Background and purposeThe aim was to study the prevalence of epilepsy in a hospital‐based systemic lupus erythematosus (SLE) cohort and to investigate the relationship between epilepsy and other manifestations of neuropsychiatric SLE (NPSLE).MethodsThe study population consisted of 440 SLE patients recruited from 1998 to 2012. An epilepsy‐screening questionnaire was sent to all patients, where those screening positive were invited to a neurological examination with documentation of NPSLE symptoms according to the American College of Rheumatology nomenclature. Occurrences of autoantibodies (double stranded DNAantibody, antinuclear antibody, lupus anticoagulant, Sjögren's syndrome A, Sjögren's syndrome B) and the antiphospholipid syndrome (APS) were tabulated.ResultsOut of 440 patients, 14% were dead and 2.7% were lost to follow‐up. The questionnaire was sent to 368 patients; 312 (85%) responded. Of these, 131 (42%) screened positive. Epilepsy was confirmed in 36 (11.5%), of whom 30 (83%) had focal onset. Ten (3.2%) patients had isolated or provoked seizures. Manifestations of NPSLE occurred in 50%. The rates of cerebrovascular disease and psychosis were elevated two‐ and three‐fold in NPSLE patients with epilepsy versus NPSLE patients without epilepsy, respectively (P = 0.001 and P = 0.0006). APS was more common in patients with epilepsy compared to epilepsy‐free SLE patients with or without NPSLE (P = 0.02). In 50% of patients with epilepsy, no other etiology than SLE was detected.ConclusionsA high prevalence of epilepsy in SLE patients is reported, with association to concurrent cerebrovascular disease, APS and psychosis. Our findings support the notion of a multifactorial background for epilepsy in SLE including both vascular disease and features consistent with autoimmunity.

Materials and methods of research.The study included 19 patients with primary a STEMI the average age was 59 6 17 years.All patients underwent PCI with reperfusion of the infarctionrelated coronary artery.The mean recanalization time was 5 (4, 6) h.Using EchoCG, final systolic volume (CSR), terminal diastolic volume (BWW), LV ejection fraction (LVEF) was msured admission, 3rd, 7th and 21st day of STEMI.The CD133þ cells level was determined by flow cytometry of venous blood samples in the same periods.There was a control group of healthy volunteers (6 males) the average age was 41 6 9 years (absolutely healthy).Results: Carrying out an individual analysis of CD 133þ cells level circulating in patients with acute myocardial infarction, they found a multidirectional, nonlinear dynamics of their content by the 3rd day of MI.Due to this dynamics, all patients were divided into 2 subgroups: the 1st subgroup consisted of the patients with initially low level and it's further increasing on the 3rd day of MI, the 2nd subgroup -patients with a decrease level to a 3-m day of illness.Intragroup and intergroup differences were revealed on different days of the disease (p <0.05).A positive correlation was established between the relative amount of CD133þ cells in patients of the 2nd subgroup on the 7th day of MI and LVEF on the 21st day (R ¼ 0.7, p ¼ 0.02).Also, a relationship was found between the low absolute amount of CD 133þ cells in the first day of acute myocardial infarction and signs of chronic heart failure (CHF) after 6 months of the disease (p <0.05).Namely: in patients of the first subgroup, 6 months after of acute MI, minimal clinical manifestations of CHF were observed.This fact confirms that a high release of CD133þ cells into the peripheral channel is associated with a favorable course and disease prognosis. Conclusion:We showed the response of marrow to acute myocardial infarction, as release of CD133þ cells into the blood.It was found that the high mobilization of these cells both on the 1st day of MI and during the whole observation period was associated with a favorable course and disease prognosis, according to improvement of clinical and echocardiographic parameters.

Immune checkpoint blockade (ICB) has been a breakthrough in the treatment of metastatic melanoma. But with only about 20–40% long-term responders and severe side-effects in about 12–17%, finding predictive markers for treatment response is of great interest.We prospectively assessed clinical data, haematologic parameters and freshly isolated peripheral blood mononuclear cells of 30 patients treated with ipilimumab (n = 21) and pembrolizumab (n = 9) prior to the first 4 cycles with ICB and before the first tumour assessment.We discovered that the baseline levels of CD45RO+CD8+ T cells significantly differed among the patients. Thirteen (43%) of our patients had normal baseline levels of CD45RO+CD8+ T cells, whereas 17 (57%) patients were low on CD45RO+CD8+ T cells. The baseline levels of CD45RO+CD8+ T cells correlated significantly with the response to ipilimumab but not pembrolizumab. Patients with baseline levels of lower/equal 25% of CD45RO+CD8+ T cells did not respond to treatment with ipilimumab. Phenotyping the CD8+ T cells in patients treated with ipilimumab revealed an activated HLA-DR+CD25− phenotype, implying antigen non-specific stimulation. The levels of the HLA-DR+CD25−CD8+ T cells were significantly higher in patients with a normal baseline of CD45RO+CD8+ T cells and even increased significantly during treatment. Furthermore, proliferation of melanoma antigen recognized by T cells 1 (MART-1)-specific CD8+ T cells was not observed. Patients with normal baseline levels of CD45RO+CD8+ T cells showed a significant longer overall survival when treated with ipilimumab but not pembrolizumab.Patients with normal baseline levels of CD45RO+CD8+ T cells respond significantly more frequently to treatment with ipilimumab and the CD8+ T cells appear to be antigen non-specifically activated. The baseline level of CD45RO+CD8+ T cells represents a promising factor as biomarker for the prediction of the response to ipilimumab.

Several lines of evidence imply early alterations in endocannabinoid and phosphodiesterase 10A (PDE10A) signaling in Huntington disease (HD). Using [18F]MK-9470 and [18F]JNJ42259152 small-animal positron emission tomography (PET), we investigated for the first time cerebral changes in type 1 cannabinoid (CB1) receptor binding and PDE10A levels in vivo in presymptomatic, early symptomatic, and late symptomatic HD (R6/2) mice, in relation to glucose metabolism ([18F]FDG PET), brain morphology (magnetic resonance imaging) and motor function. Ten R6/2 and 16 wild-type (WT) mice were investigated at 3 different time points between the age of 4 and 13 weeks. Parametric CB1 receptor and PDE10A images were anatomically standardized to Paxinos space and analyzed voxelwise. Volumetric microMRI imaging was performed to assess HD pathology. In R6/2 mice, CB1 receptor binding was decreased in comparison with WT in a cluster comprising the bilateral caudate-putamen, globus pallidus, and thalamic nucleus at week 5 (−8.1% ± 2.6%, p = 1.7 × 10−5). Longitudinal follow-up showed further progressive decline compared with controls in a cluster comprising the bilateral hippocampus, caudate-putamen, globus pallidus, superior colliculus, thalamic nucleus, and cerebellum (late vs. presymptomatic age: −13.7% ± 3.1% for R6/2 and +1.5% ± 4.0% for WT, p = 1.9 × 10−5). In R6/2 mice, PDE10A binding potential also decreased over time to reach significance at early and late symptomatic HD (late vs. presymptomatic age: −79.1% ± 1.9% for R6/2 and +2.1% ± 2.7% for WT, p = 1.5 × 10−4). The observed changes in CB1 receptor and PDE10A binding were correlated to anomalies exhibited by R6/2 animals in motor function, whereas no correlation was found with magnetic resonance imaging-based striatal volume. Our findings point to early regional dysfunctions in endocannabinoid and PDE10A signaling, involving the caudate-putamen and lateral globus pallidus, which may play a role in the progression of the disease in R6/2 animals. PET quantification of in vivo CB1 and/or PDE10A binding may thus be useful early biomarkers for HD. Our results also provide evidence of subtle motor deficits at earlier stages than previously described.

Background and aims We investigated the distribution of ischaemic stroke subtypes, classified according the Trial of Org 10 172 in Acute Stroke Treatment (TOAST) system, among patients with systemic lupus erythematosus (SLE). Genetic susceptibility in the signal transducer and activator of transcription factor 4 (STAT4) gene, defined by the single nucleotide polymorphism (SNP) rs10181656(G) were explored. Methods We identified 69/665 SLE patients with stroke. Medical charts were retrieved and brain, cardiac and vascular imaging at the time of stroke were examined. Classification was performed according to TOAST: large-artery atherosclerosis (LAA), cardioembolism (CE), small-artery occlusion (SAO), stroke of other determined aetiology (OC) and stroke of undetermined aetiology (UE). Occurrence of the anti-phospholipid syndrome (APS) was documented. Evaluators were blinded to genotypes. General population controls (n=658) and SLE patients free from previous cerebrovascular disease (n=517) were used as comparators. Results 56/69 patients with ischaemic stroke had charts with sufficient information for TOAST classification. Median age was 52 (17-84) years, 91% were female. All strokes classified as OC were attributed to APS. TOAST classification is presented in Table 1. Stroke of OE/APS and CE origin were associated with the STAT4 risk genotype as presented in Table 2. Conclusions The majority of ischaemic strokes among SLE patients were of APS or CE origin. These two subtypes were associated with genetic susceptibility in the STAT4 gene. Patients with APS associated strokes were remarkably young. STAT4 genotype could, in addition to antiphospholipid antibodies and echocardiography, add information about stroke risk and help identify patients who will benefit from prophylactic anticoagulation treatment.

Silk is a protein of interest to both biological and industrial sciences. The silkworm, Bombyx mori, forms this protein into strong threads starting from soluble silk proteins using a number of biochemical and physical cues to allow the transition from liquid to fibrous silk. A pH gradient has been measured along the gland, but the methodology employed was not able to precisely determine the pH at specific regions of interest in the silk gland. Furthermore, the physiological mechanisms responsible for the generation of this pH gradient are unknown. In this study, concentric ion selective microelectrodes were used to determine the luminal pH of B. mori silk glands. A gradient from pH 8.2 to 7.2 was measured in the posterior silk gland, with a pH 7 throughout the middle silk gland, and a gradient from pH 6.8 to 6.2 in the beginning of the anterior silk gland where silk processing into fibers occurs. The small diameter of the most anterior region of the anterior silk gland prevented microelectrode access in this region. Using a histochemical method, the presence of active carbonic anhydrase was identified in the funnel and anterior silk gland of fifth instar larvae. The observed pH gradient collapsed upon addition of the carbonic anhydrase inhibitor methazolamide, confirming an essential role for this enzyme in pH regulation in the B. mori silk gland. Plastic embedding of whole silk glands allowed clear visualization of the morphology, including the identification of four distinct epithelial cell types in the gland and allowed correlations between silk gland morphology and silk stages of assembly related to the pH gradient. B. mori silk glands have four different epithelial cell types, one of which produces carbonic anhydrase. Carbonic anhydrase is necessary for the mechanism that generates an intraluminal pH gradient, which likely regulates the assembly of silk proteins and then the formation of fibers from soluble silk proteins. These new insights into native silk formation may lead to a more efficient production of artificial or regenerated silkworm silk fibers.

The underlying cause of neutropenia may be difficult to determine due to similar clinical presentation in many neutropenic conditions. The neutrophil protein hCAP-18 (pro-LL-37) is a major component of neutrophil secondary granules and in this prospective study we assessed the use of hCAP-18 levels in blood plasma for differential diagnosis of neutropenic patients (n = 133) of various aetiologies. Plasma levels of hCAP-18 were determined using immunoblot and ELISA. Patients with severe congenital neutropenia (n = 23) presented with the lowest levels of plasma hCAP-18 and differential diagnostic accuracy revealed high sensitivity (100%) and specificity (98.8%) for hCAP-18 ELISA. The correlation coefficient of the hCAP-18 ELISA versus immunoblotting was (R = 0.831) and that of the peptide LL-37 ELISA versus immunoblotting was (R = 0.405) (P < 0.001). Plasma hCAP-18 levels thus displayed high diagnostic value in differential diagnosis of chronic neutropenia. Neutropenic patients with Shwachman-Diamond syndrome, Barth syndrome, Cohen syndrome, acute myeloid leukaemia and specific granule deficiency presented with reduced plasma hCAP-18 levels as well. The blood plasma level of hCAP-18 was thus low in conditions in which the neutrophil antibacterial propeptide hCAP-18 is deficient, i.e. severe congenital neutropenia and neutrophil-specific granule deficiency, and in conditions in which bone marrow myelopoiesis is negatively affected.

OBJECTIVE:Antimicrobial peptides (AMP) provide protection from infection by pathogenic microorganisms and restrict bacterial growth at epithelial surfaces to maintain mucosal homeostasis. In addition, they exert a significant anti-inflammatory activity. Here we analysed the anatomical distribution and biological activity of an orally administered AMP in the context of bacterial infection and host-microbial homeostasis. DESIGN:The anatomical distribution as well as antibacterial and anti-inflammatory activity of the endogenous AMP cryptdin 2 and the synthetic peptide Pep19-2.5 at the enteric mucosal surface were analysed by immunostaining, functional viability and stimulation assays, an oral Salmonella enterica subsp. enterica sv. Typhimurium (S. Typhimurium) model and comparative microbiota analysis. RESULTS:Endogenous cryptdin 2 was found attached to bacteria of the enteric microbiota within the intestinal mucus layer. Similarly, the synthetic peptide Pep19-2.5 attached rapidly to bacterial cells, exhibited a marked affinity for the intestinal mucus layer in vivo, altered the structural organisation of endotoxin in a mucus matrix and demonstrated potent anti-inflammatory and antibacterial activity. Oral Pep19-2.5 administration induced significant changes in the composition of the enteric microbiota as determined by high-throughput 16S rDNA sequencing. This may have contributed to the only transient improvement of the clinical symptoms after oral infection with S. Typhimurium. CONCLUSIONS:Our findings demonstrate the anti-inflammatory activity and mucus affinity of the synthetic AMP Pep19-2.5 and characterise the influence on microbiota composition and enteropathogen infection after oral administration.

Paneth cell-derived enteric antimicrobial peptides significantly contribute to antibacterial host defense and host-microbial homeostasis. Regulation occurs by enzymatic processing and release into the small intestinal lumen, but the stimuli involved are incompletely understood. Here, the capacity of various microbial and immune stimuli to induce antimicrobial peptide release from small intestinal tissue was systematically evaluated using antibacterial activity testing, immunostaining for Paneth cell granules and mass spectrometry. We confirmed the stimulatory activity of the muscarinic receptor agonist carbachol and the nucleotide-binding oligomerization domain ligand muramyl dipeptide. In contrast, no release of antibacterial activity was noted after treatment with the Toll-like receptor ligands poly(I:C), lipopolysaccharide or CpG, and the cytokines interleukin (IL)-15, IL-22, IL-28 and interferon-γ. Rapid Paneth cell degranulation and antimicrobial activity release, however, was observed after stimulation with the endogenous mediators IL-4 and IL-13. This process required phosphatidylinositol 3-kinase and was associated with protein kinase B phosphorylation in Paneth cells. Flow cytometric analysis confirmed expression of the IL-13 receptor α1 on isolated Paneth cells. Our findings identify a novel role of IL-13 as inducer of Paneth cell degranulation and enteric antimicrobial peptide release. IL-13 may thus contribute to mucosal antimicrobial host defense and host microbial homeostasis.

Paneth cell-derived enteric antimicrobial peptides significantly contribute to antibacterial host defense and host-microbial homeostasis. Regulation occurs by enzymatic processing and release into the small intestinal lumen, but the stimuli involved are incompletely understood. Here, the capacity of various microbial and immune stimuli to induce antimicrobial peptide release from small intestinal tissue was systematically evaluated using antibacterial activity testing, immunostaining for Paneth cell granules and mass spectrometry. We confirmed the stimulatory activity of the muscarinic receptor agonist carbachol and the nucleotide-binding oligomerization domain ligand muramyl dipeptide. In contrast, no release of antibacterial activity was noted after treatment with the Toll-like receptor ligands poly(I:C), lipopolysaccharide or CpG, and the cytokines interleukin (IL)-15, IL-22, IL-28 and interferon-γ. Rapid Paneth cell degranulation and antimicrobial activity release, however, was observed after stimulation with the endogenous mediators IL-4 and IL-13. This process required phosphatidylinositol 3-kinase and was associated with protein kinase B phosphorylation in Paneth cells. Flow cytometric analysis confirmed expression of the IL-13 receptor α1 on isolated Paneth cells. Our findings identify a novel role of IL-13 as inducer of Paneth cell degranulation and enteric antimicrobial peptide release. IL-13 may thus contribute to mucosal antimicrobial host defense and host microbial homeostasis.

Background Arthritis is known to associate with upregulation of inflammatory mediators in the central nervous system (CNS) as shown in earlier experimental studies (1). However, little is known about the possible presence of intrathecal immune activation in RA patients, which may have importance for cerebral symptoms, such as fatigue. Objectives Here we sought to investigate potential specific immune activation in RA CNS, focusing on the detection of anti-citrullinated protein antibodies (ACPA), known to be highly specific and predictive for the development of RA. Methods Thirteen female patients with ACPA-positive RA underwent lumbar puncture and collection of CSF and serum. CSF from 10 age-matched individuals with no neuroinflammatory or rheumatic disease, and 26 age-matched patients with multiple sclerosis (MS), were included as controls. IgG ACPAs were measured using a commercial anti-CCP2 ELISA assay (Euro-Diagnostica, Malmö, Sweden). Serum samples were diluted 1:100 according to the protocol, while CSF samples were used undiluted. Analyses of CSF albumin and CSF/plasma albumin ratios were performed in clinical routine. Results As expected, all RA patients were anti-CCP positive in serum (threshold for positivity: 25U/ml), with levels ranging from 34U/ml to 3200U/ml. Interestingly, 7/13 RA patients were anti-CCP positive also in CSF, with levels ranging from 29U/ml to 154U/ml in undiluted samples. Anti-CCP levels in controls, on the other hand, were significantly lower; 2.19±0.09U/ml for non-inflammatory controls, and 2.54±0.16U/ml for MS patients (undiluted samples). In RA patients, there was a correlation between presence of anti-CCP antibodies in CSF and significantly higher serum anti-CCP levels (1453±82U/ml in RA with CCP positivity both serum/CSF versus 90±46U/ml; CCP negative in CSF, p<0.01). No difference in CSF albumin or the CSF/plasma albumin ratio could be detected between these two RA groups. All RA patients had normal CSF albumine and CSF/plasma albumine ratios, consistent with an intact blood brain barrier. No RA patient had any history of, or symptoms of, neurologic disease. Conclusions Our data show, for the first time, the presence of ACPAs in CSF of RA patients. The association with high serum ACPA levels in these patients may suggest a leakage of antibodies, though data demonstrate intact blood brain barriers. Interestingly, certain ACPA specificities are known to recognize the citrullinated form of the neuronal antigen myelinic base protein (cit-MBP) (2). Moreover, endogenous cit-MBP is thought to be play an important role in the autoimmune mechanisms of multiple sclerosis (3). Thus, presence of autoantibodies like ACPAs in RA CSF may predispose for autoimmune reactions in the central nervous system, potentially affecting fatigue and other cerebral symptoms. References Inglis JJ et al, Arthritis Rheum 2007 Dec;56(12):4015-23. Ioan-Facsinay A et al, Ann Rheum Dis 2011;70:188–193 Moscarello MA et al, Neurochem Res (2007) 32:251–256 Disclosure of Interest None Declared

OBJECTIVE:The objective of this report is to assess the presence and viral load of JC polyomavirus (JCV) DNA in cerebrospinal fluid (CSF) and plasma from neuropsychiatric systemic lupus erythematosus (NPSLE) patients in comparison to controls and to investigate if different types of immunosuppressive treatments were correlated to detection and viral load of JCV DNA in SLE.BACKGROUND:Reactivation of a latent JCV infection with subsequent development of the fatal disease progressive multifocal leukoencephalopathy (PML) has become an increasing problem in patients with autoimmune diseases treated with newer immunosuppressants. Accumulating data point out that SLE patients are at particular risk for PML compared to patients with other rheumatic diseases.METHODS:CSF samples (n = 69) and plasma samples (n = 51) from 71 SLE patients and 58 controls (53 CSF samples and 50 plasma samples) with other non-inflammatory neurological disease (OND) were analyzed for JCV DNA with a quantitative PCR method.RESULTS:All CSF and plasma samples from NPSLE patients and controls were negative for JCV DNA.CONCLUSION:JCV DNA was absent in CSF and plasma in NPSLE patients and controls and consequently we were not able to identify any correlation between the occurrence of JCV DNA and type of immunosuppressive medication.