Myo-differentiation reporter screen reveals NF-Y as an activator of PAX3-FOXO1 in rhabdomyosarcoma.

Recurrent chromosomal rearrangements found in rhabdomyosarcoma (RMS) produce the PAX3-FOXO1 fusion protein, which is an oncogenic driver and a dependency in this disease. One important function of PAX3-FOXO1 is to arrest myogenic differentiation, which is linked to the ability of RMS cells to gain an unlimited proliferation potential. Here, we developed a phenotypic screening strategy for identifying factors that collaborate with PAX3-FOXO1 to block myo-differentiation in RMS. Unlike most genes evaluated in our screen, we found that loss of any of the three subunits of the Nuclear Factor Y (NF-Y) complex leads to a myo-differentiation phenotype that resembles the effect of inactivating PAX3-FOXO1. While the transcriptomes of NF-Y- and PAX3-FOXO1-deficient RMS cells bear remarkable similarity to one another, we found that these two transcription factors occupy nonoverlapping sites along the genome: NF-Y preferentially occupies promoters, whereas PAX3-FOXO1 primarily binds to distal enhancers. By integrating multiple functional approaches, we map the PAX3 promoter as the point of intersection between these two regulators. We show that NF-Y occupies CCAAT motifs present upstream of PAX3 to function as a transcriptional activator of PAX3-FOXO1 expression in RMS. These findings reveal a critical upstream role of NF-Y in the oncogenic PAX3-FOXO1 pathway, highlighting how a broadly essential transcription factor can perform tumor-specific roles in governing cellular state.

The Cellular and Molecular Basis of Translational Immunometabolism.

The immune response requires major changes to metabolic processes, and indeed, energy metabolism and functional activation are fully integrated in immune cells to determine their ability to divide, differentiate, and carry out effector functions. Immune cell metabolism has therefore become an attractive target area for therapeutic purposes. A neglected aspect in the translation of immunometabolism is the critical connection between systemic and cellular metabolism. Here, we discuss the importance of understanding and manipulating the integration of systemic and immune cell metabolism through in-depth analysis of immune cell phenotype and function in human metabolic diseases and, in parallel, of the effects of conventional metabolic drugs on immune cell differentiation and function. We examine how the recent identification of selective metabolic programs operating in distinct immune cell subsets and functions has the potential to deliver tools for cell- and function-specific immunometabolic targeting.

Prevalence of self-reported severe periodontitis: Data from the population-based CONSTANCES cohort.

AIM: To assess the prevalence of severe periodontitis based on the population-based CONSTANCES cohort using a validated self-reported questionnaire. MATERIALS AND METHODS: Individuals were selected from the adult population in France using a random sampling scheme. Analyses were restricted to those invited in 2013-2014 who completed the periodontal health questionnaire at the 2017 follow-up. The risk of severe periodontitis was assessed using the periodontal screening score (PESS) and weighting coefficients were applied to provide representative results in the general French population. RESULTS: The study included 19,859 participants (9204 men, mean age: 52.8 ± 12.6 years). Based on a PESS ≥ 5, 7106 participants were at risk of severe periodontitis, corresponding to a weighted prevalence of 31.6% (95% confidence interval: 30.6%-32.7%). This prevalence was higher among participants aged 55 and over, those with lower socio-economic status as well as current smokers, e-cigarette users and heavy drinkers. Among individuals at risk of severe periodontitis, only 18.8% (17.3%-20.4%) thought they had gum disease, although 50.5% (48.6%-52.5%) reported that their last dental visit was less than 6 months. CONCLUSIONS: The present survey indicates that (1) self-reported severe periodontitis is highly prevalent with marked disparities between groups in the general French adult population, and (2) periodontitis could frequently be under-diagnosed given the low awareness.

Thrombopoietin-Dependent Myelo-Megakaryopoiesis Fuels Thromboinflammation and Worsens Antibody-Mediated Chronic Renal Microvascular Injury.

SIGNIFICANCE STATEMENT: Kidney-derived thrombopoietin (TPO) increases myeloid cell and platelet production during antibody-mediated chronic kidney disease (AMCKD) in a mouse model, exacerbating chronic thromobinflammation in microvessels. The effect is mirrored in patients with extracapillary glomerulonephritis associated with thromboinflammation, TGF ß -dependent glomerulosclerosis, and increased bioavailability of TPO. Neutralization of TPO in mice normalized hematopoiesis, reduced chronic thromboinflammation, and ameliorated renal disease. The findings suggest that TPO is a relevant biomarker and a promising therapeutic target for patients with CKD and other chronic thromboinflammatory diseases.Neutralization of TPO in mice normalized hematopoiesis, reduced chronic thromboinflammation, and ameliorated renal disease. The findings suggest that TPO is a relevant biomarker and a promising therapeutic target for patients with CKD and other chronic thromboinflammatory diseases. BACKGROUND: Chronic thromboinflammation provokes microvascular alterations and rarefaction, promoting organ dysfunction in individuals with various life-threatening diseases. Hematopoietic growth factors (HGFs) released by the affected organ may sustain emergency hematopoiesis and fuel the thromboinflammatory process. METHODS: Using a murine model of antibody-mediated chronic kidney disease (AMCKD) and pharmacological interventions, we comprehensively monitored the response to injury in the circulating blood, urine, bone marrow, and kidney. RESULTS: Experimental AMCKD was associated with chronic thromboinflammation and the production of HGFs, especially thrombopoietin (TPO), by the injured kidney, which stimulated and skewed hematopoiesis toward myelo-megakaryopoiesis. AMCKD was characterized by vascular and kidney dysfunction, TGF ß -dependent glomerulosclerosis, and microvascular rarefaction. In humans, extracapillary glomerulonephritis is associated with thromboinflammation, TGF ß -dependent glomerulosclerosis, and increased bioavailability of TPO. Analysis of albumin, HGF, and inflammatory cytokine levels in sera from patients with extracapillary glomerulonephritis allowed us to identify treatment responders. Strikingly, TPO neutralization in the experimental AMCKD model normalized hematopoiesis, reduced chronic thromboinflammation, and ameliorated renal disease. CONCLUSION: TPO-skewed hematopoiesis exacerbates chronic thromboinflammation in microvessels and worsens AMCKD. TPO is both a relevant biomarker and a promising therapeutic target in humans with CKD and other chronic thromboinflammatory diseases.

Protein instability associated with AARS1 and MARS1 mutations causes trichothiodystrophy.

Trichothiodystrophy (TTD) is a rare hereditary neurodevelopmental disorder defined by sulfur-deficient brittle hair and nails and scaly skin, but with otherwise remarkably variable clinical features. The photosensitive TTD (PS-TTD) forms exhibits in addition to progressive neuropathy and other features of segmental accelerated aging and is associated with impaired genome maintenance and transcription. New factors involved in various steps of gene expression have been identified for the different non-photosensitive forms of TTD (NPS-TTD), which do not appear to show features of premature aging. Here, we identify alanyl-tRNA synthetase 1 and methionyl-tRNA synthetase 1 variants as new gene defects that cause NPS-TTD. These variants result in the instability of the respective gene products alanyl- and methionyl-tRNA synthetase. These findings extend our previous observations that TTD mutations affect the stability of the corresponding proteins and emphasize this phenomenon as a common feature of TTD. Functional studies in skin fibroblasts from affected individuals demonstrate that these new variants also impact on the rate of tRNA charging, which is the first step in protein translation. The extension of reduced abundance of TTD factors to translation as well as transcription redefines TTD as a syndrome in which proteins involved in gene expression are unstable.

Periodontitis and atherosclerotic cardiovascular disease: A critical appraisal.

In spite of intensive research efforts driving spectacular advances in terms of prevention and treatments, cardiovascular diseases (CVDs) remain a leading health burden, accounting for 32% of all deaths (World Health Organization. "Cardiovascular Diseases (CVDs)." WHO, February 1, 2017, https://www.who.int/news-room/fact-sheets/detail/cardiovascular-diseases-(cvds)). Cardiovascular diseases are a group of disorders affecting the heart and blood vessels. They encompass a collection of different conditions, among which atherosclerotic cardiovascular disease (ASCVD) is the most prevalent. CVDs caused by atherosclerosis, that is, ASCVD, are particularly fatal: with heart attack and stroke being together the most prevalent cause of death in the world. To reduce the health burden represented by ASCVD, it is urgent to identify the nature of the "residual risk," beyond the established risk factors (e.g., hypertension) and behavioral factors already maximally targeted by drugs and public health campaigns. Remarkably, periodontitis is increasingly recognized as an independent cardiovascular risk factor.

Bi-allelic TARS Mutations Are Associated with Brittle Hair Phenotype.

Brittle and "tiger-tail" hair is the diagnostic hallmark of trichothiodystrophy (TTD), a rare recessive disease associated with a wide spectrum of clinical features including ichthyosis, intellectual disability, decreased fertility, and short stature. As a result of premature abrogation of terminal differentiation, the hair is brittle and fragile and contains reduced cysteine content. Hypersensitivity to UV light is found in about half of individuals with TTD; all of these individuals harbor bi-allelic mutations in components of the basal transcription factor TFIIH, and these mutations lead to impaired nucleotide excision repair and basal transcription. Different genes have been found to be associated with non-photosensitive TTD (NPS-TTD); these include MPLKIP (also called TTDN1), GTF2E2 (also called TFIIEß), and RNF113A. However, a relatively large group of these individuals with NPS-TTD have remained genetically uncharacterized. Here we present the identification of an NPS-TTD-associated gene, threonyl-tRNA synthetase (TARS), found by next-generation sequencing of a group of uncharacterized individuals with NPS-TTD. One individual has compound heterozygous TARS variants, c.826A>G (p.Lys276Glu) and c.1912C>T (p.Arg638∗), whereas a second individual is homozygous for the TARS variant: c.680T>C (p.Leu227Pro). We showed that these variants have a profound effect on TARS protein stability and enzymatic function. Our results expand the spectrum of genes involved in TTD to include genes implicated in amino acid charging of tRNA, which is required for the last step in gene expression, namely protein translation. We previously proposed that some of the TTD-specific features derive from subtle transcription defects as a consequence of unstable transcription factors. We now extend the definition of TTD from a transcription syndrome to a "gene-expression" syndrome.

Targeting DNA Damage Response and Replication Stress in Pancreatic Cancer.

BACKGROUND & AIMS: Continuing recalcitrance to therapy cements pancreatic cancer (PC) as the most lethal malignancy, which is set to become the second leading cause of cancer death in our society. The study aim was to investigate the association between DNA damage response (DDR), replication stress, and novel therapeutic response in PC to develop a biomarker-driven therapeutic strategy targeting DDR and replication stress in PC. METHODS: We interrogated the transcriptome, genome, proteome, and functional characteristics of 61 novel PC patient-derived cell lines to define novel therapeutic strategies targeting DDR and replication stress. Validation was done in patient-derived xenografts and human PC organoids. RESULTS: Patient-derived cell lines faithfully recapitulate the epithelial component of pancreatic tumors, including previously described molecular subtypes. Biomarkers of DDR deficiency, including a novel signature of homologous recombination deficiency, cosegregates with response to platinum (P < .001) and PARP inhibitor therapy (P < .001) in vitro and in vivo. We generated a novel signature of replication stress that predicts response to ATR (P < .018) and WEE1 inhibitor (P < .029) treatment in both cell lines and human PC organoids. Replication stress was enriched in the squamous subtype of PC (P < .001) but was not associated with DDR deficiency. CONCLUSIONS: Replication stress and DDR deficiency are independent of each other, creating opportunities for therapy in DDR-proficient PC and after platinum therapy.

CD31 as a Therapeutic Target in Atherosclerosis.

The potential of CD31 as a therapeutic target in atherosclerosis has been considered ever since its cloning in the 1990s, but the exact role played by this molecule in the biologic events underlying atherosclerosis has remained controversial, resulting in the stalling of any therapeutic perspective. Due to the supposed cell adhesive properties of CD31, specific monoclonal antibodies and recombinant proteins were regarded as blocking agents because their use prevented the arrival of leukocytes at sites of acute inflammation. However, the observed effect of those compounds likely resulted from the engagement of the immunomodulatory function of CD31 signaling. This was acknowledged only later though, upon the discovery of CD31's 2 intracytoplasmic tyrosine residues called immunoreceptor tyrosine inhibitory motifs. A growing body of evidence currently points at a therapeutic potential for CD31 agonists in atherothrombosis. Clinical observations show that CD31 expression is altered at the surface of leukocytes infiltrating unhealed atherothrombotic lesions and that the physiological immunomodulatory functions of CD31 are lost at the surface of blood leukocytes in patients with acute coronary syndromes. On the contrary, translational studies using candidate therapeutic molecules in laboratory animals have provided encouraging results: synthetic peptides administered to atherosclerotic mice as systemic drugs in the acute phases of atherosclerotic complications favor the healing of wounded arteries, whereas the immobilization of CD31 agonist peptides onto coronary stents implanted in farm pigs favors their peaceful integration within the coronary arterial wall.

Coronary stent CD31-mimetic coating favours endothelialization and reduces local inflammation and neointimal development in vivo.

AIMS: The rapid endothelialization of bare metal stents (BMS) is counterbalanced by inflammation-induced neointimal growth. Drug-eluting stents (DES) prevent leukocyte activation but impair endothelialization, delaying effective device integration into arterial walls. Previously, we have shown that engaging the vascular CD31 co-receptor is crucial for endothelial and leukocyte homeostasis and arterial healing. Furthermore, we have shown that a soluble synthetic peptide (known as P8RI) acts like a CD31 agonist. The aim of this study was to evaluate the effect of CD31-mimetic metal stent coating on the in vitro adherence of endothelial cells (ECs) and blood elements and the in vivo strut coverage and neointimal growth. METHODS AND RESULTS: We produced Cobalt Chromium discs and stents coated with a CD31-mimetic peptide through two procedures, plasma amination or dip-coating, both yielding comparable results. We found that CD31-mimetic discs significantly reduced the extent of primary human coronary artery EC and blood platelet/leukocyte activation in vitro. In vivo, CD31-mimetic stent properties were compared with those of DES and BMS by coronarography and microscopy at 7 and 28 days post-implantation in pig coronary arteries (n = 9 stents/group/timepoint). Seven days post-implantation, only CD31-mimetic struts were fully endothelialized with no activated platelets/leukocytes. At day 28, neointima development over CD31-mimetic stents was significantly reduced compared to BMS, appearing as a normal arterial media with the absence of thrombosis contrary to DES. CONCLUSION: CD31-mimetic coating favours vascular homeostasis and arterial wall healing, preventing in-stent stenosis and thrombosis. Hence, such coatings seem to improve the metal stent biocompatibility.

CD31 Mimetic Coating Enhances Flow Diverting Stent Integration into the Arterial Wall Promoting Aneurysm Healing.

BACKGROUND AND PURPOSE: Beyond aneurysmal occlusion, metallic flow diverters (FDs) can induce an adverse endovascular reaction due to the foreignness of metal devices, hampering FD endothelialization across the aneurysm neck, and arterial healing of intracranial aneurysms. Here, we evaluated the potential benefits of an FD coating mimicking CD31, a coreceptor critically involved in endothelial function and endovascular homeostasis, on the endothelialization of FDs implanted in vivo. METHODS: Nitinol FD (Silk Vista Baby) and flat disks were dip-coated with a CD31-mimetic peptide via an intermediate layer of polydopamine. Disks were used to assess the reaction of endothelial cells and blood elements in vitro. An aneurysm rabbit model was used to compare in vivo effects on the arterial wall of CD31-mimetic-coated (CD31-mimetic, n=6), polydopamine-coated (polydopamine, n=6), and uncoated FDs (bare, n=5) at 4 weeks post-FD implantation. In addition, long-term safety was assessed at 12 weeks. RESULTS: In vitro, CD31-mimetic coated disks displayed reduced adhesion of blood elements while favoring endothelial cell attachment and confluence, compared to bare and polydopamine disks. Strikingly, in vivo, the neoarterial wall formed over the CD31-mimetic-FD struts at the aneurysm neck was characteristic of an arterial tunica media, with continuous differentiated endothelium covering a significantly thicker layer of collagen and smooth muscle cells as compared to the controls. The rates of angiographic complete occlusion and covered branch arterial patency were similar in all 3 groups. CONCLUSIONS: CD31-mimetic coating favors the colonization of metallic endovascular devices with endothelial cells displaying a physiological phenotype while preventing the adhesion of platelets and leukocytes. These biological properties lead to a rapid and improved endothelialization of the neoarterial wall at the aneurysm neck. CD31-mimetic coating could therefore represent a valuable strategy for FD biocompatibility improvement and aneurysm healing.

Haemodynamic stress-induced breaches of the arterial intima trigger inflammation and drive atherogenesis.

AIMS: Inflammatory mediators, including blood cells and their products, contribute critically to atherogenesis, but the igniting triggers of inflammation remain elusive. Atherosclerosis develops at sites of flow perturbation, where the enhanced haemodynamic stress could initiate the atherogenic inflammatory process due to the occurrence of mechanic injury. We investigated the role of haemodynamic stress-induced breaches, allowing the entry of blood cells in the arterial intima, in triggering inflammation-driven atherogenesis. METHODS AND RESULTS: Human coronary samples isolated from explanted hearts, (n = 47) displayed signs of blood entry (detected by the presence of iron, ferritin, and glycophorin A) in the subintimal space (54%) as assessed by histology, immunofluorescence, high resolution episcopic microscopy, and scanning electron microscopy. Computational flow dynamic analysis showed that intimal haemorrhagic events occurred at sites of flow disturbance. Experimental carotid arteries from Apoe deficient mice showed discrete endothelial breaches and intimal haemorrhagic events specifically occurring at the site of flow perturbation, within 3 days after the exacerbation of the local haemodynamic stress. Endothelial tearing was associated with increased VCAM-1 expression and, within 7 days, substantial Ly6G+ leucocytes accumulated at the sites of erythrocyte-derived iron and lipids droplets accumulation, pathological intimal thickening and positive oil red O staining. The formation of fatty streaks at the sites of intimal breaches was prevented by the depletion of Ly6G+ leucocytes, suggesting that the local injury driven by haemodynamic stress-induced breaches triggers atherogenic inflammation. CONCLUSION: Haemodynamic-driven breaches of the arterial intima drive atherogenic inflammation by triggering the recruitment of leucocyte at sites of disturbed arterial flow.

GTF2E2 Mutations Destabilize the General Transcription Factor Complex TFIIE in Individuals with DNA Repair-Proficient Trichothiodystrophy.

The general transcription factor IIE (TFIIE) is essential for transcription initiation by RNA polymerase II (RNA pol II) via direct interaction with the basal transcription/DNA repair factor IIH (TFIIH). TFIIH harbors mutations in two rare genetic disorders, the cancer-prone xeroderma pigmentosum (XP) and the cancer-free, multisystem developmental disorder trichothiodystrophy (TTD). The phenotypic complexity resulting from mutations affecting TFIIH has been attributed to the nucleotide excision repair (NER) defect as well as to impaired transcription. Here, we report two unrelated children showing clinical features typical of TTD who harbor different homozygous missense mutations in GTF2E2 (c.448G>C [p.Ala150Pro] and c.559G>T [p.Asp187Tyr]) encoding the beta subunit of transcription factor IIE (TFIIEß). Repair of ultraviolet-induced DNA damage was normal in the GTF2E2 mutated cells, indicating that TFIIE was not involved in NER. We found decreased protein levels of the two TFIIE subunits (TFIIEα and TFIIEß) as well as decreased phosphorylation of TFIIEα in cells from both children. Interestingly, decreased phosphorylation of TFIIEα was also seen in TTD cells with mutations in ERCC2, which encodes the XPD subunit of TFIIH, but not in XP cells with ERCC2 mutations. Our findings support the theory that TTD is caused by transcriptional impairments that are distinct from the NER disorder XP.

Adipocytes orchestrate the formation of tertiary lymphoid organs in the creeping fat of Crohn's disease affected mesentery.

The formation of tertiary lymphoid organs (TLOs) is orchestrated by the stromal cells of tissues chronically submitted to inflammatory stimuli, in order to uphold specific adaptive immune responses. We have recently shown that the smooth muscle cells of the arterial wall orchestrate the formation of the TLOs associated with atherosclerosis in response to the local release of TNF-α. Observational studies have recently documented the presence of structures resembling TLOs the creeping fat that develops in the mesentery of patients with Crohn's disease (CD), an inflammatory condition combining a complex and as yet not elucidated infectious and autoimmune responses. We have performed a comprehensive analysis of the TLO structures in order to decipher the mechanism leading to their formation in the mesentery of CD patients, and assessed the effect of infectious and/or inflammatory inducers on the potential TLO-organizer functions of adipocytes. Quantitative analysis showed that both T and B memory cells, as well as plasma cells, are enriched in the CD-affected mesentery, as compared with tissue from control subjects. Immunohistochemistry revealed that these cells are concentrated within the creeping fat of CD patients, in the vicinity of transmural lesions; that T and B cells are compartmentalized in clearly distinct areas; that they are supplied by post-capillary high endothelial venules and drained by lymphatic vessels indicating that these nodules are fully mature TLOs. Organ culture showed that mesenteric tissue samples from CD patients contained greater amounts of adipocyte-derived chemokines and the use of the conditioned medium from these cultures in functional assays was able to actively recruit T and B lymphocytes. Finally, the production of chemokines involved in TLO formation by 3T3-L1 adipocytes was directly elicited by a combination of TNF-α and LPS in vitro. We therefore propose a mechanism in which mesenteric adipocyte, through their production of key chemokines in response to inflammatory/bacterial stimuli, may orchestrate the formation of functional TLOs developing in CD-affected mesentery.

Translational Relevance and Recent Advances of Animal Models of Abdominal Aortic Aneurysm.

Human abdominal aortic aneurysm (AAA) pathophysiology is not yet completely understood. In conductance arteries, the insoluble extracellular matrix, synthesized by vascular smooth muscle cells, assumes the function of withstanding the intraluminal arterial blood pressure. Progressive loss of this function through extracellular matrix proteolysis is a main feature of AAAs. As most patients are now treated via endovascular approaches, surgical AAA specimens have become rare. Animal models provide valuable complementary insights into AAA pathophysiology. Current experimental AAA models involve induction of intraluminal dilation (nondissecting AAAs) or a contained intramural rupture (dissecting models). Although the ideal model should reproduce the histological characteristics and natural history of the human disease, none of the currently available animal models perfectly do so. Experimental models try to represent the main pathophysiological determinants of AAAs: genetic or acquired defects in extracellular matrix, loss of vascular smooth muscle cells, and innate or adaptive immune response. Nevertheless, most models are characterized by aneurysmal stabilization and healing after a few weeks because of cessation of the initial stimulus. Recent studies have focused on ways to optimize existing models to allow continuous aneurysmal growth. This review aims to discuss the relevance and recent advances of current animal AAA models. VISUAL OVERVIEW: An online visual overview is available for this article.

Self-report assessment of severe periodontitis: Periodontal screening score development.

AIM: To derive from a validated questionnaire a periodontal screening score (PESS), intended as a user-friendly tool to identify individuals at risk of periodontitis in epidemiological studies. METHODS: A French 12-item self-reported questionnaire was developed by translating English questions previously used for periodontitis screening and surveillance. After a cognitive evaluation, the questionnaire was validated in a sample of 232 individuals (mean age: 46.1 ± 12.6 years) receiving full-mouth periodontal examination, including probing pocket depth and clinical attachment level recordings. Case definition was based on the American Academy of Periodontology/Centers for Disease Control and Prevention criteria. Logistic regression analyses and C-statistics were used to assess the validity and accuracy of the questionnaire and to develop the PESS. RESULTS: The sample was constituted of 109 individuals with severe periodontitis, who were compared with 123 individuals with no/moderate periodontitis. The questionnaire had moderate-to-high accuracy in identifying severe cases; the PESS (calculated on five self-report items, age, and smoking) showed a sensitivity of 78.9% and a specificity of 74.8%, with an area under the receiver operating characteristics curve of 0.821. CONCLUSION: The PESS represents a valuable and accurate tool to screen for severe periodontitis at the population level.

CD31 is a key coinhibitory receptor in the development of immunogenic dendritic cells.

CD31 is a transhomophilic tyrosine-based inhibitory motif receptor and is expressed by both dendritic cells (DCs) and T lymphocytes. Previous studies have established that the engagement of CD31 drives immune-inhibitory signaling in T lymphocytes, but the effect exerted by CD31 signaling in DCs remains elusive. Here, we show that CD31 is a key coinhibitory receptor on stimulated DCs, favoring the development of tolerogenic functions and finally resulting in T-cell tolerance. The disruption of CD31 signaling favored the immunogenic maturation and migration of resident DCs to the draining lymph nodes. In contrast, sustaining the CD31/SHP-1 signaling during DC maturation resulted in reduced NF-κB nuclear translocation, expression of costimulatory molecules, and production of immunogenic cytokines (e.g., IL-12, IL-6), whereas the expression of TGF-ß and IL-10 were increased. More importantly, CD31-conditioned DCs purified from the draining lymph nodes of ovalbumin-immunized mice favored the generation of antigen-specific regulatory T cells (CD25(+) forkhead box P3(+)) at the expense of effector (IFN-γ(+)) cells upon coculture with naive ovalbumin-specific CD4(+) T lymphocytes ex vivo. Finally, the adoptive transfer of CD31-conditioned myelin oligodendrocyte glycoprotein-loaded DCs carried immune tolerance against the subsequent development of MOG-induced experimental autoimmune encephalomyelitis in vivo. The key coinhibitory role exerted by CD31 on DCs highlighted by the present study may have important implications both in settings where the immunogenic function of DCs is desirable, such as infection and cancer, and in settings where tolerance-driving DCs are preferred, such as autoimmune diseases and transplantation.

Iron alters valvular interstitial cell function and is associated with calcification in aortic stenosis.

AIMS: Aortic valve stenosis (AS) is the most common valvulopathy and is characterized by inflammation, extracellular matrix (ECM) remodelling and calcification, causing a narrowing of the valve and the consequential obstruction of the cardiac outflow. Although intraleaflet haemorrhage is associated with AS progression, the mechanisms involved are not known. The aims of this study were to identify valvular iron in relation to pathological changes associated with AS and the effects on valvular interstitial cells (VIC) in terms of iron uptake and iron-induced responses. METHODS AND RESULTS: Valvular iron accumulation was detected by Perls' staining on aortic valve sections and shown to increase with the extent of calcification. Furthermore, qRT-PCR analysis revealed that iron-containing valve regions exhibited increased expression of genes involved in ECM remodelling and calcification. In addition, we demonstrate that iron transporters are regulated by pathways with major impact on AS and that VIC can take up and accumulate iron, which resulted in increased proliferation and decreased elastin production. CONCLUSION: Iron, which may accumulate in the aortic valve by means of intraleaflet haemorrhages, can be taken up by VIC in a pro-inflammatory environment and actively contribute to VIC proliferation, ECM remodelling and calcification. These findings suggest a possible mechanism through which iron uptake by VIC may favour AS progression.