healing in myocardial infarct patients but interferon alpha application deteriorates myocardial healing in rats.

Monocytes are involved in adverse left ventricular (LV) remodelling following myocardial infarction (MI). To provide therapeutic opportunities we aimed to identify gene transcripts in monocytes that relate to post-MI healing and evaluated intervention with the observed gene activity in a rat MI model. In 51 MI patients treated by primary percutaneous coronary intervention (PCI), the change in LV end-diastolic volume index (EDVi) from baseline to 4-month follow-up was assessed using cardiovascular magnetic resonance imaging (CMR). Circulating monocytes were collected at day 5 (Arterioscler Thromb Vasc Biol 35:1066-1070, 2015; Cell Stem Cell 16:477-487, 2015; Curr Med Chem 13:1877-1893, 2006) after primary PCI for transcriptome analysis. Transcriptional profiling and pathway analysis revealed that patients with a decreased LV EDVi showed an induction of type I interferon (IFN) signalling (type I IFN pathway: P value < 0.001; false discovery rate < 0.001). We subsequently administered 15,000 Units of IFN-α subcutaneously in a rat MI model for three consecutive days following MI. Cardiac function was measured using echocardiography and infarct size/cardiac inflammation using (immuno)-histochemical analysis. We found that IFN-α application deteriorated ventricular dilatation and increased infarct size at day 28 post-MI. Moreover, IFN-α changed the peripheral monocyte subset distribution towards the pro-inflammatory monocyte subset whereas in the myocardium, the presence of the alternative macrophage subset was increased at day 3 post-MI. Our findings suggest that induction of type I IFN signalling in human monocytes coincides with adverse LV remodelling. In rats, however, IFN-α administration deteriorated post-MI healing. These findings underscore important but also contradictory roles for the type I IFN response during cardiac healing following MI.

Interferon Gamma-Induced Protein (IP-10) as Potential Biomarker for Cancer-Related-Fatigue: Results from a 6-month Randomized Controlled Trial.

We examined if serum concentrations Interferon gamma-induced protein (IP-10) is a potential clinical biomarker for cancer-related-fatigue (CRF). Fatigue scores and IP-10 concentrations were measured from curatively treated fatigued cancer patients randomized to either cognitive behavioral therapy (CBT, n = 26) or waiting-list (WL, n = 13). No correlation was found between baseline IP-10 level and fatigue severity and no significant differences in IP-10 serum levels were observed between fatigued and matched non-fatigued patients (n = 22). Relative changes in IP-10 concentrations from baseline to six-month follow-up were not significantly different between the CBT and WL conditions. In this study, IP-10 showed low potential as clinical CRF biomarker. TRIAL REGISTRATION: This study is registered at ClinicalTrials.gov (NCT01096641).

CCR7-CCL19/CCL21 Axis is Essential for Effective Arteriogenesis in a Murine Model of Hindlimb Ischemia.

BACKGROUND: In order to identify factors that stimulate arteriogenesis after ischemia, we followed gene expression profiles in two extreme models for collateral artery formation over 28 days after hindlimb ischemia, namely "good-responding" C57BL/6 mice and "poor-responding" BALB/c mice. METHODS AND RESULTS: Although BALB/c mice show very poor blood flow recovery after ischemia, most known proarteriogenic genes were upregulated more excessively and for a longer period than in C57BL/6 mice. In clear contrast, chemokine genes Ccl19, Ccl21a, and Ccl21c and the chemokine receptor CCR7 were upregulated in C57BL/6 mice 1 day after hindlimb ischemia, but not in BALB/C mice. CCL19 and CCL21 regulate migration and homing of T lymphocytes via CCR7. When subjecting CCR7-/-/LDLR-/- mice to hindlimb ischemia, we observed a 20% reduction in blood flow recovery compared with that in LDLR-/- mice. Equal numbers of α-smooth muscle actin-positive collateral arteries were found in the adductor muscles of both mouse strains, but collateral diameters were smaller in the CCR7-/-/LDLR-/-. Fluorescence-activated cell sorter analyses showed that numbers of CCR7+ T lymphocytes (both CD4+ and CD8+) were decreased in the spleen and increased in the blood at day 1 after hindlimb ischemia in LDLR-/- mice. At day 1 after hindlimb ischemia, however, numbers of activated CD4+ T lymphocytes were decreased in the draining lymph nodes of LDLR-/- mice compared with CCR7-/-/LDLR-/- mice. CONCLUSIONS: These data show that CCR7-CCL19/CCL21 axis facilitates retention CD4+ T lymphocytes at the site of collateral artery remodeling, which is essential for effective arteriogenesis.

Identification of HIF-2α-regulated genes that play a role in human microvascular endothelial sprouting during prolonged hypoxia in vitro.

During prolonged hypoxic conditions, endothelial cells change their gene expression to adjust to the low oxygen environment. This process is mainly regulated by the hypoxia-inducible factors, HIF-1α and HIF-2α. Although endothelial cells do not form sprouts during prolonged hypoxic culturing, silencing of HIF-2α partially restores sprout formation. The present study identifies novel HIF-2α-target genes that may regulate endothelial sprouting during prolonged hypoxia. The gene expression profile of primary human microvascular endothelial cells (hMVECs) that were cultured at 20 % oxygen was compared to hMVECs that were cultured at 1 % oxygen for 14 days by using genome-wide RNA-sequencing. The differentially regulated genes in hypoxia were compared to the genes that were differentially regulated upon silencing of HIF-2α in hypoxia. Surprisingly, KEGG pathway analysis showed that metabolic pathways were enriched within genes upregulated in response to hypoxia and enriched within genes downregulated upon HIF-2α silencing. Moreover, 51 HIF-2α-regulated genes were screened for their role in endothelial sprouting in hypoxia, of which four genes ARRDC3, MME, PPARG and RALGPS2 directly influenced endothelial sprouting during prolonged hypoxic culturing. The manipulation of specific downstream targets of HIF-2α provides a new, but to be further evaluated, perspective for restoring reduced neovascularization in several pathological conditions, such as diabetic ulcers or other chronic wounds, for improvement of vascularization of implanted tissue-engineered scaffolds.

The emerging role of galectins in cardiovascular disease.

Galectins are an ancient family of ß-galactoside-specific lectins and consist of 15 different types, each with a specific function. They play a role in the immune system, inflammation, wound healing and carcinogenesis. In particular the role of galectin in cancer is widely studied. Lately, the role of galectins in the development of cardiovascular disease has gained attention. Worldwide cardiovascular disease is still the leading cause of death. In ischemic heart disease, atherosclerosis limits adequate blood flow. Angiogenesis and arteriogenesis are highly important mechanisms relieving ischemia by restoring perfusion to the post-stenotic myocardial area. Galectins act ambiguous, both relieving ischemia and accelerating atherosclerosis. Atherosclerosis can ultimately lead to myocardial infarction or ischemic stroke, which are both associated with galectins. There is also a role for galectins in the development of myocarditis by their influence on inflammatory processes. Moreover, galectin acts as a biomarker for the severity of myocardial ischemia and heart failure. This review summarizes the association between galectins and the development of multiple cardiovascular diseases such as myocarditis, ischemic stroke, myocardial infarction, heart failure and atrial fibrillation. Furthermore it focuses on the association between galectin and more general mechanisms such as angiogenesis, arteriogenesis and atherosclerosis.

Physiological evidence for diversification of IFNα- and IFNß-mediated response programs in different autoimmune diseases.

BACKGROUND: Activation of the type I interferon (IFN) response program is described for several autoimmune diseases, including systemic lupus erythematosus (SLE), multiple sclerosis (MS), myositis (IIM) and rheumatoid arthritis (RA). While IFNα contributes to SLE pathology, IFNß therapy is often beneficial in MS, implying different immunoregulatory roles for these IFNs. This study was aimed to investigate potential diversification of IFNα-and IFNß-mediated response programs in autoimmune diseases. METHODS: Peripheral blood gene expression of 23 prototypical type I IFN response genes (IRGs) was determined in 54 healthy controls (HCs), 69 SLE (47 test, 22 validation), 149 IFNß-treated MS (71 test, 78 validation), 160 untreated MS, 78 IIM and 76 RA patients. Patients with a type I IFN signature were selected for analysis. RESULTS: We identified IFNα- and IFNß-specific response programs (GC-A and GC-B, respectively) in SLE and IFNß-treated MS patients. Concordantly, the GC-A/GC-B log-ratio was positive for all SLE patients and negative for virtually all IFNß-treated MS patients, which was confirmed in additional cohorts. Applying this information to other autoimmune diseases, IIM patients displayed positive GC-A/GC-B log-ratios, indicating predominant IFNα activity. The GC-A/GC-B log-ratio in RA was lower and approached zero in part of the patients, implying relative importance of both clusters. Remarkably, GC-A/GC-B log-ratios appeared most heterogeneous in untreated MS; half of the patients displayed GC-A dominance, whereas others showed GC-B dominance or log-ratios near zero. CONCLUSIONS: Our findings show diversification of the type I IFN response in autoimmune diseases, suggesting different pathogenic roles of the type I IFNs.

IFN-ß affects the angiogenic potential of circulating angiogenic cells by activating calpain 1.

Circulating angiogenic cells (CACs) are monocyte-derived cells with endothelial characteristics, which contribute to both angiogenesis and arteriogenesis in a paracrine way. Interferon-ß (IFN-ß) is known to inhibit these divergent processes in animals and patients. We hypothesized that IFN-ß might act by affecting the differentiation and function of CACs. CACs were cultured from peripheral blood mononuclear cells and phenotypically characterized by surface expression of monocytic and endothelial markers. IFN-ß significantly reduced the number of CACs by 18-64%. Apoptosis was not induced by IFN-ß, neither in mononuclear cells during differentiation, nor after maturation to CACs. Rather, IFN-ß impaired adhesion to, and spreading on, fibronectin, which was dependent on α5ß1 (VLA-5)-integrin. IFN-ß affected the function of VLA-5 in mature CACs, leading to rounding and detachment of cells, by induction of calpain 1 activity. Cell rounding and detachment was completely reversed by inhibition of calpain 1 activity in mature CACs. During in vitro capillary formation, CAC addition and calpain 1 inhibition enhanced sprouting of endothelial cells to a comparable extent, but were not sufficient to rescue tube formation in the presence of IFN-ß. We show that the IFN-ß-induced reduction of the numbers of in vitro differentiated CACs is based on activation of calpain 1, resulting in an attenuated adhesion to extracellular matrix proteins via VLA-5. In vivo, this could lead to inhibition of vessel formation due to reduction of the locally recruited CAC numbers and their paracrine angiogenic factors.

Metabolic changes in type 2 diabetes are reflected in peripheral blood cells, revealing aberrant cytotoxicity, a viral signature, and hypoxia inducible factor activity.

BACKGROUND: Metabolic syndrome (MetS) is characterized by central obesity, insulin resistance, dysglycemia, and a pro-atherogenic plasma lipid profile. MetS creates a high risk for development of type 2 diabetes (T2DM) and cardiovascular disease (CVD), presumably by altering inflammatory responses. Presently, it is unknown how the chronic metabolic disturbances in acute hyperglycemia, MetS and T2DM affect the immune activity of peripheral blood cells. METHODS: We performed genome-wide expression analysis of peripheral blood cells obtained from patients with T2DM (n = 6) and age-, sex- , BMI- and blood pressure-matched obese individuals with MetS (n = 4) and lean healthy normoglycemic controls (n = 3), both under fasting conditions and after controlled induction of acute hyperglycemia during a 70 min hyperglycemic clamp. Differential gene expression during fasting conditions was confirmed by real-time PCR, for which we included additional age-, sex-, BMI-, and blood pressure-matched obese individuals with (n = 4) or without (n = 4) MetS. RESULTS: Pathway and Gene ontology analysis applied to baseline expression profiles of peripheral blood cells from MetS and T2DM patients revealed metabolic changes, highly similar to a reoviral infection gene signature in T2DM patients. Transcription factor binding site analysis indicated that increased HIF-1α activity, a transcription factor induced by either hypoxia or oxidative stress, is responsible for this aberrant metabolic profile in peripheral blood cells from T2DM patients. Acute hyperglycemia in healthy controls resulted in reduced expression of cytotoxicity-related genes, representing NK- and CD8(+) cells. In obese controls, MetS and especially T2DM patients, baseline expression of genes involved in cytotoxicity was already low, compared to healthy controls and did not further decrease upon acute hyperglycemia. CONCLUSIONS: The reduced activity of cytotoxic genes in T2DM is explained by chronic hyperglycemia, but its acute effects are restricted to healthy controls. Genome expression of circulating leukocytes from T2DM patients differs from MetS individuals by a specific reovirus signature. Our data thus suggest a role for suppressed anti-viral capacity in the etiology of diabetes.

Galectin-2 induces a proinflammatory, anti-arteriogenic phenotype in monocytes and macrophages.

Galectin-2 is a monocyte-expressed carbohydrate-binding lectin, for which increased expression is genetically determined and associated with decreased collateral arteriogenesis in obstructive coronary artery disease patients. The inhibiting effect of galectin-2 on arteriogenesis was confirmed in vivo, but the mechanism is largely unknown. In this study we aimed to explore the effects of galectin-2 on monocyte/macrophage phenotype in vitro and vivo, and to identify the receptor by which galectin-2 exerts these effects. We now show that the binding of galectin-2 to different circulating human monocyte subsets is dependent on monocyte surface expression levels of CD14. The high affinity binding is blocked by an anti-CD14 antibody but not by carbohydrates, indicating a specific protein-protein interaction. Galectin-2 binding to human monocytes modulated their transcriptome by inducing proinflammatory cytokines and inhibiting pro-arteriogenic factors, while attenuating monocyte migration. Using specific knock-out mice, we show that galectin-2 acts through the CD14/toll-like receptor (TLR)-4 pathway. Furthermore, galectin-2 skews human macrophages to a M1-like proinflammatory phenotype, characterized by a reduced motility and expression of an anti-arteriogenic cytokine/growth factor repertoire. This is accompanied by a switch in surface protein expression to CD40-high and CD206-low (M1). In a murine model we show that galectin-2 administration, known to attenuate arteriogenesis, leads to increased numbers of CD40-positive (M1) and reduced numbers of CD206-positive (M2) macrophages surrounding actively remodeling collateral arteries. In conclusion galectin-2 is the first endogenous CD14/TLR4 ligand that induces a proinflammatory, non-arteriogenic phenotype in monocytes/macrophages. Interference with CD14-Galectin-2 interaction may provide a new intervention strategy to stimulate growth of collateral arteries in genetically compromised cardiovascular patients.

Interferon-Beta, a Decisive Factor in Angiogenesis and Arteriogenesis.

In this review we discuss the current literature on the effects of type I interferons (IFN) and their downstream effectors on vascular growth in experimental models in vitro and in vivo. In addition to its well-documented role in angiogenesis, that is, the growth of new capillaries from existing vessels, we will also describe emerging evidence and mechanisms by which type I IFN may inhibit arteriogenesis, that is, the expansive remodeling of existing collateral arteries. Crucial in both processes is the common role of circulating monocytes, which are known to act as pivotal cellular modulators in revascularization through secreted chemokines, proteases, and growth factors. These secreted molecules, which are all modulated by IFN signaling, act via degradation of the extracellular matrix and by stimulating the proliferation of vascular smooth muscle cells and endothelial cells. Thus, next to the antiviral and immunomodulatory activities of type I IFNs, a potent role of IFN-ß as modulator of revascularization is now emerging and may be considered a potential clinical target for the stimulation of angiogenesis and arteriogenesis in ill-perfused tissues.

Systemic toll-like receptor and interleukin-18 pathway activation in patients with acute ST elevation myocardial infarction.

Acute myocardial infarction (AMI) is accompanied by increased expression of Toll like receptors (TLR)-2 and TLR4 on circulating monocytes. In animal models, blocking TLR2/4 signaling reduces inflammatory cell influx and infarct size. The clinical consequences of TLR activation during AMI in humans are unknown, including its role in long-term cardiac functional outcome Therefore, we analyzed gene expression in whole blood samples from 28 patients with an acute ST elevation myocardial infarction (STEMI), enrolled in the EXenatide trial for AMI patients (EXAMI), both at admission and after 4-month follow-up, by whole genome expression profiling and real-time PCR. Cardiac function was determined by cardiac magnetic resonance (CMR) imaging at baseline and after 4-month follow-up. TLR pathway activation was shown by increased expression of TLR4 and its downstream genes, including IL-18R1, IL-18R2, IL-8, MMP9, HIF1A, and NFKBIA. In contrast, expression of the classical TLR-induced genes, TNF, was reduced. Bioinformatics analysis and in vitro experiments explained this noncanonical TLR response by identification of a pivotal role for HIF-1α. The extent of TLR activation and IL-18R1/2 expression in circulating cells preceded massive troponin-T release and correlated with the CMR-measured ischemic area (R=0.48, p=0.01). In conclusion, we identified a novel HIF-1-dependent noncanonical TLR activation pathway in circulating leukocytes leading to enhanced IL-18R expression which correlated with the magnitude of the ischemic area. This knowledge may contribute to our mechanistic understanding of the involvement of the innate immune system during STEMI and may yield diagnostic and prognostic value for patients with myocardial infarction.

MicroRNAs regulate human brain endothelial cell-barrier function in inflammation: implications for multiple sclerosis.

Blood-brain barrier (BBB) dysfunction is a major hallmark of many neurological diseases, including multiple sclerosis (MS). Using a genomics approach, we defined a microRNA signature that is diminished at the BBB of MS patients. In particular, miR-125a-5p is a key regulator of brain endothelial tightness and immune cell efflux. Our findings suggest that repair of a disturbed BBB through microRNAs may represent a novel avenue for effective treatment of MS.

Pharmacological induction of interferon type I activity following treatment with rituximab determines clinical response in rheumatoid arthritis.

OBJECTIVE: Despite the fact that rituximab depletes B cells in all treated patients with RA, not all patients show a favourable clinical response. The goal of this study was to provide insight into pharmacological changes in peripheral blood that are associated with clinical response to rituximab. METHODS: Gene expression profiling was performed on peripheral blood RNA of 13 patients with RA (test group) using Illumina HumanHT beadchip microarrays. An independent group of nine patients was used for validation using TaqMan quantitative PCR. Clinical responder status was determined after 6 months using change in 28-joint Disease Activity Score (ΔDAS28) and European League Against Rheumatism (EULAR) response criteria. Significance analysis of microarrays and ontology analysis were used for data analysis and interpretation. RESULTS: Pharmacogenomic analyses demonstrated marked interindividual differences in the pharmacological responses at 3 and 6 months after start of treatment with rituximab. Interestingly, only differences in the regulation of type I interferon (IFN)-response genes after 3 months correlated with the ΔDAS28 response. Good responders (DAS>1.2; n=7) exhibited a selective increase in the expression of type I IFN-response genes, whereas this activity was unchanged or hardly changed in non-responders (DAS<1.2; n=6) (p=0.0040 at a cut-off of 1.1-fold induction). Similar results were obtained using EULAR response criteria. These results were validated in an independent cohort of nine patients (five non-responders and four responders, p=0.0317). CONCLUSIONS: A good clinical response to rituximab in RA is associated with a selective drug-induced increase in type I IFN-response activity in patients with RA. This finding may provide insight in the biological mechanism underlying the therapeutic response to rituximab.

Expression of a retinoic acid signature in circulating CD34 cells from coronary artery disease patients.

BACKGROUND: Circulating CD34+ progenitor cells have the potential to differentiate into a variety of cells, including endothelial cells. Knowledge is still scarce about the transcriptional programs used by CD34+ cells from peripheral blood, and how these are affected in coronary artery disease (CAD) patients. RESULTS: We performed a whole genome transcriptome analysis of CD34+ cells, CD4+ T cells, CD14+ monocytes, and macrophages from 12 patients with CAD and 11 matched controls. CD34+ cells, compared to other mononuclear cells from the same individuals, showed high levels of KRAB box transcription factors, known to be involved in gene silencing. This correlated with high expression levels in CD34+ cells for the progenitor markers HOXA5 and HOXA9, which are known to control expression of KRAB factor genes. The comparison of expression profiles of CD34+ cells from CAD patients and controls revealed a less naïve phenotype in patients' CD34+ cells, with increased expression of genes from the Mitogen Activated Kinase network and a lowered expression of a panel of histone genes, reaching levels comparable to that in more differentiated circulating cells. Furthermore, we observed a reduced expression of several genes involved in CXCR4-signaling and migration to SDF1/CXCL12. CONCLUSIONS: The altered gene expression profile of CD34+ cells in CAD patients was related to activation/differentiation by a retinoic acid-induced differentiation program. These results suggest that circulating CD34+ cells in CAD patients are programmed by retinoic acid, leading to a reduced capacity to migrate to ischemic tissues.

Synovial tissue heterogeneity in rheumatoid arthritis in relation to disease activity and biomarkers in peripheral blood.

OBJECTIVE: To investigate the clinical relevance of synovial tissue subtypes in rheumatoid arthritis (RA) and to search for peripheral blood (PB) markers that may serve as biomarkers for tissue subtypes. METHODS: Gene expression analysis using complementary DNA microarrays was applied on paired synovial tissue biopsy and PB samples obtained from 17 RA patients. Molecular tissue subtypes were correlated with histologic parameters (CD3, CD22, CD38, CD68, CD163, tumor necrosis factor alpha, intercellular adhesion molecule 1, vascular cell adhesion molecule, and E-selectin), disease characteristics, and PB markers. PANTHER classification was used for pathway analysis. RESULTS: Genomic subtyping of high- and low-inflammation rheumatoid synovial tissues based on gene expression profiles exactly matched immunohistochemical classification. The patients with the high-inflammation tissue type had higher Disease Activity Scores in 28 joints, higher C-reactive protein levels, higher erythrocyte sedimentation rates, increased numbers of platelets, and shorter disease durations. Comparative analysis of PB gene expression profiles yielded no statistically significant differences between the 2 tissue groups at the single-gene expression level. PANTHER pathway analysis revealed a significant association of increased protein biosynthesis with high-inflammation tissue. CONCLUSION: High-inflammation tissue is associated with more severe disease and shorter disease duration. While pathway-level analysis revealed that coordinate differential expression of genes involved in protein synthesis in PB is associated with high-inflammation tissue types, differential tissue pathology was not reflected in the PB by differential expression of single genes.

Gene expression profiling in autoantibody-positive patients with arthralgia predicts development of arthritis.

OBJECTIVE: To identify molecular features associated with the development of rheumatoid arthritis (RA), to understand the pathophysiology of preclinical development of RA, and to assign predictive biomarkers. METHODS: The study group comprised 109 anti-citrullinated protein antibody (ACPA)- and/or rheumatoid factor-positive patients with arthralgia who did not have arthritis but were at risk of RA, and 25 patients with RA. The gene expression profiles of blood samples obtained from these patients were determined by DNA microarray analysis and quantitative polymerase chain reaction. RESULTS: In 20 of the 109 patients with arthralgia who were at risk of RA, arthritis developed after a median of 7 months. Gene expression profiling of blood cells revealed heterogeneity among the at-risk patients, based on differential expression of immune-related genes. This report is the first to describe gene signatures relevant to the development of arthritis. Signatures significantly associated with arthritis development were involved in interferon (IFN)-mediated immunity, hematopoiesis, and chemokine/cytokine activity. Logistic regression analysis revealed that the odds ratio (OR) for developing arthritis within 12 months was 21.0 (95% confidence interval [95% CI] 2.8-156.1 [P = 0.003]) for the subgroup characterized by increased expression of genes involved in IFN-mediated immunity and/or cytokine/chemokine-activity. Genes involved in B cell immunology were associated with protection against progression to arthritis (OR 0.38, 95% CI 0.21-0.70 [P = 0.002]). These processes were reminiscent of those in patients with RA, implying that the preclinical phase of disease is associated with features of established disease. CONCLUSION: The results of this study indicate that IFN-mediated immunity, hematopoiesis, and cell trafficking specify processes relevant to the progression of arthritis independent of ACPA positivity. These findings strongly suggest that certain gene signatures have value for predicting the progression to arthritis, which will pave the way to preventive medicine.

Type I interferons have no major influence on humoral autoimmunity in rheumatoid arthritis.

OBJECTIVE: Type I IFNs have recently been implicated in autoantibody-mediated diseases such as SLE. As half the RA patients display a type I IFN(high) signature, we investigated in a pilot study if type I IFN determines the autoantibody response in RA. METHODS: Serum and peripheral blood cells were obtained from 52 RA patients, with paired samples before and after infliximab treatment in 21 patients. Additional samples were collected from 8 anti-citrullinated protein antibody (ACPA)-positive individuals without arthritis and from 10 ACPA-negative healthy controls. The type I IFN signature was determined by peripheral blood cell gene expression analysis and quantitative RT-PCR. ACPA IgG and IgM, RF IgM, anti-nucleosome IgM and anti-dsDNA were measured by ELISA. RESULTS: The type I IFN signature was not related to the presence and titers of ACPA and RF during active disease. TNF blockade induced a similar rise of ANAs, and a similar decrease in RF titers in both groups. ACPA IgG and IgM levels appeared to be down-modulated only in the type I IFN(low) group. These changes were independent of the changes in type I IFN response gene activity after TNF blockade. Furthermore, the ACPA response in individuals without arthritis and inflammation was not related to an increase of type I IFN. CONCLUSIONS: In this explorative study, type I IFN signature does not appear to have a major impact on the humoral autoimmune response in RA. Replication of these data remains warranted.

Loss of imprinting of IGF2 characterises high IGF2 mRNA-expressing type of fibroblast-like synoviocytes in rheumatoid arthritis.

OBJECTIVE: Increased expression of insulin-like growth factor 2 (IGF2) by fibroblast-like synoviocytes (FLS) was associated with low inflammatory synovium of patients with rheumatoid arthritis (RA). The aim of this study was to analyse whether the differential expression of IGF2, whose expression is normally restricted to one allele, is due to activation of the normally suppressed allele. METHODS: IGF2 gene expression of RA FLS was quantified by quantitative real-time PCR. FLS heterozygous for a 3'-untranslated region IGF2 polymorphism were selected to measure the relative contribution of the allelic transcripts by allele-specific transcript quantification assay. Proliferation was determined by [(3)H]thymidine incorporation. RESULTS: IGF2 was shown to contribute to RA FLS proliferation. FLS could be classified in IGF2 high and IGF2 low-expressing cell lines. Allelic IGF2 transcript quantification analysis revealed that in part of the RA FLS the normally suppressed allele was activated, resulting in biallelic expression of the IGF2 gene. Biallelic expression was associated with increased levels of IGF2 mRNA production. CONCLUSION: The findings indicate that the imprinting status of IGF2 might underlie the increased expression of IGF2, which may contribute to autonomous growth of RA FLS of low inflammatory synovial tissues.

Transcription profiling of rheumatic diseases.

Rheumatic diseases are a diverse group of disorders. Most of these diseases are heterogeneous in nature and show varying responsiveness to treatment. Because our understanding of the molecular complexity of rheumatic diseases is incomplete and criteria for categorization are limited, we mainly refer to them in terms of group averages. The advent of DNA microarray technology has provided a powerful tool to gain insight into the molecular complexity of these diseases; this technology facilitates open-ended survey to identify comprehensively the genes and biological pathways that are associated with clinically defined conditions. During the past decade, encouraging results have been generated in the molecular description of complex rheumatic diseases, such as rheumatoid arthritis, systemic lupus erythematosus, Sjögren syndrome and systemic sclerosis. Here, we describe developments in genomics research during the past decade that have contributed to our knowledge of pathogenesis, and to the identification of biomarkers for diagnosis, patient stratification and prognostication.

Acute experimental colitis and human chronic inflammatory diseases share expression of inflammation-related genes with conserved Ets2 binding sites.

BACKGROUND: Ulcerative colitis (UC) and Crohn's disease (CD) are characterized by chronic inflammation of the gastrointestinal tract, with overlapping clinical characteristics and unknown etiology. We reasoned that in intestinal inflammation the initial activation of the innate immune response fails to resolve, finally resulting in uncontrolled chronic inflammatory bowel disease. METHODS: To identify the early inflammatory events in colitis that remain active in human chronic colitis, we analyzed the changes of the colonic transcriptome during acute experimental colitis and compared the outcome with previously published profiles of affected tissues from patients with UC and CD, and as a control for intestinal inflammation in general, tissues from celiac disease patients. Rheumatoid arthritis synovial tissues were included as a nonintestinal inflammatory disease. The expression profiles of each disease were analyzed separately, in which diseased tissues were compared to unaffected tissues from the same anatomical location. RESULTS: Gene ontology analysis of significantly regulated genes revealed a marked activation of immunity and defense processes in all diseases, except celiac disease, where immune activation is less prominent. The control region of upregulated genes contained an increase in Ets2 binding sites in experimental colitis, UC, and rheumatoid arthritis, and were associated with upregulated immune activity. In contrast, upregulated genes in celiac disease harbored the transcription factor binding site GLI, which binds to the Gli family of transcription factors involved in hedgehog signaling, affecting development and morphogenesis. CONCLUSION: Ets2 may be an important transcription factor driving inflammation in acute as well as chronic inflammatory disease