Impaired Butyrate Induced Regulation of T Cell Surface Expression of CTLA-4 in Patients with Ulcerative Colitis.

Patients with ulcerative colitis (UC) have reduced intestinal levels of short-chain fatty acids (SCFAs), including butyrate, which are important regulators of host-microbiota crosstalk. The aim was therefore to determine effects of butyrate on blood and intestinal T cells from patients with active UC. T cells from UC patients and healthy subjects were polyclonally stimulated together with SCFAs and proliferation, activation, cytokine secretion, and surface expression of cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) were analyzed. Butyrate induced comparable, dose dependent inhibition of activation and proliferation in blood T cells and activation in intestinal T cells from UC patients and healthy subjects. However, surface expression of the inhibitory molecule CTLA-4 on stimulated blood and intestinal T cells was impaired in UC patients and was not restored following butyrate treatment. Furthermore, unlike intestinal T cells from healthy subjects, butyrate was unable to downregulate secretion of interferon gamma (IFNγ), interleukin (IL)-4, IL-17A, and IL-10 in UC patients. Although seemingly normal inhibitory effects on T cell activation and proliferation, butyrate has an impaired ability to reduce cytokine secretion and induce surface expression of CTLA-4 in T cells from UC patients with active disease. Overall, these observations indicate a dysfunction in butyrate induced immune regulation linked to CTLA-4 signaling in T cells from UC patients during a flare.

Perioperative anesthetic management of a combined right atrial thrombectomy with living donor liver transplantation.

Hepatocellular carcinoma (HCC) with vascular invasion is usually considered inoperable. We describe a case of HCC with vascular invasion and right atrial thrombus that was successfully down staged. Patient underwent combined right atrial thrombectomy and living donor liver transplantation (LDLT) in the same setting. Perioperative anesthesia management and perioperative concerns of two major combined procedures are discussed.

Redox regulation of PTPN22 affects the severity of T-cell-dependent autoimmune inflammation.

Chronic autoimmune diseases are associated with mutations in PTPN22, a modifier of T cell receptor (TCR) signaling. As with all protein tyrosine phosphatases, the activity of PTPN22 is redox regulated, but if or how such regulation can modulate inflammatory pathways in vivo is not known. To determine this, we created a mouse with a cysteine-to-serine mutation at position 129 in PTPN22 (C129S), a residue proposed to alter the redox regulatory properties of PTPN22 by forming a disulfide with the catalytic C227 residue. The C129S mutant mouse showed a stronger T-cell-dependent inflammatory response and development of T-cell-dependent autoimmune arthritis due to enhanced TCR signaling and activation of T cells, an effect neutralized by a mutation in Ncf1, a component of the NOX2 complex. Activity assays with purified proteins suggest that the functional results can be explained by an increased sensitivity to oxidation of the C129S mutated PTPN22 protein. We also observed that the disulfide of native PTPN22 can be directly reduced by the thioredoxin system, while the C129S mutant lacking this disulfide was less amenable to reductive reactivation. In conclusion, we show that PTPN22 functionally interacts with Ncf1 and is regulated by oxidation via the noncatalytic C129 residue and oxidation-prone PTPN22 leads to increased severity in the development of T-cell-dependent autoimmunity.

Secretome screening reveals immunomodulating functions of IFNα-7, PAP and GDF-7 on regulatory T-cells.

Regulatory T cells (Tregs) are the key cells regulating peripheral autoreactive T lymphocytes. Tregs exert their function by suppressing effector T cells. Tregs have been shown to play essential roles in the control of a variety of physiological and pathological immune responses. However, Tregs are unstable and can lose the expression of FOXP3 and suppressive functions as a consequence of outer stimuli. Available literature suggests that secreted proteins regulate Treg functional states, such as differentiation, proliferation and suppressive function. Identification of secreted proteins that affect Treg cell function are highly interesting for both therapeutic and diagnostic purposes in either hyperactive or immunosuppressed populations. Here, we report a phenotypic screening of a human secretome library in human Treg cells utilising a high throughput flow cytometry technology. Screening a library of 575 secreted proteins allowed us to identify proteins stabilising or destabilising the Treg phenotype as suggested by changes in expression of Treg marker proteins FOXP3 and/or CTLA4. Four proteins including GDF-7, IL-10, PAP and IFNα-7 were identified as positive regulators that increased FOXP3 and/or CTLA4 expression. PAP is a phosphatase. A catalytic-dead version of the protein did not induce an increase in FOXP3 expression. Ten interferon proteins were identified as negative regulators that reduced the expression of both CTLA4 and FOXP3, without affecting cell viability. A transcriptomics analysis supported the differential effect on Tregs of IFNα-7 versus other IFNα proteins, indicating differences in JAK/STAT signaling. A conformational model experiment confirmed a tenfold reduction in IFNAR-mediated ISG transcription for IFNα-7 compared to IFNα-10. This further strengthened the theory of a shift in downstream messaging upon external stimulation. As a summary, we have identified four positive regulators of FOXP3 and/or CTLA4 expression. Further exploration of these Treg modulators and their method of action has the potential to aid the discovery of novel therapies for both autoimmune and infectious diseases as well as for cancer.

Tissue Infiltrating LTi-Like Group 3 Innate Lymphoid Cells and T Follicular Helper Cells in Graves' and Hashimoto's Thyroiditis.

Background: Hashimoto's thyroiditis (HT) and Graves' disease (GD) are autoimmune thyroid disorders (AITDs). These conditions have been associated to abnormalities in circulating regulatory T cells (Tregs). We postulated that immune perturbations could be more pronounced at the thyroid tissue level. Methods: The phenotype of PBMCs and immune cells infiltrating thyroid tissue from 19 patients with HT, 21 patients with GD, and 30 controls has been analyzed by flow cytometry. Results: We report that blood and thyroid Treg cell subsets are similarly represented in all AITDs patients and controls. Increased Lymphoid tissue inducer (LTi)-like ILC3 and CXCR5+ PD-1hi CD4+ T follicular helper cells (Tfh) tissue-infiltrating cells, together with the prevalence of tertiary lymphoid structures (TLS) and germinal centers (GCs) represented a typical immune signature in all HT and 60% of GD patients. In the remaining group of GD patients, the absence of the aforementioned abnormalities was associated with a higher prevalence of ophthalmopathy. Conclusion: Tissue infiltrating Lymphoid Tissue inducer-like group 3 Innate Lymphoid cells and T follicular helper cells are increased in most thyroid autoimmune disease.

A Phenotypic Screening Approach Using Human Treg Cells Identified Regulators of Forkhead Box p3 Expression.

Regulatory T (Treg) cells, expressing the transcription factor forkhead box p3 (FOXP3), are the key cells regulating peripheral autoreactive T lymphocytes by suppressing effector T cells. FOXP3+ Treg cells play essential roles controlling immune responses in autoimmune diseases and cancer. Several clinical approaches (e.g., polyclonal expansion of Treg cells with anti-CD3 and anti-CD28 coated beads in the presence of drugs) are under evaluation. However, expression of FOXP3, recognized as the master regulator of Treg cells, in induced Treg cells have been shown to be instable, and molecular targets involved in regulating FOXP3 expression and Treg cell function have not been well-defined. Thus, new targets directly regulating FOXP3 expression and the expression of its downstream genes (e.g., cytotoxic T-lymphocyte-associated protein 4 (CTLA4)) have the potential to stabilize the Treg cell phenotype and function. This report describes the development of an automated medium-throughput 384-well plate flow cytometry phenotypic assay meauring the protein expression of FOXP3 and CTLA4 in human Treg cells. Screening a library of 4213 structurally diverse compounds allowed us to identify a variety of compounds regulating FOXP3 and CTLA4 expression. Further evaluation of these and related small molecules, followed by confirmation using siRNA-mediated gene knockdown, revealed three targets: euchromatic histone-lysine N-methyltransferase (EHMT2) and glycogen synthase kinase 3 alpha/beta (GSK3α/ß) as potent positive regulators of FOXP3 expression, and bromodomain and extra-terminal domain (BET) inhibitors as negative regulators of FOXP3 and CTLA4 expression. These targets have potential implications for establishing novel therapies for autoimmune diseases and cancer.

Comparison between transcatheter closure and minimally invasive surgery for fossa ovalis atrial septal defect: a single institutional experience.

BACKGROUND: Although, conventional surgical closure of atrial septal defect (ASD) provides excellent results with very low mortality and morbidity, it leaves the scar of incision and postoperative pain. Newer treatment modalities like minimal invasive surgery and percutaneous closure are being increasingly used nowadays where available. AIM: To compare the patient population, success, safety, and efficacy of transcatheter closure of ASD (Group A) with that of minimally invasive surgery (Port Access) (Group B). METHODS: In this retrospective non-randomized study, a record of a total of 640 patients with diagnosis of ASD secundum between May 1997 and October 2006 were reviewed. A total of 470 out of 640 patients were selected for transcatheter closure (Group A) while 170 patients were taken for surgical closure by minimally invasive port access surgery (Group B). The safety and efficacy of two groups was evaluated on the basis of morbidity and mortality, duration of intensive care unit (ICU) stay, total duration of hospital stay, post-procedural complications, residual sequel at time of discharge, and residual flow across the ASD. RESULTS: Success rate in two groups was 97.1% and 99.4%, respectively and had no statistically significant difference. Similarly major complication rate also had no difference in statistical significance (1.8% and 2.9% for Group A and B, respectively). Group B patients had longer hospital stay. A small but significant number of patients were not found suitable for device closure. This number is likely to decrease as experience with technique increases. Port access surgery is currently not possible in small children (femoral artery diameter 35 mm) due to difficulty in cannulation. CONCLUSION: Percutaneous device closure of ASD can be offered as a treatment option in suitable patients. Port access is minimally invasive and an equally safe and effective alternative choice in ASDs with deficient rim in patient with appropriate age and weight.

Human FOXP3+ T regulatory cell heterogeneity.

FOXP3-expressing CD4+ T regulatory (Treg) cells are instrumental for the maintenance of self-tolerance. They are also involved in the prevention of allergy, allograft rejection, foetal rejection during pregnancy and of exaggerated immune response towards commensal pathogens in mucosal tissues. They can also prevent immune responses against tumors and promote tumor progression. FOXP3-expressing Treg cells are not a homogenous population. The different subsets of Treg cells can have different functions or roles in the maintenance of immune homeostasis and can therefore be differentially targeted in the management of autoimmune diseases or in cancer. We discuss here how Treg cell subsets can be differentiated phenotypically, functionally and developmentally in humans.

Differential gene expression in human tissue resident regulatory T cells from lung, colon, and blood.

As we learn more about how immune responses occur in situ, it is becoming clear that each organ/tissue is characterized with its own anatomy and microenvironment which may affect and even determine the outcome of the immune responses. With emerging data from animal studies showing that regulatory T cells infiltrating non-lymphoid tissues exhibit unique phenotypes and transcriptional signatures and display functions beyond their well-established suppressive roles, there is an urgent need to explore the function of tissue Treg cells in humans. Here we characterized the transcriptome of Treg residing at the human mucosal tissue obtained from the normal area of cancer resections and their peripheral blood counterparts, identifying human lung and colon tissue Treg signature genes and their upstream regulators. Pathway analysis highlighted potential differences in the cross-talk between tissue Treg cells and other non-immune tissue-specific cell types. For example, genes associated with wnt pathway were differentially regulated in lung Treg cells compared to blood or colon indicating a potential role for lung Treg cells in epithelium repair and regeneration. Moreover, we identified several non-coding RNAs specifically expressed by tissue-resident Tregs. These results provide a comprehensive view of lung and colon tissue Treg transcriptional landscape.

Early hemodynamic performance of the Trifecta™ surgical bioprosthesis aortic valve in Indian patient population: 12 month outcomes of the EVEREST post-market study.

BACKGROUND: Indian patients undergoing surgical aortic valve replacement (SAVR) differ from western populations with respect to aortic annulus size and valve disease morphology. The purpose of this post-market, non-randomized observational study was to evaluate the early hemodynamic performance of the Trifecta™ bioprosthesis (Abbott, previously St. Jude Medical, Minneapolis, US) in an Indian patient population. METHODS: From January 2014 to September 2015, 100 patients (mean age 64.4 ± 7.1 years, 62% male) undergoing SAVR for valve disease (68% stenosis, 7% insufficiency, 25% mixed pathology) were enrolled across 10 centers in India. Patients implanted with a 19-27 mm Trifecta™ valve were eligible to participate and were prospectively followed for 12-months post-implantation. Echocardiographic hemodynamic performance was evaluated at pre-implant, pre-discharge and at 12-months by an independent core laboratory. Adverse events were adjudicated by the study sponsor. Functional status at 12-months was assessed according to NYHA classification. Continuous data was summarized using descriptive statistics (mean &standard deviation,) and categorical data was summarized using frequencies and percentages. RESULT: Ninety patients (mean age 64.5, 62.2% male) completed the 12-month follow up. Significant improvements in hemodynamic valve performance were reported in 81 patients with available echocardiographic data at 12 months. Compared to baseline at 12-month follow up visit, mean effective orifice area increased from 0.75cm2 to 1.61cm2 (p < 0.0001), mean pressure gradient reduced to 10.42 mmHg from 51.47 mmHg (p < 0.0001), cardiac output increased from 4.46 l/min to 4.85 l/min (P 0.9254). Compared to baseline, functional status improved by ≥1 NYHA class in 75% of patients at 12 months (95% Clopper-Pearson (Exact) confidence limit [64.6%, 83.6%]). No instances of early mortality (< 30 days from index procedure) or structural valve dysfunction were reported. CONCLUSION: In an Indian patient population, implantation of the Trifecta™ bioprosthesis is shown to be safe and associated with favorable early hemodynamic performance and improved functional status at 12 months. TRIAL REGISTRATION: The clinical study has been registered under Clinical Trial Registry-India ( http://www.ctri.nic.in ) and registration number is CTRI/2014/02/004434 registered on 25 February 2014 retrospectively registered.

Application of a phenotypic drug discovery strategy to identify biological and chemical starting points for inhibition of TSLP production in lung epithelial cells.

Thymic stromal lymphopoietin (TSLP) is a cytokine released by human lung epithelium in response to external insult. Considered as a master switch in T helper 2 lymphocyte (Th2) mediated responses, TSLP is believed to play a key role in allergic diseases including asthma. The aim of this study was to use a phenotypic approach to identify new biological and chemical starting points for inhibition of TSLP production in human bronchial epithelial cells (NHBE), with the objective of reducing Th2-mediated airway inflammation. To this end, a phenotypic screen was performed using poly I:C / IL-4 stimulated NHBE cells interrogated with a 44,974 compound library. As a result, 85 hits which downregulated TSLP protein and mRNA levels were identified and a representative subset of 7 hits was selected for further characterization. These molecules inhibited the activity of several members of the MAPK, PI3K and tyrosine kinase families and some of them have been reported as modulators of cellular phenotypic endpoints like cell-cell contacts, microtubule polymerization and caspase activation. Characterization of the biological profile of the hits suggested that mTOR could be a key activity involved in the regulation of TSLP production in NHBE cells. Among other targeted kinases, inhibition of p38 MAPK and JAK kinases showed different degrees of correlation with TSLP downregulation, while Syk kinase did not seem to be related. Overall, inhibition of TSLP production by the selected hits, rather than resulting from inhibition of single isolated targets, appeared to be due to a combination of activities with different levels of relevance. Finally, a hit expansion exercise yielded additional active compounds that could be amenable to further optimization, providing an opportunity to dissociate TSLP inhibition from other non-desired activities. This study illustrates the potential of phenotypic drug discovery to complement target based approaches by providing new chemistry and biology leads.

Altered regulation and expression of genes by BET family of proteins in COPD patients.

BACKGROUND: BET proteins (BRD2, BRD3, BRDT and BRD4) belong to the family of bromodomain containing proteins, which form a class of transcriptional co-regulators. BET proteins bind to acetylated lysine residues in the histones of nucleosomal chromatin and function either as co-activators or co-repressors of gene expression. An imbalance between HAT and HDAC activities resulting in hyperacetylation of histones has been identified in COPD. We hypothesized that pan-BET inhibitor (JQ1) treatment of BET protein interactions with hyperacetylated sites in the chromatin will regulate excessive activation of pro-inflammatory genes in key inflammatory drivers of alveolar macrophages (AM) in COPD. METHODS AND FINDINGS: Transcriptome analysis of AM from COPD patients indicated up-regulation of macrophage M1 type genes upon LPS stimulation. Pan-BET inhibitor JQ1 treatment attenuated expression of multiple genes, including pro-inflammatory cytokines and regulators of innate and adaptive immune cells. We demonstrated for the first time that JQ1 differentially modulated LPS-induced cytokine release from AM or peripheral blood mononuclear cells (PBMC) of COPD patients compared to PBMC of healthy controls. Using the BET regulated gene signature, we identified a subset of COPD patients, which we propose to benefit from BET inhibition. CONCLUSIONS: This work demonstrates that the effects of pan-BET inhibition through JQ1 treatment of inflammatory cells differs between COPD patients and healthy controls, and the expression of BET protein regulated genes is altered in COPD. These findings provide evidence of histone hyperacetylation as a mechanism driving chronic inflammatory changes in COPD.

Correction: Altered regulation and expression of genes by BET family of proteins in COPD patients.

[This corrects the article DOI: 10.1371/journal.pone.0173115.].

Cardiac herniation following atrial septal defect closure using port access surgery.

Cardiac herniation, a rare entity, is seen most commonly after traumatic rupture of the pericardium or following pneu-monectomy with partial pericardiactomy. It is rarely seen to occur following closure of a sinus venosus atrial septal defect through the port access technique. A review of the literature in English did not reveal any such case report.

Predictors of operative mortality following primary coronary artery bypass surgery.

BACKGROUND: Although quality assessment of coronary artery patients can be done by 30 days risk-adjusted operative mortality, it is still insufficient to study the outcome after primary coronary artery bypass graft surgery (CABG). In our study, we attempted to determine the factors, which can help predict operative mortality before and after CABG. METHODS: The study population consisted of 1000 prospective patients who underwent primary isolated CABG. Assessment was done by dividing the patients into two groups, i.e. non-survivors ( n= 12) and survivors ( n= 988). Data were analyzed using both univariate and multivariate models. RESULTS: On univariate analysis, recent acute myocardial infarction, intra-aortic balloon counterpulsation (IABC), left ventricular ejection fraction (LVEF) <25%, ventilator-associated pneumonia (VAP), tracheostomy, re-exploration, ventricular arrhythmias, low cardiac output (CO), multiple blood transfusions, post-operative renal dysfunction and longer intensive care unit and hospital stay were found as risk factors for mortality. Multivariate analysis showed that LVEF <25%,VAP, ventricular arrhythmias and low CO independently predicted mortality. Prior knowledge of these risk factors can help not only in predicting the outcome and the risks but also helps to plan the surgical and post-operative course of the patients to improve the morbidity and mortality. CONCLUSION: Our data suggest that operative mortality can be predicted prior to and after surgery considering factors such as LVEF, use of IABC, onset of ventricular arrhythmias and low CO.

Functional role of lipid rafts in CD20 activity?

CD20 is a B-lymphocyte-specific integral membrane protein, implicated in the regulation of transmembrane calcium conductance, cell-cycle progression and B-lymphocyte proliferation. CD20 is proposed to function as a SOCC (store-operated calcium channel). SOCCs are activated by receptor-stimulated calcium depletion of intracellular stores. Sustained calcium conductivity across the plasma membrane mediated by SOCC activity is required for long-term calcium-dependent processes, such as transcriptional control and gene expression. Cross-linking of CD20 by antibodies (e.g. Rituxan) has been reported to induce a rapid redistribution of CD20 into specialized microdomains at the plasma membrane, known as lipid rafts. Recruitment of CD20 into lipid rafts and its homo-oligomerization are suggested to be crucial for CD20 activity and regulation. This review outlines recent biochemical studies characterizing the role of CD20 in calcium signalling in B-lymphocytes and evaluates an engagement of lipid rafts in the regulation of CD20-mediated calcium conductivity.

Regulation of B lymphocytes in health and disease. Meeting review.

Signal transduction by the B cell receptor (BCR) is an absolute requirement for the selection and development of B lymphocytes at multiple checkpoints. Binding to antigen via the BCR is complemented by a co-stimulus delivered through accessory and co-stimulatory cell surface molecules that regulate the signalling threshold. In addition, identification of genes associated with immunodeficiency syndromes has highlighted the importance of genetic regulation, particularly in immunoglobulin class-switching and somatic hypermutation. A 1-day symposium organised by the Biochemical Society considered some of the recent advances in our understanding of the molecules and regulatory pathways involved in B lymphocyte activation, differentiation and survival and the health consequences when threshold settings malfunction.

Immunology, genetics and microbiota in the COPD pathophysiology: potential scope for patient stratification.

Chronic obstructive pulmonary disease (COPD) is characterized by sustained inflammation of the airways, leading to destruction of lung tissue and declining pulmonary function. Although smoking is the most obvious risk factor for COPD, only about 20% of smokers develop COPD and smoking cessation does not reverse progression of COPD, indicating that while smoking is an important cause or initiating factor, it is not the only driver of ongoing chronic inflammation and disease progression in COPD patients. We hypothesize that smoking-induced changes in lung microbiota, epithelial integrity and epigenetic control of gene expression result in autoantigen induction and perturbed immune regulation in genetically vunerable individuals. In our view, COPD patients may be stratified according to their immunological and inflammatory status related to specific changes in the lung microbiota (innate and adaptive immunity), presence of autoantigens (adaptive immunity: Th1-B-cell axis) and epigenetic modifications (inflammation and structural changes).