Prediction of underlying atrial fibrillation in patients with a cryptogenic stroke: results from the NOR-FIB Study.

BACKGROUND: Atrial fibrillation (AF) detection and treatment are key elements to reduce recurrence risk in cryptogenic stroke (CS) with underlying arrhythmia. The purpose of the present study was to assess the predictors of AF in CS and the utility of existing AF-predicting scores in The Nordic Atrial Fibrillation and Stroke (NOR-FIB) Study. METHOD: The NOR-FIB study was an international prospective observational multicenter study designed to detect and quantify AF in CS and cryptogenic transient ischaemic attack (TIA) patients monitored by the insertable cardiac monitor (ICM), and to identify AF-predicting biomarkers. The utility of the following AF-predicting scores was tested: AS5F, Brown ESUS-AF, CHA2DS2-VASc, CHASE-LESS, HATCH, HAVOC, STAF and SURF. RESULTS: In univariate analyses increasing age, hypertension, left ventricle hypertrophy, dyslipidaemia, antiarrhythmic drugs usage, valvular heart disease, and neuroimaging findings of stroke due to intracranial vessel occlusions and previous ischemic lesions were associated with a higher likelihood of detected AF. In multivariate analysis, age was the only independent predictor of AF. All the AF-predicting scores showed significantly higher score levels for AF than non-AF patients. The STAF and the SURF scores provided the highest sensitivity and negative predictive values, while the AS5F and SURF reached an area under the receiver operating curve (AUC) > 0.7. CONCLUSION: Clinical risk scores may guide a personalized evaluation approach in CS patients. Increasing awareness of the usage of available AF-predicting scores may optimize the arrhythmia detection pathway in stroke units.

Atrial fibrillation in cryptogenic stroke and TIA patients in The Nordic Atrial Fibrillation and Stroke (NOR-FIB) Study: Main results.

Introduction: Secondary stroke prevention depends on proper identification of the underlying etiology and initiation of optimal treatment after the index event. The aim of the NOR-FIB study was to detect and quantify underlying atrial fibrillation (AF) in patients with cryptogenic stroke (CS) or transient ischaemic attack (TIA) using insertable cardiac monitor (ICM), to optimise secondary prevention, and to test the feasibility of ICM usage for stroke physicians. Patients and methods: Prospective observational international multicenter real-life study of CS and TIA patients monitored for 12 months with ICM (Reveal LINQ) for AF detection. Results: ICM insertion was performed in 91.5% by stroke physicians, within median 9 days after index event. Paroxysmal AF was diagnosed in 74 out of 259 patients (28.6%), detected early after ICM insertion (mean 48 ± 52 days) in 86.5% of patients. AF patients were older (72.6 vs 62.2; p < 0.001), had higher pre-stroke CHA2DS2-VASc score (median 3 vs 2; p < 0.001) and admission NIHSS (median 2 vs 1; p = 0.001); and more often hypertension (p = 0.045) and dyslipidaemia (p = 0.005) than non-AF patients. The arrhythmia was recurrent in 91.9% and asymptomatic in 93.2%. At 12-month follow-up anticoagulants usage was 97.3%. Discussion and conclusions: ICM was an effective tool for diagnosing underlying AF, capturing AF in 29% of the CS and TIA patients. AF was asymptomatic in most cases and would mainly have gone undiagnosed without ICM. The insertion and use of ICM was feasible for stroke physicians in stroke units.

Underlying causes of cryptogenic stroke and TIA in the nordic atrial fibrillation and stroke (NOR-FIB) study - the importance of comprehensive clinical evaluation.

BACKGROUND: Cryptogenic stroke is a heterogeneous condition, with a wide spectrum of possible underlying causes for which the optimal secondary prevention may differ substantially. Attempting a correct etiological diagnosis to reduce the stroke recurrence should be the fundamental goal of modern stroke management. METHODS: Prospective observational international multicenter study of cryptogenic stroke and cryptogenic transient ischemic attack (TIA) patients clinically monitored for 12 months to assign the underlying etiology. For atrial fibrillation (AF) detection continuous cardiac rhythm monitoring with insertable cardiac monitor (Reveal LINQ, Medtronic) was performed. The 12-month follow-up data for 250 of 259 initially included NOR-FIB patients were available for analysis. RESULTS: After 12 months follow-up probable stroke causes were revealed in 43% patients, while 57% still remained cryptogenic. AF and atrial flutter was most prevalent (29%). In 14% patients other possible causes were revealed (small vessel disease, large-artery atherosclerosis, hypercoagulable states, other cardioembolism). Patients remaining cryptogenic were younger (p < 0.001), had lower CHA2DS2-VASc score (p < 0.001) on admission, and lower NIHSS score (p = 0.031) and mRS (p = 0.016) at discharge. Smoking was more prevalent in patients that were still cryptogenic (p = 0.014), while dyslipidaemia was less prevalent (p = 0.044). Stroke recurrence rate was higher in the cryptogenic group compared to the group where the etiology was revealed, 7.7% vs. 2.8%, (p = 0.091). CONCLUSION: Cryptogenic stroke often indicates the inability to identify the cause in the acute phase and should be considered as a working diagnosis until efforts of diagnostic work up succeed in identifying a specific underlying etiology. Timeframe of 6-12-month follow-up may be considered as optimal. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT02937077, EudraCT 2018-002298-23.

Randomized controlled trial of intra-articular ketorolac on pain and inflammation after minor arthroscopic knee surgery.

BACKGROUND: Ketorolac is an effective non-steroidal anti-inflammatory drug, commonly used with local anaesthetics as part of local infiltration analgesia protocols following orthopaedic surgery. However, systemic uptake and drug action may be the major mechanism after local infiltration. The aims of this project were to study the effects of a small, systemically ineffective dose of ketorolac given intra-articularly for post-operative pain and also to study synovial inflammatory biomarkers. We investigated whether ketorolac affects pro-inflammatory biomarkers in an in vitro model, as well. METHODS: In this placebo-controlled, blind, randomized study, we analysed intra-articular ketorolac (5 mg) in ambulatory minor knee surgery patients with moderate or severe pain (n = 44). We assessed post-operative pain intensity (n = 44) and analysed microdialysis samples taken from knee synovial tissue every 20 min (n = 34). We also tested cyclooxygenase-independent effects of ketorolac in synovial cells stimulated by prostaglandin E2 and chondroitin sulphate in vitro. RESULTS: Intra-articular ketorolac (5 mg) administration did not reduce pain or synovial pro-inflammatory cytokines CXCL1, IL-8, and MCP-1, 0-120 min after knee arthroscopy. Female gender was a risk factor for moderate or severe pain (relative risk 1.45, 95% confidence interval 1.04-2.01). Paradoxically, ketorolac increased the release of CXCL1 and IL-8 in prostaglandin E2 and chondroitin sulphate-stimulated synovial cells in vitro. CONCLUSION: Ketorolac prescribed at a low dose intra-articularly does not produce any detectable analgesic effect after minor knee surgery.

Increased expression of chemokines in patients with Wegener's granulomatosis - modulating effects of methylprednisolone in vitro.

Chemokines, a group of cytokines that attracts and activates leucocyte subpopulations in inflamed tissue, have been associated with the pathogenesis of a number of inflammatory diseases, and some recent reports have suggested their involvement in Wegener's granulomatosis (WG). To elucidate further the possible role of chemokines in WG we examined serum levels of several CC- and CXC-chemokines in WG patients and assessed the ability of corticosteroids to modulate the expression of these mediators in vitro. Our main findings were: (i) WG patients (n = 14) had elevated serum levels of several inflammatory chemokines [i.e. regulated upon activation normal T cell expressed and secreted (RANTES), monocyte chemoattractant protein (MCP)-1 and interleukin (IL)-8] compared to healthy controls (n = 9), as assessed by enzyme immunoassays (EIAs); (ii) by using EIAs and real-time reverse transcription-polymerase chain reaction (RT-PCR), we demonstrated the ability of methylprednisolone (MP) to down-regulate both the spontaneous and the staphylococcal enterotoxin B (SEB)-induced release of chemokines from peripheral blood mononuclear cells (PBMC) in vitro in both WG patients and controls, possibly involving both transcriptional and post-transcriptional mechanisms; and (iii) the ability of MP to attenuate chemokine secretion was less pronounced in WG patients than in controls, particularly with regard to inhibition of spontaneous release. Our findings suggest a role for chemokines in the pathogenesis of WG. The diminished MP-mediated suppression of chemokines in PBMC from WG patients suggests that more specific modulators of chemokine levels should be investigated in this disorder.

Expression of matrix metalloproteinases in patients with Wegener's granulomatosis.

BACKGROUND: Enhanced activity of matrix metalloproteinases (MMPs) has been reported to have a pathogenic role in several diseases such as cancer and cardiovascular disorders, and seems also to play a part in certain autoimmune diseases. OBJECTIVE: To examine whether enhanced MMP activity may also have a role in the pathogenesis of Wegener's granulomatosis (WG). METHODS: In a study group of 15 patients with WG and 15 controls, plasma levels and gene expression were measured in freshly isolated peripheral blood mononuclear cells (PBMCs) of several MMPs and their endogenous inhibitors (that is, tissue inhibitors of metalloproteinases (TIMPs)) by enzyme immunoassays and RNase protection assay, respectively. RESULTS: Whereas patients with WG in remission had enhanced gene expression of several MMPs and TIMPs in PBMCs, those with active disease had a selective up regulation of MMP-2 and MMP-8 compared with healthy controls, and a down regulation of TIMP-1 and TIMP-3 compared with other patients with WG. Moreover, plasma levels of TIMP-1 and MMP-8 correlated significantly with C reactive protein levels, further supporting an association between activation of the MMP/TIMP system and disease activity in WG. Finally, these changes in MMP/TIMP expression in WG were accompanied by increased total MMP activity in PBMC supernatants, particularly in those with active disease, suggesting a matrix degrading net effect. CONCLUSION: These findings suggest that disturbed MMP and TIMP activity has a role in the pathogenesis of WG.

Gene expression analysis of peripheral T cells in a subgroup of common variable immunodeficiency shows predominance of CCR7(-) effector-memory T cells.

Common variable immunodeficiency (CVID) represents a heterogeneous group of antibody deficiency syndromes, characterized by defective antibody production in which T cell deficiency may play a pathogenic role. A subgroup of CVID patients has impaired in vitro T cell proliferation. Using microarray analyses of T cells from these patients, we found a gene expression pattern different from healthy controls and patients with X-linked agammaglobulinaemia. The profile of the differentially expressed genes suggests enhanced cytotoxic effector functions, antigen experienced or chronically activated T cells and a predominance of CCR7(-) T cells. Further experiments using flow cytometry revealed a striking predominance of CCR7(-) T cells in a subgroup of CVID patients, and an association with impaired T cell proliferation. Our observations indicate that a predominance of CCR7(-) T cells with effector-memory cell features and with reduced proliferative capacity may characterize a subgroup of CVID.

Raised serum levels of interleukin-18 is associated with disease progression and may contribute to virological treatment failure in HIV-1-infected patients.

To gain further insight into the possible role of interleukin (IL)-18 in HIV-1 infection we examined serum levels of IL-18 in various clinical and immunological stages of HIV-1 infection during cross-sectional (n = 41) and longitudinal testing (n = 20) and during HAART (n = 21, 24 months follow-up). Our main findings were that HIV-1-infected patients had significantly raised IL-18 levels comparing healthy controls, particularly in those with advanced disease, that while HAART induced a marked decline in IL-18, virological treatment failure was associated with persistently raised IL-18 levels during such therapy and that our in vitro experiments showed an IL-18-mediated up-regulation of the HIV-1 coreceptor CXCR4 and the pro-apoptotic mediator TRAIL in PBMC from HIV-1-infected patients receiving HAART. HIV-1 infection appears to be characterized by persistently raised IL-18 levels and during HAART, such a pattern was associated with virological treatment failure, possibly contributing to immunodeficiency and HIV-1 replication in these patients.

Effects of interferon-alpha on gene expression of chemokines and members of the tumour necrosis factor superfamily in HIV-infected patients.

We examined the effect of interferon (IFN)-alpha on the expression of 375 genes relevant to inflammatory and immunological reactions in peripheral blood mononuclear cells (PBMC) from HIV-infected patients by cDNA expression array and real-time quantitative RT-PCR. Our main findings were: (i) IFN-alpha induced up-regulation of several genes in the tumour necrosis factor (TNF) superfamily including the ligands APRIL, FasL, TNF-alpha and TRAIL, with particularly enhancing effects on the latter in HIV-infected patients. (ii) While IFN-alpha markedly up-regulated the expression of anti-angionetic ELR- CXC-chemokines (e.g. MIG and IP-10), it suppressed the expression of angiogenic ELR+ CXC-chemokines (e.g. GRO-alpha, IL-8 and ENA-78), with similar patterns in both patients and controls. (iii) IFN-alpha induced a marked increase in gene expression of the HIV co-receptor CCR5 in both patients and controls. We suggest that these effects may contribute to both the therapeutic and toxic effects of IFN-alpha. Moreover, our findings underscore that the biological effects of IFN-alpha in HIV infection are complex and that the clinical net effects of IFN-alpha treatment may be difficult to predict. However, the potent enhancing effect of IFN-alpha on several pro-apoptotic genes in the TNF superfamily and the enhancing effect on CCR5 expression suggest a possible pathogenic role of IFN-alpha in the progression of HIV-related immunodeficiency and suggests caution in the therapeutic use of IFN-alpha in HIV-infected -individuals.

Myocardial gene expression of leukaemia inhibitory factor, interleukin-6 and glycoprotein 130 in end-stage human heart failure.

BACKGROUND: Studies in different animal models and plasma analyses in humans suggest that members of the interleukin-6 (IL-6) cytokine family may be involved in the pathogenesis of congestive heart failure (CHF). Accordingly, we have examined IL-6-related cytokines in chronic CHF in humans by analysing gene and protein expression in myocardium derived from patients with end-stage heart failure and donor hearts. METHODS: Gene expression of cytokines/receptors of the IL-6 family was documented in myocardial samples using cDNA array hybridization and RNase protection assays. Immunohistochemistry was used to detect leukaemia inhibitory factor (LIF), IL-6 and glycoprotein 130 (gp130) in myocardial tissues. RESULTS: Myocardial gene activity was documented for the majority of IL-6 family cytokines and their receptors. Immunohistochemical analysis localized IL-6, LIF and their common receptor subunit gp130 to myocytes and vascular smooth muscle cells. LIF mRNA levels were enhanced in the left ventricles of CHF patients relative to the left ventricles of donor hearts (patients 4.6 +/- 4.7 vs. donors 0.3 +/- 0.3, P < 0.005). Myocardial IL-6 and gp130 mRNA levels were not statistically different between patients and donors, but in contrast to LIF mRNA expression in heart explants, gp130 mRNA levels were significantly higher in left atrium compared with left ventricle in both patients and donors. CONCLUSIONS: Both mRNA and proteins of gp130 and its ligands IL-6 and LIF are expressed in both nonfailing and failing human myocardium. The elevated LIF mRNA levels in left ventricles from patients with end-stage heart failure suggest a role for LIF in the pathogenesis of CHF.

Relation between food provocation and systemic immune activation in patients with food intolerance.

We found that food provocation in food intolerant patients was characterised by a general and systemic immune activation accompanied by an increase in systemic symptoms. Our findings might be important for the understanding of the mechanisms involved in the pathogenesis of food intolerance.

Myocardial expression of CC- and CXC-chemokines and their receptors in human end-stage heart failure.

OBJECTIVES: Chemokines regulate several biological processes, such as chemotaxis, collagen turnover, angiogenesis and apoptosis. Based on the persistent immune activation with elevated circulating levels of chemokines in patients with congestive heart failure (CHF), we have hypothesised a pathogenic role for chemokines in the development of CHF. The objective of this study was to examine mRNA levels and cellular localisation of chemokines and chemokine receptors in human CHF. METHODS: We examined explanted hearts from ten patients with end-stage heart failure (all chambers) and in ten organ donors using an RNase protection assays and immunohistochemical techniques. RESULTS: Our main findings were: (i) expression of eight chemokine and nine chemokine receptor genes in both failing and nonfailing myocardium, (ii) particularly high mRNA levels of monocyte chemoattractant protein (MCP)-1 and CXC-chemokine receptor 4 (CXCR4), in both chronic failing and nonfailing myocardium, (iii) decreased mRNA levels of MCP-1 and interleukin (IL)-8 in the failing left ventricles compared to failing left atria, (iv) decreased chemokine (e.g., MCP-1 and IL-8) and increased chemokine receptor (e.g., CCR2, CXCR1) mRNA levels in failing left ventricles and failing left atria compared to corresponding chambers in the nonfailing hearts and (v) immunolocalisation of MCP-1, IL-8 and CXCR4 to cardiomyocytes. CONCLUSION: The present study demonstrates for the first time chemokine and chemokine receptor gene expression and protein localisation in the human myocardium, introducing a new family of mediators with potentially important effects on the myocardium. The observation of chemokine dysregulation in human end-stage heart failure may represent a previously unknown mechanism involved in progression of chronic heart failure.

Dysregulation of membrane-bound tumor necrosis factor-alpha and tumor necrosis factor receptors on mononuclear cells in human immunodeficiency virus type 1 infection: low percentage of p75-tumor necrosis factor receptor positive cells in patients with advanced disease and high viral load.

The correlation of persistent tumor necrosis factor-alpha (TNF-alpha) activation with disease progression in patients infected with human immunodeficiency virus type 1 (HIV-1), suggests a role for TNF-alpha in the pathogenesis of HIV-1 infection. In the present study, we examined by flow cytometry the expression of membrane-bound (m) components of the TNF system in 33 HIV-1-infected patients and 12 healthy controls. While peripheral blood mononuclear cells (PBMC) from asymptomatic and symptomatic non-acquired immune deficiency syndrome (AIDS) patients showed a significantly increased percentage of mTNF-alpha+ and mTNF receptor (TNFR)+ cells compared with controls, this was not found in the AIDS group. Compared with healthy controls, AIDS patients had a significantly decreased percentage of both monocytes and lymphocytes expressing p75-TNFR. PBMC from AIDS patients showed a higher p75-TNFR mRNA level and a higher spontaneous release of soluble p75-TNFR than healthy individuals, suggesting enhanced cell surface turnover of this TNFR. The low expression of TNFRs on both lymphocytes and monocytes in the AIDS group was associated with high numbers of HIV-1 RNA copies in plasma, low numbers of CD4+ lymphocytes, and high serum levels of soluble TNFRs. AIDS patients had a decreased percentage of CD8+ lymphocytes expressing TNFRs compared with healthy controls. In contrast, these patients, as well as symptomatic non-AIDS patients, had an increased percentage of TNF-alpha+ and TNFRs+ cells among remaining CD4+ lymphocytes. The pattern of abnormalities seen in AIDS patients suggests a role for persistent activation of the TNF system in the accelerated CD4+ lymphocyte destruction, the enhanced HIV-1 replication, and the markedly impaired antimicrobial defense in advanced HIV-1-related disease.

Effects of intravenous immunoglobulin in vivo on abnormally increased tumor necrosis factor-alpha activity in human immunodeficiency virus type 1 infection.

The effect of a single bolus injection (0.4 g/kg) of intravenous immunoglobulin (IVIG) on the tumor necrosis factor (TNF) system in human immunodeficiency virus type 1 (HIV-1)-infected patients was investigated. At 140 h after infusion, there was a significant decrease in levels of TNF-alpha and a significant increase in levels of soluble TNF receptors (sTNFR) in both plasma and lipopolysaccharide-stimulated peripheral blood mononuclear cells (PBMC). A rapid (within 1 h) decline in expression of membrane-bound TNF-alpha and p55-TNFR on PBMC persisted throughout the study. In contrast, there was an increased expression of membrane-bound p75-TNFR after 140 h. IVIG administration also resulted in significantly increased numbers of circulating CD4 lymphocytes, correlated with down-regulation of TNF-alpha activity in PBMC supernatants. Thus, down-regulation of the abnormally increased TNF-alpha activity may be achieved by IVIG administration. Studies evaluating the possible therapeutic role of long-term TNF-alpha suppression by IVIG may be warranted in HIV-1-infected patients.