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Nagy, N.

Publications and source records attributed to Nagy, N..

At least 19 recordsLinked to original sources

Hyaluronan and CD44 targeting reverses early matrix changes, proinflammatory signals and fibrosis in primary sclerosing cholangitis

Primary sclerosing cholangitis (PSC) is a rare, progressive liver disease characterized by biliary inflammation and bile duct strictures and no approved medical therapy. Despite its clinical severity, the pathological mechanisms underlying PSC remain poorly understood, largely due to early diagnostic challenges. Here we provide complementary evidence in human PSC samples, transcriptomic data, mouse models, and 3D cholangiocyte cultures that underscore the importance of hyaluronic acid (HA) and its cognate receptor CD44 in PSC pathogenesis. HA is a glycosaminoglycan abundant in the extracellular matrix in inflammatory disorders, yet its role in PSC has not been well characterized. We demonstrate that in early-stage PSC, cholangiocytes aberrantly produce high molecular weight HA that accumulates in the peribiliary matrix, increasing local tissue stiffness. This mechanical signal is transduced by a CD44/Integrin {beta}1 receptor complex in cholangiocytes, driving cell proliferation, YAP mechanosignaling, pro-inflammatory cytokine production with a transition to a ductular reactive phenotype. CD44 knockdown in cholangiocyte cell lines and mouse models significantly lowered stiffness, and attenuated inflammation. Together, these findings reveal a mechano-inflammatory axis in which HA-driven matrix stiffening perpetuates biliary inflammation and disease progression, identifying HA targeting and CD44 as promising therapeutic strategies. One Sentence SummaryHyaluronan and CD44 mediate matrix changes and progressive fibrosis in primary sclerosing cholangitis.

pathology↗

Targeted depletion of CD38+ autoreactive T cells prevents type 1 diabetes

Type 1 diabetes (T1D) is caused by T cell-mediated autoimmune destruction of insulin-producing islet {beta}-cells. Treatment with T-cell depleting therapies delays the progression of stage 2 and 3 T1D, but these agents exert broad immunosuppressive effects on T cell populations, including T regulatory cells (Tregs), which are key in promoting immune tolerance. We evaluated non-obese diabetic (NOD) mice and recently diagnosed T1D patients and identified CD38 as a marker for pathogenic T cell populations. Using adoptive T-cell transfer in Recombination Activating Gene 1 knockout NOD mice and in a humanized mouse model of autoimmune diabetes, we demonstrated that CD38-expressing autoreactive T cells drive diabetes pathogenesis. Furthermore, we found that selective depletion of CD38+ cells, using an anti-CD38 monoclonal antibody (mAb), prevents insulitis and diabetes onset without depleting CD4+CD25+ Tregs. Administration of anti-CD38 mAb did not adversely affect islet function and may selectively eliminate immunogenic senescent islet {beta}-cells. These results support the strategy of selectively depleting diabetogenic T cells using an anti-CD38 mAb to treat T1D and restore immune tolerance. Therefore, transient depletion of autoreactive T cells using anti-CD38 mAb may provide a novel strategy to prevent or abrogate {beta}-cell autoimmunity in T1D. One sentence summaryPathogenic autoreactive are characterized by CD38 overexpression, and their selective depletion with anti-CD38 monoclonal antibody prevents autoimmune diabetes.

immunology↗

Donor-derived CD8+CD122+ Tregs generated in mixed donor chimeric NOD mice delete autoreactive T cells

The establishment of mixed hematopoietic chimerism is a promising way to induce immune tolerance for islet replacement therapy and to treat the underlying autoimmunity in type 1 diabetes (T1D). Mixed chimerism not only promotes effective thymic negative selection of autoreactive cells but also restores regulatory T cell (Treg) function and peripheral tolerance. In the current study, we determined that a novel class of donor-derived CD8+CD44+CD122+ Tregs (d-CD8+CD122+ Tregs) plays a crucial role in controlling autoimmunity in non-obese diabetic (NOD) mice with induced mixed chimerism. Using adoptive T cell transfer experiments, we showed that d-CD8+CD122+ Tregs abrogate autoimmunity by selectively depleting the exogenously injected diabetogenic T cells in Recombination-Activating Gene deficient NOD mice. These d-CD8+CD122+ Tregs from NOD chimeras show upregulation of Helios, Programmed cell death protein 1, perforin, granzyme-B, CD39, Folate receptor 4, and downregulation of proinflammatory markers like Scart1 and Scart2. Using in vitro assays, we show that d-CD8+CD122+ Tregs respond specifically to a Complementarity-Determining Region-3 peptide sequence derived from T cell receptors of islet antigen-specific autoreactive T cells. Similarly, we found that individuals with T1D have a deficiency in CD8+CD122+ Tregs, suggesting a potential loss of regulatory function accompanies disease onset. Revitalizing CD8+CD122+ Tregs may offer a new therapeutic strategy of restoring immune tolerance in autoimmune diabetes.

immunology↗

Recognizing dUTPase as a mitotic factor essential for early embryonic development

dUTPase is universally regarded as a metabolic sanitizing enzyme that protects genomes by preventing the incorporation of uracil into DNA. Despite its essentiality across eukaryotes, no function beyond nucleotide sanitization has been demonstrated. Here, we uncover a conserved, non-canonical role for dUTPase as a regulator of mitosis. Using Drosophila and mouse models, we demonstrate that dUTPase loss causes early embryonic lethality characterized by severe mitotic failure that, cannot be rescued by disabling uracil-DNA repair, uncoupling dUTPase essentiality from DNA repair pathways. Mechanistically, dUTPase dynamically associates with the mitotic spindle and centrosomes, and its depletion induces centrosome amplification and chromosome segregation defects. Beyond cell division, dUTPase dosage bidirectionally controls cell migration, linking its mitotic function to cellular behaviors relevant for metastasis. Together, our findings redefine dUTPase as a moonlighting mitotic factor that coordinates centrosome integrity and spindle dynamics, expanding its known repertoire beyond nucleotide metabolism. HighlightsdUTPase deficiency leads to mitotic defects during early embryonic development in both Drosophila and mouse models dUTPase shows dynamic spatiotemporal localization associated with microtubules through mitosis dUTPase is essential for the normal number and intracellular localization of centrosomes dUTPase deficiency counteracts while its overexpression enhances cell migration

molecular biology↗

T-World: A highly general computational model of a human ventricular myocyte

Cardiovascular disease is the leading cause of death, demanding new tools to improve mechanistic understanding and overcome limitations of stem cell and animal-based research. We introduce T-World, a highly general virtual model of human ventricular cardiomyocyte suitable for multiscale studies. T-World shows comprehensive agreement with human physiology, from electrical activation to contraction, and is the first to replicate all key cellular mechanisms driving life-threatening arrhythmias. Extensively validated on unseen data, it demonstrates strong predictivity across applications and scales. Using T-World we revealed a likely sex-specific arrhythmia risk in females related to restitution properties, identified arrhythmia drivers in type 2 diabetes, and describe unexpected pro-arrhythmic role of NaV1.8 in heart failure. T-World demonstrates strong performance in predicting drug-induced arrhythmia risk and opens new opportunities for predicting and explaining drug efficacy, demonstrated by unpicking effects of mexiletine in Long QT syndrome 2. T-World is available as open-source code and an online app.

cell biology↗

A549 tumorigenic and BEAS-2B non-tumorigenic cell line derived small extracellular vesicles show distinct proteomic, N-glycoproteomic and chondroitin/dermatan sulfate profiles

Extracellular vesicles (EVs) are critical mediators of intercellular communication and hold promise as biomarkers and therapeutic targets in cancer, but their molecular alterations remain poorly understood. Protein glycosylation is a frequent post-translational modification; however, most EV studies focus only on proteomics, while mapping glycosylation changes of proteins are still underrepresented. To address this shortcoming, we analyzed the proteomic, N-glycoproteomic, and chondroitin/dermatan sulfate (CS/DS) glycosaminoglycan (GAG) profiles of small EVs (sEVs) derived from A549 lung adenocarcinoma and BEAS-2B non-tumorigenic epithelial cell lines. Principal component analysis and hierarchical clustering revealed that all three profiles are highly dependent on the origin of sEV, highlighting fundamental differences not only at the proteomic but also at the N-glycopeptide and CS/DS levels. Protein expression differences were primarily associated with the upregulation of cell cycle regulation, DNA repair, metabolism, and protein synthesis, while immune-related processes were predominantly downregulated. Proteomics revealed differential expressions of 5 CS proteoglycans, anticipating that their CS profile may also change. N-glycoproteomics highlighted a shift from complex to hybrid N-glycans in cancer sEVs, alongside a significant decrease in fucosylation. Prominent glycoproteins characterized with multiple glycosylation sites included versican, galectin-3-binding protein and laminins. The total amount of CS/DS increased 3.4-fold in cancer sEVs, while the ratio of the two monosulfated disaccharides changed 2-fold, suggesting altered sulfation mechanisms. These findings highlight the potential of N-glycoproteomics and GAG profiling to enhance biomarker discovery and EV-based cancer diagnostics. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/643059v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@c2dfdeorg.highwire.dtl.DTLVardef@1f79452org.highwire.dtl.DTLVardef@12a9dbborg.highwire.dtl.DTLVardef@d992da_HPS_FORMAT_FIGEXP M_FIG C_FIG Proteomic, N-glycoproteomic and chondroitin/dermatan sulfate disaccharide profiles differ between A549 lung adenocarcinoma and BEAS-2B non-tumorigenic epithelial cell derived small extracellular vesicles.

cancer biology↗

Endurance exercise remodels pulmonary vein sleeve myocytes and promotes a proarrhythmic atrial substrate

BACKGROUNDAtrial fibrillation (AF) susceptibility is heightened in endurance athletes but the underlying mechanisms are incompletely understood. Because pulmonary vein (PV) myocyte triggers are critical determinants of AF, we investigated PV electrophysiological remodelling in animal models of the athletes heart. METHODSThe following experiments were performed in canines and mice after 16 or 6 weeks, respectively, of daily exercise training (ExT), and compared to sedentary (Sed) controls: ECG recording, echocardiography, pharmacological autonomic block, extrastimulus pacing, multielectrode array mapping, monophasic and intracellular action potential (AP) recording with custom-designed pattern recognition analysis, histology, RNAseq and spatial in situ transcriptomics. RESULTSAF propensity was significantly increased in ExT animals. Mapping studies identified heightened rotational activity in the PV-left atrial (LA) junction of ExT vs. Sed canines in vivo, and enhanced automaticity, triggered activity and AP duration variability ex vivo in ExT canines and mice. Intracellular recordings in mouse PV cardiomyocytes determined at least six AP subtypes with increased frequency of pacemaker-like APs in ExT PV, concomitant with increased expression of pacemaking HCN4, Cav1.3 and Cav3.1 channels. PV spontaneous excitability was also significantly enhanced. Subcellular resolution spatial transcriptomics in mouse PV-LA identified diffuse ion channel remodelling and activation of established AF-promoting pro-inflammatory and pro-fibrotic cytokines and chemokines in ExT PV cardiomyocytes. Conduction slowing in the ExT PV-LA junction was attributable to: gap junction remodelling, reduced Na+ channel expression and increased extracellular matrix deposition with enhanced myofibroblast number and proximity to PV cardiomyocytes. CONCLUSIONSEndurance exercise elicits proarrhythmic electro-anatomical remodelling of the PV-LA junction with enhanced pacemaking ion channel expression and immune-inflammatory pathway activation in PV myocytes as prominent contributors. CLINICAL PERSPECTIVEO_ST_ABSWhat is new?C_ST_ABSO_LIThis work is the first demonstration that endurance training results in proarrhythmic electrophysiological remodelling of PV sleeve myocytes and extracellular matrix deposition in the PV-LA junction. C_LIO_LIWe register electrical and molecular heterogeneity of the PV-LA junction at single cell and subcellular resolution, and for the first time identify the molecular events that underlie increased proarrhythmic activity of the trained PV. These include enhanced pacemaking ion channel expression (e.g., HCN4, Cav1.3, and Cav3.1), pro-inflammatory cytokine activation (e.g., TNF, IL-6), increased myofibroblasts and extracellular matrix deposition. C_LI What are the clinical implications?O_LIWe identify the molecular determinants of PV proarrhythmic activity in the trained heart and present new therapeutic targets for AF prevention in athletes. C_LIO_LIOur findings provide mechanistic rationale for the efficacy of pulmonary vein isolation for AF in athletes. C_LI

physiology↗

Targeting non-canonical NF-κB signalling in CYLD cutaneous syndrome by selective inhibition of IκB kinase alpha.

CYLD cutaneous syndrome (CCS) skin tumors develop from puberty onwards, can number in the hundreds and progressively grow over time. CCS patients lack medical therapies and require repeated surgery to control tumor burden. CYLD loss of heterozygosity (LOH) drives tumor growth, and CCS tumors have previously been shown to demonstrate increased canonical NF-{kappa}B and Wnt signalling. Here, we demonstrate evidence of non-canonical NF-{kappa}B signalling in CCS tumor keratinocytes, with increased p100 to p52 processing and RelB protein expression compared to normal skin. Utilizing complementary transcriptomics and proteomics on patient derived CCS tumor cell fractions, we identify I{kappa}B kinase alpha (IKK) as a candidate target in the non-canonical NF-{kappa}B signalling pathway. A novel, highly selective, IKK inhibitor (SU1644) used in patient derived CCS tumor spheroid cultures demonstrated that IKK inhibition reduced tumor spheroid viability. These data provide the pre-clinical rationale for the assessment of topical IKK inhibitors as a novel preventative treatment for CCS. TeaserTopical IKK inhibition emerges as a potential therapy for CYLD cutaneous syndrome by targeting non-canonical NF-{kappa}B signalling Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/635629v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@829218org.highwire.dtl.DTLVardef@459f5dorg.highwire.dtl.DTLVardef@e198f6org.highwire.dtl.DTLVardef@1015685_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

Sympathetic Stimulation Can Compensate for Hypocalcaemia-Induced Bradycardia in Human and Rabbit Sinoatrial Node Cells

Regular activation of the heart originates from cyclic spontaneous depolarisations of sinoatrial node cells (SANC). Variations in electrolyte levels, commonly observed in haemodialysis (HD) patients, and the autonomic nervous system (ANS) profoundly affect the SANC function. Thus, we investigated the effects of hypocalcaemia and sympathetic stimulation on SANC beating rate (BR). The {beta}-adrenergic ({beta}-AR) signalling cascade, as described by Behar et al., was incorporated into the SANC models of Severi et al. (rabbit) and Fabbri et al. (human). Simulations were conducted across various extracellular calcium ([Ca2+]o) (0.6 to 1.8 mM) and isoprenaline concentrations [ISO] (0 to 1000 nM) for a sufficient time to allow transient oscillations to equilibrate and reach a limit cycle. The {beta}-AR cell response of the extended models was validated against new Langendorff-perfused rabbit heart experiments and literature data. The extended models revealed that decreased [Ca2+]o necessitated an exponential-like increase in [ISO] to restore the basal BR. Specifically, at 1.2 mM [Ca2+]o, the Severi and Fabbri model required 15.5 and 7.3 nM [ISO], respectively, to restore the initial BR. Further reduction of [Ca2+]o to 0.6 mM required 60.0 and 41.7 nM [ISO] to compensate for hypocalcaemia. A sudden loss of sympathetic tone at low [Ca2+]o resulted in extreme bradycardia or even loss of automaticity within seconds. These findings suggest that hypocalcaemic bradycardia can be compensated for by an elevated sympathetic tone. The integration of the {beta}-AR pathways led to a logarithmic BR increase and offers insights into potential pathomechanisms underlying sudden cardiac death (SCD) in HD patients. Key Points- We extended the sinoatrial node cell (SANC) models of Severi et al. (rabbit) and Fabbri et al. (human) with the {beta}-adrenergic ({beta}-AR) signalling cascade Behar et al. described. - Simulations were conducted across various extracellular calcium ([Ca2+]o) (0.6 to 1.8 mM) and isoprenaline concentrations [ISO] (0 to 1000 nM) to mimic conditions in haemodialysis patients. - An exponential-like increase in [ISO] compensated for hypocalcaemia-induced bradycardia in both models, while inter-species differences lead to more sensitivity of the extended Fabbri model towards hypocalcaemia and increased sympathetic tone. - The extended models may help to further understand the pathomechanisms of several cardiovascular diseases affecting pacemaking, such as the high occurrence of sudden cardiac death (SCD) in chronic kidney disease (CKD) patients.

cell biology↗

Inhibition of hyaluronan synthesis prevents β-cell loss in obesity-associated type 2 diabetes

Pancreatic {beta}-cell dysfunction and death are central to the pathogenesis of type 2 diabetes (T2D). We have identified a novel role for the inflammatory extracellular matrix polymer hyaluronan (HA) in this pathophysiology. Low levels of HA are present in healthy pancreatic islets. However, HA substantially accumulates in cadaveric islets of human T2D and islets of the db/db mouse model of T2D in response to hyperglycemia. Treatment with 4-methylumbelliferone (4-MU), an inhibitor of HA synthesis, or the deletion of the major HA receptor CD44, preserve glycemic control and insulin levels in db/db mice despite ongoing weight gain, indicating a critical role for this pathway in T2D pathogenesis. 4-MU treatment and the deletion of CD44 likewise preserve glycemic control in other settings of {beta}-cell injury including streptozotocin treatment and islet transplantation. Mechanistically, we find that 4-MU increases the expression of the apoptosis inhibitor survivin, a downstream transcriptional target of CD44 dependent on HA/CD44 signaling, on {beta}-cells such that caspase 3 activation does not result in {beta}-cell apoptosis. These data indicate a role for HA accumulation in diabetes pathogenesis and suggest that it may be a viable target to ameliorate {beta}-cell loss in T2D. These data are particularly exciting, because 4-MU is already an approved drug (also known as hymecromone), which could accelerate translation of these findings to clinical studies.

immunology↗

FOXP3+ regulatory T cells use heparanase to access IL-2 bound to ECM in inflamed tissues

FOXP3+ regulatory T cells (Treg) depend on exogenous IL-2 for their survival and function, but circulating levels of IL-2 are low, making it unclear how Treg access this critical resource in vivo. Here, we show that Treg use heparanase (HPSE) to access IL-2 sequestered by heparan sulfate (HS) within the extracellular matrix (ECM) of inflamed central nervous system tissue. HPSE expression distinguishes human and murine Treg from conventional T cells and is regulated by the availability of IL-2. HPSE-/- Treg have impaired stability and function in vivo, including the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis. Conversely, endowing Treg with HPSE enhances their ability to access HS-sequestered IL-2 and their tolerogenic function in vivo. Together, these data identify novel roles for HPSE and the ECM in immune tolerance, providing new avenues for improving Treg-based therapy of autoimmunity. One-Sentence SummaryRegulatory T cells use heparanase to strip IL-2 bound to extracellular matrix within inflamed tissues, thereby supporting their homeostasis and function.

immunology↗

Remodeling of the Ribosomal Quality Control and Protein Translation by a Viral Ubiquitin Deconjugase

The strategies adopted by viruses to reprogram the protein translation and quality control machineries to promote infection are poorly understood. Here, we discovered that the viral ubiquitin deconjugase (vDUB) encoded in the large tegument protein of Epstein- Barr virus (EBV) regulates ribosomal stress responses. The vDUB participates in protein complexes that include the ubiquitin ligases ZNF598 and LTN1 and the UFM1 ligase UFL1. Upon ribosomal stalling, the vDUB counteracts the ubiquitination of 40S ribosome subunits, inhibits the degradation of translation-stalled polypeptides by the proteasome, and prevents UFMylation of the 60S particle, which impairs the ER-phagy- dependent clearance of stalled products. Inhibition of the ribosome quality control activates a GCN2-dependent integrated stress response that decreases global protein translation while promoting the readthrough of stall-inducing mRNAs. The vDUB enhances viral mRNAs translation and virus release during productive infection, pointing to a pivotal role in cell reprogramming that enables virus production and underlies the pathogenesis of EBV-associated cancers and autoimmune diseases. GRAPHIC SUMMARY O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/526464v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@1c7421dorg.highwire.dtl.DTLVardef@c59b8dorg.highwire.dtl.DTLVardef@1676b4eorg.highwire.dtl.DTLVardef@b5e111_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

The pharmacokinetics and pharmacodynamics of 4-methylumbelliferone and its glucuronide metabolite in mice

Hyaluronan (HA) is an extracellular matrix glycosaminoglycan, with important roles in chronic inflammation, cancer and autoimmunity. 4-methylumbelliferone (4-MU), a small molecule inhibitor of HA synthases, is widely used to study HAs interactions with the surrounding tissues and the immune cells. There is substantial experimental and therapeutic interest in using oral 4-MU to inhibit HA synthesis, but pharmacokinetic and pharmacodynamic data on treatment routes have been lacking. Moreover, it recently became clear that the main metabolite of 4-MU, 4-methlyumbelliferyl glucuronide (4-MUG), is bioactive. We therefore sought to define the pharmacokinetics and pharmacodynamics of 4-MU and its active metabolite 4-MUG in mice. Single dose mouse studies showed that 4-MU administered intravenously (i.v.) resulted in 100-fold higher 4-MU exposure compared to oral (p.o.) administration. The 4-MU ratio AUC i.v./AUC p.o. was 96/1. 4-MUG exposures were much higher than 4-MU exposures after both 4-MU i.v. and p.o. administration, but only small differences in 4-MUG exposure were seen after 4-MU i.v. versus p.o. administration. The 4-MUG metabolite was also administered as a single dose both i.v. and p.o. and showed a 25.9% bioavailability. Compared to 4-MUG p.o. dosing, 1.14 higher 4-MUG exposures were seen after 4-MU p.o. dosing. 4-MU exposure after 4-MUG p.o. administration was minimal but similar to 4-MU exposure after 4-MU p.o. administration. In mice treated for several weeks with 4-MU in chow, the 4-MU concentration immediately drops after treatment was stopped, whereas the 4-MUG concentration showed a peak 1 hour after treatment stop. In a build-up study, 4-MU and 4-MUG treatment in mice lead to a plateau of 4-MU concentration starting at 4 days post treatment start. These 4-MU and 4-MUG concentration findings in vivo will inform future clinical studies and experimental work with 4-MU.

pharmacology and toxicology↗

Cardiac electrophysiological remodeling associated with enhanced arrhythmia susceptibility in a canine model of elite exercise

The health benefits of regular physical exercise are well known. Even so, there is increasing evidence that the exercise regimes of elite athletes can evoke cardiac arrhythmias including ventricular fibrillation and even sudden cardiac death (SCD). The mechanism of exercise-induced arrhythmia and SCD is poorly understood. While some studies after endurance training have been performed in small animals these have limited translation value. Here, we show that chronic training in a canine model (12 sedentary and 12 trained dogs) that mimics the regime of elite athletes induces electrophysiological remodeling (measured by ECG, patch-clamp and immunocytochemical techniques) resulting in increases of both the trigger and the substrate for ventricular arrhythmias. Thus, 4 months sustained training lengthened ventricular repolarization (QTc: 213.6{+/-}2.8 ms vs. 237.1{+/-}3.4 ms, n=12; APD90: 370.1{+/-}32.7 ms vs. 472.8{+/-}29.6 ms, n=25 vs. 29), decreased transient outward potassium current (8.8{+/-}0.9 pA/pF vs. 6.4{+/-}0.5 pA/pF at 50 mV, n=42 vs. 54) and increased the short term variability of repolarization (17.5{+/-}4.0 ms vs. 29.5{+/-}3.8 ms, n=18 vs. 27). Left ventricular fibrosis and HCN4 protein expression were also enhanced. These changes were associated with enhanced ectopic activity (number of extrasystoles: 4/hour vs. 366/hour) in vivo and arrhythmia susceptibility (elicited ventricular fibrillation: 3 of 10 sedentary dogs vs. 6 of 10 trained dogs). Our findings provide in vivo, cellular electrophysiological and molecular biological evidence for the enhanced susceptibility to ventricular arrhythmia in an experimental large animal model of endurance training.

physiology↗

TALPID3/KIAA0586 regulates multiple aspects of neuromuscular patterning during gastrointestinal development in animal models and human

TALPID3/KIAA0586 is an evolutionary conserved protein, which plays an essential role in protein trafficking. Its role during gastrointestinal (GI) and enteric nervous system (ENS) development has not been studied previously. Here, we analysed chicken, mouse and human embryonic GI tissues with TALPID3 mutations. The GI tract of TALPID3 chicken embryos was shortened and malformed. Histologically, the gut smooth muscle was mispatterned and enteric neural crest cells were scattered throughout the gut wall. Analysis of the Hedgehog pathway and gut extracellular matrix provided causative reasons for these defects. Interestingly, chicken intra-species grafting experiments and a conditional knockout mouse model showed that ENS formation did not require TALPID3, but was dependent on correct environmental cues. Surprisingly, the lack of TALPID3 in enteric neural crest cells (ENCC) affected smooth muscle and epithelial development in a non cell-autonomous manner. Analysis of human gut fetal tissues with a KIAA0586 mutation showed strikingly similar findings compared to the animal models demonstrating conservation of TALPID3 and its necessary role in human GI tract development and patterning

developmental biology↗

TLR2 on CD4+ and CD8+ T cells promotes late control of Mycobacterium tuberculosis infection

Although a role for TLR2 on T cells has been indicated in prior studies, in vivo stimulation of TLR2 on T cells by Mtb and its impact on Mtb infection has not been tested. Furthermore, it is not known if the enhanced susceptibility to Mtb of Tlr2 gene knockout (ko) mice is due to its role in macrophages, on T cells or both. To address TLR2 on T cells, we generated Tlr2fl/flxCd4cre/cre mice, which lack expression of TLR2 on both CD4 and CD8 T cells, to study the in vivo role of TLR2 on T cells after aerosol infection with virulent Mtb. Deletion of TLR2 in CD4+ and CD8+ T cells reduces their ability to be co-stimulated by TLR2 ligands for cytokine production. These include both pro-(IFN-{gamma}, TNF-) and anti-inflammatory cytokines (IL-10). Deletion of TLR2 in T cells did not affect early control but did result in decreased late control of Mtb in the lungs of infected mice. This suggests that T cell co-stimulation by mycobacterial TLR2 ligands in vivo is important for control of infection during the chronic phase of Mtb infection in the lung.

immunology↗

The Epstein-Barr virus ubiquitin deconjugase BPLF1 regulates the activity of Topoisomerase II during virus replication

Topoisomerases are essential for the replication of herpesviruses but the mechanisms by which the viruses hijack the cellular enzymes are largely unknown. We found that topoisomerase-II (TOP2) is a substrate of the Epstein-Barr virus (EBV) ubiquitin deconjugase BPLF1. BPLF1 selectively inhibited the ubiquitination of TOP2 following treatment with topoisomerase poisons, interacted with TOP2 and TOP2{beta} in co-immunoprecipitation and in vitro pull-down, stabilized Etoposide-trapped TOP2 cleavage complexes (TOP2cc) and promoted TOP2 SUMOylation, which halted the DNA-damage response and reduced Etoposide toxicity. Induction of the productive virus cycle promoted the accumulation of TOP2{beta}cc, enhanced TOP2{beta} SUMOylation, and reduced Etoposide toxicity in lymphoblastoid cell lines carrying recombinant EBV encoding the active enzyme. Attenuation of this phenotype upon expression of a catalytic mutant BPLF1-C61A impaired viral DNA synthesis and virus release. These findings highlight a previously unrecognized function of BPLF1 in promoting non-proteolytic pathways for TOP2cc debulking that favor cell survival and virus production.

microbiology↗

Hyaluronan Synthesis Inhibition Impairs Antigen Presentation and Delays Transplantation Rejection

A coat of pericellular hyaluronan surrounds mature dendritic cells (DC) and contributes to cell-cell interactions. We asked whether 4-methylumbelliferone (4MU), an oral inhibitor of HA synthesis, could inhibit antigen presentation. We find that 4MU treatment reduces pericellular hyaluronan, destabilizes interactions between DC and T-cells, and prevents T-cell proliferation in vitro and in vivo. These effects were observed only when 4MU was added prior to initial antigen presentation but not later, consistent with 4MU-mediated inhibition of de novo antigenic responses. Building on these findings, we find that 4MU delays rejection of allogeneic pancreatic islet transplant and allogeneic cardiac transplants in mice and suppresses allogeneic T-cell activation in human mixed lymphocyte reactions. We conclude that 4MU, an approved drug, may have benefit as an adjunctive agent to delay transplantation rejection.

immunology↗