Sortilin enhances fibrosis and calcification in aortic valve disease by inducing interstitial cell heterogeneity.

AIMS: Calcific aortic valve disease (CAVD) is the most common valve disease, which consists of a chronic interplay of inflammation, fibrosis, and calcification. In this study, sortilin (SORT1) was identified as a novel key player in the pathophysiology of CAVD, and its role in the transformation of valvular interstitial cells (VICs) into pathological phenotypes is explored. METHODS AND RESULTS: An aortic valve (AV) wire injury (AVWI) mouse model with sortilin deficiency was used to determine the effects of sortilin on AV stenosis, fibrosis, and calcification. In vitro experiments employed human primary VICs cultured in osteogenic conditions for 7, 14, and 21 days; and processed for imaging, proteomics, and transcriptomics including single-cell RNA-sequencing (scRNA-seq). The AVWI mouse model showed reduced AV fibrosis, calcification, and stenosis in sortilin-deficient mice vs. littermate controls. Protein studies identified the transition of human VICs into a myofibroblast-like phenotype mediated by sortilin. Sortilin loss-of-function decreased in vitro VIC calcification. ScRNA-seq identified 12 differentially expressed cell clusters in human VIC samples, where a novel combined inflammatory myofibroblastic-osteogenic VIC (IMO-VIC) phenotype was detected with increased expression of SORT1, COL1A1, WNT5A, IL-6, and serum amyloid A1. VICs sequenced with sortilin deficiency showed decreased IMO-VIC phenotype. CONCLUSION: Sortilin promotes CAVD by mediating valvular fibrosis and calcification, and a newly identified phenotype (IMO-VIC). This is the first study to examine the role of sortilin in valvular calcification and it may render it a therapeutic target to inhibit IMO-VIC emergence by simultaneously reducing inflammation, fibrosis, and calcification, the three key pathological processes underlying CAVD.

Proprotein Convertase Subtilisin/Kexin 9 (PCSK9) Promotes Macrophage Activation via LDL Receptor-Independent Mechanisms.

BACKGROUND: Activated macrophages contribute to the pathogenesis of vascular disease. Vein graft failure is a major clinical problem with limited therapeutic options. PCSK9 (proprotein convertase subtilisin/kexin 9) increases low-density lipoprotein (LDL)-cholesterol levels via LDL receptor (LDLR) degradation. The role of PCSK9 in macrophage activation and vein graft failure is largely unknown, especially through LDLR-independent mechanisms. This study aimed to explore a novel mechanism of macrophage activation and vein graft disease induced by circulating PCSK9 in an LDLR-independent fashion. METHODS: We used Ldlr-/- mice to examine the LDLR-independent roles of circulating PCSK9 in experimental vein grafts. Adeno-associated virus (AAV) vector encoding a gain-of-function mutant of PCSK9 (rAAV8/D377Y-mPCSK9) induced hepatic PCSK9 overproduction. To explore novel inflammatory targets of PCSK9, we used systems biology in Ldlr-/- mouse macrophages. RESULTS: In Ldlr-/- mice, AAV-PCSK9 increased circulating PCSK9, but did not change serum cholesterol and triglyceride levels. AAV-PCSK9 promoted vein graft lesion development when compared with control AAV. In vivo molecular imaging revealed that AAV-PCSK9 increased macrophage accumulation and matrix metalloproteinase activity associated with decreased fibrillar collagen, a molecular determinant of atherosclerotic plaque stability. AAV-PCSK9 induced mRNA expression of the pro-inflammatory mediators IL-1ß (interleukin-1 beta), TNFα (tumor necrosis factor alpha), and MCP-1 (monocyte chemoattractant protein-1) in peritoneal macrophages underpinned by an in vitro analysis of Ldlr-/- mouse macrophages stimulated with endotoxin-free recombinant PCSK9. A combination of unbiased global transcriptomics and new network-based hyperedge entanglement prediction analysis identified the NF-κB (nuclear factor-kappa B) signaling molecules, lectin-like oxidized LOX-1 (LDL receptor-1), and SDC4 (syndecan-4) as potential PCSK9 targets mediating pro-inflammatory responses in macrophages. CONCLUSIONS: Circulating PCSK9 induces macrophage activation and vein graft lesion development via LDLR-independent mechanisms. PCSK9 may be a potential target for pharmacologic treatment for this unmet medical need.

Progression of Mitral Regurgitation in Rheumatic Valve Disease: Role of Left Atrial Remodeling.

Introduction: Mitral regurgitation (MR) is the most common valve abnormality in rheumatic heart disease (RHD) often associated with stenosis. Although the mechanism by which MR develops in RHD is primary, longstanding volume overload with left atrial (LA) remodeling may trigger the development of secondary MR, which can impact on the overall progression of MR. This study is aimed to assess the incidence and predictors of MR progression in patients with RHD. Methods: Consecutive RHD patients with non-severe MR associated with any degree of mitral stenosis were selected. The primary endpoint was a progression of MR, which was defined as an increase of one grade in MR severity from baseline to the last follow-up echocardiogram. The risk of MR progression was estimated accounting for competing risks. Results: The study included 539 patients, age of 46.2 ± 12 years and 83% were women. At a mean follow-up time of 4.2 years (interquartile range [IQR]: 1.2-6.9 years), 54 patients (10%) displayed MR progression with an overall incidence of 2.4 per 100 patient-years. Predictors of MR progression by the Cox model were age (adjusted hazard ratio [HR] 1.541, 95% CI 1.222-1.944), and LA volume (HR 1.137, 95% CI 1.054-1.226). By considering competing risk analysis, the direction of the association was similar for the rate (Cox model) and incidence (Fine-Gray model) of MR progression. In the model with LA volume, atrial fibrillation (AF) was no longer a predictor of MR progression. In the subgroup of patients in sinus rhythm, 59 had an onset of AF during follow-up, which was associated with progression of MR (HR 2.682; 95% CI 1.133-6.350). Conclusions: In RHD patients with a full spectrum of MR severity, progression of MR occurs over time is predicted by age and LA volume. LA enlargement may play a role in the link between primary MR and secondary MR in patients with RHD.

Prothymosin Alpha: A Novel Contributor to Estradiol Receptor Alpha-Mediated CD8+ T-Cell Pathogenic Responses and Recognition of Type 1 Collagen in Rheumatic Heart Valve Disease.

BACKGROUND: Rheumatic heart valve disease (RHVD) is a leading cause of cardiovascular death in low- and middle-income countries and affects predominantly women. The underlying mechanisms of chronic valvular damage remain unexplored and regulators of sex predisposition are unknown. METHODS: Proteomics analysis of human heart valves (nondiseased aortic valves, nondiseased mitral valves [NDMVs], valves from patients with rheumatic aortic valve disease, and valves from patients with rheumatic mitral valve disease; n=30) followed by system biology analysis identified ProTα (prothymosin alpha) as a protein associated with RHVD. Histology, multiparameter flow cytometry, and enzyme-linked immunosorbent assay confirmed the expression of ProTα. In vitro experiments using peripheral mononuclear cells and valvular interstitial cells were performed using multiparameter flow cytometry and quantitative polymerase chain reaction. In silico analysis of the RHVD and Streptococcuspyogenes proteomes were used to identify mimic epitopes. RESULTS: A comparison of NDMV and nondiseased aortic valve proteomes established the baseline differences between nondiseased aortic and mitral valves. Thirteen unique proteins were enriched in NDMVs. Comparison of NDMVs versus valves from patients with rheumatic mitral valve disease and nondiseased aortic valves versus valves from patients with rheumatic aortic valve disease identified 213 proteins enriched in rheumatic valves. The expression of the 13 NDMV-enriched proteins was evaluated across the 213 proteins enriched in diseased valves, resulting in the discovery of ProTα common to valves from patients with rheumatic mitral valve disease and valves from patients with rheumatic aortic valve disease. ProTα plasma levels were significantly higher in patients with RHVD than in healthy individuals. Immunoreactive ProTα colocalized with CD8+ T cells in RHVD. Expression of ProTα and estrogen receptor alpha correlated strongly in circulating CD8+ T cells from patients with RHVD. Recombinant ProTα induced expression of the lytic proteins perforin and granzyme B by CD8+ T cells as well as higher estrogen receptor alpha expression. In addition, recombinant ProTα increased human leukocyte antigen class I levels in valvular interstitial cells. Treatment of CD8+ T cells with specific estrogen receptor alpha antagonist reduced the cytotoxic potential promoted by ProTα. In silico analysis of RHVD and Spyogenes proteomes revealed molecular mimicry between human type 1 collagen epitope and bacterial collagen-like protein, which induced CD8+ T-cell activation in vitro. CONCLUSIONS: ProTα-dependent CD8+ T-cell cytotoxicity was associated with estrogen receptor alpha activity, implicating ProTα as a potential regulator of sex predisposition in RHVD. ProTα facilitated recognition of type 1 collagen mimic epitopes by CD8+ T cells, suggesting mechanisms provoking autoimmunity.

Systemic cytokines, chemokines and growth factors reveal specific and shared immunological characteristics in infectious cardiomyopathies.

Heart disease is a major cause of death worldwide. Chronic Chagas cardiomyopathy (CCC) caused by infection with Trypanosoma cruzi leading to high mortality in adults, and rheumatic heart disease (RHD), resulting from infection by Streptococcus pyogenes affecting mainly children and young adults, are amongst the deadliest heart diseases in low-middle income countries. Despite distinct etiology, the pathology associated with both diseases is a consequence of inflammation. Here we compare systemic immune profile in patients with these cardiopathies, to identify particular and common characteristics in these infectious heart diseases. We evaluated the expression of 27 soluble factors, employing single and multivariate analysis combined with machine-learning approaches. We observed that, while RHD and CCC display higher levels of circulating mediators than healthy individuals, CCC is associated with stronger immune activation as compared to RHD. Despite distinct etiologies, univariate analysis showed that expression of TNF, IL-17, IFN-gamma, IL-4, CCL4, CCL3, CXCL8, CCL11, CCL2, PDGF-BB were similar between CCC and RHD, consistent with their inflammatory nature. Network analysis revealed common inflammatory pathways between CCC and RHD, while highlighting the broader reach of the inflammatory response in CCC. The final multivariate model showed a 100% discrimination power for the combination of the cytokines IL-12p70, IL-1Ra, IL-4, and IL-7 between CCC and RHD groups. Thus, while clear immunological distinctions were identified between CCC and RHD, similarities indicate shared inflammatory pathways in these infectious heart diseases. These results contribute to understanding the pathogenesis of CCC and RHD and may impact the design of immune-based therapies for these and other inflammatory cardiopathies that may also share immunological characteristics.

Integration of Functional Imaging, Cytometry, and Unbiased Proteomics Reveals New Features of Endothelial-to-Mesenchymal Transition in Ischemic Mitral Valve Regurgitation in Human Patients.

Background: Following myocardial infarction, mitral regurgitation (MR) is a common complication. Previous animal studies demonstrated the association of endothelial-to-mesenchymal transition (EndMT) with mitral valve (MV) remodeling. Nevertheless, little is known about how MV tissue responds to ischemic heart changes in humans. Methods: MVs were obtained by the Cardiothoracic Surgical Trials Network from 17 patients with ischemic mitral regurgitation (IMR). Echo-doppler imaging assessed MV function at time of resection. Cryosections of MVs were analyzed using a multi-faceted histology and immunofluorescence examination of cell populations. MVs were further analyzed using unbiased label-free proteomics. Echo-Doppler imaging, histo-cytometry measures and proteomic analysis were then integrated. Results: MVs from patients with greater MR exhibited proteomic changes associated with proteolysis-, inflammatory- and oxidative stress-related processes compared to MVs with less MR. Cryosections of MVs from patients with IMR displayed activated valvular interstitial cells (aVICs) and double positive CD31+ αSMA+ cells, a hallmark of EndMT. Univariable and multivariable association with echocardiography measures revealed a positive correlation of MR severity with both cellular and geometric changes (e.g., aVICs, EndMT, leaflet thickness, leaflet tenting). Finally, proteomic changes associated with EndMT showed gene-ontology enrichment in vesicle-, inflammatory- and oxidative stress-related processes. This discovery approach indicated new candidate proteins associated with EndMT regulation in IMR. Conclusion: We describe an atypical cellular composition and distinctive proteome of human MVs from patients with IMR, which highlighted new candidate proteins implicated in EndMT-related processes, associated with maladaptive MV fibrotic remodeling.

Histopathological Characterization of Mitral Valvular Lesions from Patients with Rheumatic Heart Disease. / Caracterização Histológica das Lesões da Valva Mitral de Pacientes com Cardiopatia Reumática.

BACKGROUND: The underlying mechanisms by which rheumatic heart disease (RHD) lead to severe valve dysfunction are not completely understood. OBJECTIVE: The present study evaluated the histopathological changes in mitral valves (MV) seeking an association between the pattern of predominant valvular dysfunction and histopathological findings. METHODS: In 40 patients who underwent MV replacement due to RHD, and in 20 controls that underwent heart transplant, histological aspects of the excised MV were analyzed. Clinical and echocardiographic data were also collected. Histological analyses were performed using hematoxylin-eosin staining. Inflammation, fibrosis, neoangiogenesis, calcification and adipose metaplasia were determined. A p value<0.05 was considered to be statistically significant. RESULTS: The mean age of RHD patients was 53±13 years, 36 (90%) were female, whereas the mean age of controls was 50±12 years, similar to the cases, with the majority of males (70%). The rheumatic valve endocardium presented greater thickness than the controls (1.3±0.5 mm versus 0.90±0.4 mm, p=0.003, respectively), and a more intense inflammatory infiltrate in the endocardium (78% versus 36%; p=0.004), with predominance of mononuclear cells. Moderate to marked fibrosis occurred more frequently in rheumatic valves than in control valves (100% vs. 29%; p<0.001). Calcification occurred in 35% of rheumatic valves, especially among stenotic valves, which was associated with the mitral valve area (p=0.003). CONCLUSIONS: Despite intense degree of fibrosis, the inflammatory process remains active in the rheumatic mitral valve, even at late disease with valve dysfunction. Calcification predominated in stenotic valves and in patients with right ventricular dysfunction.

FUNDAMENTOS: Os mecanismos subjacentes pelos quais a doença cardíaca reumática (DCR) levam à disfunção valvar grave não são totalmente compreendidos. OBJETIVO: O presente estudo avaliou as alterações histopatológicas nas valvas mitrais (VM) buscando uma associação entre o padrão de disfunção valvar predominante e os achados histopatológicos. MÉTODOS: Em 40 pacientes submetidos à troca da VM devido a DCR e em 20 controles submetidos a transplante cardíaco, foram analisados os aspectos histológicos da VM excisada. Dados clínicos e ecocardiográficos também foram coletados. As análises histológicas foram realizadas usando coloração com hematoxilina-eosina. Determinou-se inflamação, fibrose, neoangiogênese, calcificação e metaplasia adiposa. Valores de p<0,05 foram considerados estatisticamente significativos. RESULTADOS: A idade média dos pacientes com DCR foi de 53±13 anos, sendo 36 (90%) do sexo feminino, enquanto a idade média dos controles foi de 50±12 anos, semelhante aos casos, sendo a maioria do sexo masculino (70%). O endocárdio valvar reumático apresentou espessura maior que os controles (1,3±0,5 mm versus 0,90±0,4 mm, p=0,003, respectivamente), e infiltrado inflamatório mais intenso no endocárdio (78% versus 36%; p=0,004), com predominância de células mononucleares. Ocorreu fibrose moderada a acentuada mais frequentemente em válvulas reumáticas do que em válvulas controle (100% vs. 29%; p<0,001). Ocorreu calcificação em 35% das valvas reumáticas, principalmente entre as valvas estenóticas, associada à área valvar mitral (p=0,003). CONCLUSÕES: Apesar do intenso grau de fibrose, o processo inflamatório permanece ativo na valva mitral reumática, mesmo em doença tardia com disfunção valvar. A calcificação predominou em valvas estenóticas e em pacientes com disfunção ventricular direita.

Caracterização Histológica das Lesões da Valva Mitral de Pacientes com Cardiopatia Reumática / Histopathological Characterization of Mitral Valvular Lesions from Patients with Rheumatic Heart Disease

Resumo Fundamentos: Os mecanismos subjacentes pelos quais a doença cardíaca reumática (DCR) levam à disfunção valvar grave não são totalmente compreendidos. Objetivo: O presente estudo avaliou as alterações histopatológicas nas valvas mitrais (VM) buscando uma associação entre o padrão de disfunção valvar predominante e os achados histopatológicos. Métodos: Em 40 pacientes submetidos à troca da VM devido a DCR e em 20 controles submetidos a transplante cardíaco, foram analisados os aspectos histológicos da VM excisada. Dados clínicos e ecocardiográficos também foram coletados. As análises histológicas foram realizadas usando coloração com hematoxilina-eosina. Determinou-se inflamação, fibrose, neoangiogênese, calcificação e metaplasia adiposa. Valores de p<0,05 foram considerados estatisticamente significativos. Resultados: A idade média dos pacientes com DCR foi de 53±13 anos, sendo 36 (90%) do sexo feminino, enquanto a idade média dos controles foi de 50±12 anos, semelhante aos casos, sendo a maioria do sexo masculino (70%). O endocárdio valvar reumático apresentou espessura maior que os controles (1,3±0,5 mm versus 0,90±0,4 mm, p=0,003, respectivamente), e infiltrado inflamatório mais intenso no endocárdio (78% versus 36%; p=0,004), com predominância de células mononucleares. Ocorreu fibrose moderada a acentuada mais frequentemente em válvulas reumáticas do que em válvulas controle (100% vs. 29%; p<0,001). Ocorreu calcificação em 35% das valvas reumáticas, principalmente entre as valvas estenóticas, associada à área valvar mitral (p=0,003). Conclusões: Apesar do intenso grau de fibrose, o processo inflamatório permanece ativo na valva mitral reumática, mesmo em doença tardia com disfunção valvar. A calcificação predominou em valvas estenóticas e em pacientes com disfunção ventricular direita.

Abstract Background: The underlying mechanisms by which rheumatic heart disease (RHD) lead to severe valve dysfunction are not completely understood. Objective: The present study evaluated the histopathological changes in mitral valves (MV) seeking an association between the pattern of predominant valvular dysfunction and histopathological findings. Methods: In 40 patients who underwent MV replacement due to RHD, and in 20 controls that underwent heart transplant, histological aspects of the excised MV were analyzed. Clinical and echocardiographic data were also collected. Histological analyses were performed using hematoxylin-eosin staining. Inflammation, fibrosis, neoangiogenesis, calcification and adipose metaplasia were determined. A p value<0.05 was considered to be statistically significant. Results: The mean age of RHD patients was 53±13 years, 36 (90%) were female, whereas the mean age of controls was 50±12 years, similar to the cases, with the majority of males (70%). The rheumatic valve endocardium presented greater thickness than the controls (1.3±0.5 mm versus 0.90±0.4 mm, p=0.003, respectively), and a more intense inflammatory infiltrate in the endocardium (78% versus 36%; p=0.004), with predominance of mononuclear cells. Moderate to marked fibrosis occurred more frequently in rheumatic valves than in control valves (100% vs. 29%; p<0.001). Calcification occurred in 35% of rheumatic valves, especially among stenotic valves, which was associated with the mitral valve area (p=0.003). Conclusions: Despite intense degree of fibrosis, the inflammatory process remains active in the rheumatic mitral valve, even at late disease with valve dysfunction. Calcification predominated in stenotic valves and in patients with right ventricular dysfunction.

Proinflammatory Matrix Metalloproteinase-1 Associates With Mitral Valve Leaflet Disruption Following Percutaneous Mitral Valvuloplasty.

Mitral regurgitation (MR) is a major complication of the percutaneous mitral valvuloplasty (PMV). Despite high technical expertise and cumulative experience with the procedure, the incidence rate of severe MR has not decreased. Although some of MR can be anticipated by echocardiographic analysis; leaflet tearing, which leads to the most dreaded type of MR, remains unpredictable. Irregular valvular collagen remodeling is likely to compromise tissue architecture and increase the tearing risk during PMV balloon inflation. In this study, we evaluated histological and molecular characteristics of excised mitral valves from patients with rheumatic mitral stenosis (MS) who underwent emergency surgery after PMV due to severe MR caused by leaflet tear. Those findings were compared with patients who underwent elective mitral valve replacement surgery owing to severe MS, in whom PMV was not indicated. In vitro assay using peripheral blood mononuclear cells was performed to better understand the impact of the cellular and molecular alterations identified in leaflet tear mitral valve specimens. Our analysis showed that focal infiltration of inflammatory cells contributes to accumulation of MMP-1 and IFN-γ in valve leaflets. Moreover, we showed that IFN-γ increase the expression of MMP-1 in CD14+ cells (monocytes) in vitro. Thus, inflammatory cells contribute to unevenly remodel collagen resulting in variable thickening causing abnormalities in leaflet architecture making them more susceptible to laceration.

Human CD8+ T Cells Release Extracellular Traps Co-Localized With Cytotoxic Vesicles That Are Associated With Lesion Progression and Severity in Human Leishmaniasis.

Cell death plays a fundamental role in mounting protective and pathogenic immunity. Etosis is a cell death mechanism defined by the release of extracellular traps (ETs), which can foster inflammation and exert microbicidal activity. While etosis is often associated with innate cells, recent studies showed that B cells and CD4+ T cells can release ETs. Here we investigate whether CD8+ T cells can also release ETs, which might be related to cytotoxicity and tissue pathology. To these ends, we first employed an in vitro system stimulating human CD8+ T cells isolated from healthy volunteers with anti-CD3/anti-CD28. Using time-frame video, confocal and electron microscopy, we demonstrate that human CD8+ T cells release ETs upon stimulation (herein LETs - lymphocyte extracellular traps), which display unique morphology and functional characteristics. CD8+ T cell-derived LETs form long strands that co-localize with CD107a, a marker of vesicles containing cytotoxic granules. In addition, these structures connect the LET-releasing cell to other neighboring cells, often resulting in cell death. After demonstrating the release of LETs by human CD8+ T cells in vitro, we went on to study the occurrence of CD8-derived LETs in a human disease setting. Thus, we evaluated the occurrence of CD8-derived LETs in lesions from patients with human tegumentary leishmaniasis, where CD8+ T cells play a key role in mediating pathology. In addition, we evaluated the association of these structures with the intensity of the inflammatory infiltrate in early and late cutaneous, as well as in mucosal leishmaniasis lesions. We demonstrated that progression and severity of debilitating and mutilating forms of human tegumentary leishmaniasis are associated with the frequency of CD8+ T cells in etosis, as well as the occurrence of CD8-derived LETs carrying CD107a+ vesicles in the lesions. We propose that CD8+ T cell derived LETs may serve as a tool for delivering cytotoxic vesicles to distant target cells, providing insights into mechanisms of CD8+ T cell mediated pathology.

Mitral Regurgitation After Percutaneous Mitral Valvuloplasty: Insights Into Mechanisms and Impact on Clinical Outcomes.

OBJECTIVES: The aim of this study was to assess the incidence, mechanisms, and outcomes of mitral regurgitation (MR) after percutaneous mitral valvuloplasty (PMV). BACKGROUND: Significant MR continues to be a major complication of PMV, with a wide range in clinical presentation and prognosis. METHODS: Consecutive patients with mitral stenosis undergoing PMV were prospectively enrolled. MR severity was evaluated by using quantitative echocardiographic criteria, and its mechanism was characterized by 3-dimensional transesophageal echocardiography, divided broadly into 4 categories based on the features contributing to the valve damage. B-type natriuretic peptide levels were obtained before and 24 h after the procedure. Endpoints estimated cardiovascular death or mitral valve (MV) replacement due to predominant MR. RESULTS: A total of 344 patients, ages 45.1 ± 12.1 years, of whom 293 (85%) were women, were enrolled. Significant MR after PMV was found in 64 patients (18.6%). The most frequent mechanism of MR was commissural, which occurred in 22 (34.4%) patients, followed by commissural with posterior leaflet in 16 (25.0%), leaflets at central scallop or subvalvular damage in 15 (23.4%), and central MR in 11 (17.2%). During the mean follow-up period of 3 years (range 1 day to 10.6 years), 60 patients reached the endpoint. The event-free survival rates were similar among patients with mild or commissural MR, whereas patients with damaged central leaflet scallop or subvalvular apparatus had the worst outcome, with an event-free survival rate at 1 year of only 7%. Long-term outcome was predicted by net atrioventricular compliance (Cn) at baseline and post-procedural variables, including valve area, mean gradient, and magnitude of decrease in B-type natriuretic peptide levels, adjusted for the mechanism of MR. CONCLUSIONS: Significant MR following PMV is a frequent event, mainly related to commissural splitting, with favorable clinical outcome. Parameters that express the relief of valve obstruction and the mechanism by which MR develops were predictors of long-term outcomes.

Innate and adaptive immunity in cardiovascular calcification.

Despite the focus placed on cardiovascular research, the prevalence of vascular and valvular calcification is increasing and remains a leading contributor of cardiovascular morbidity and mortality. Accumulating studies provide evidence that cardiovascular calcification is an inflammatory disease in which innate immune signaling becomes sustained and/or excessive, shaping a deleterious adaptive response. The triggering immune factors and subsequent inflammatory events surrounding cardiovascular calcification remain poorly understood, despite sustained significant research interest and support in the field. Most studies on cardiovascular calcification focus on innate cells, particularly macrophages' ability to release pro-osteogenic cytokines and calcification-prone extracellular vesicles and apoptotic bodies. Even though substantial evidence demonstrates that macrophages are key components in triggering cardiovascular calcification, the crosstalk between innate and adaptive immune cell components has not been adequately addressed. The only therapeutic options currently used are invasive procedures by surgery or transcatheter intervention. However, no approved drug has shown prophylactic or therapeutic effectiveness. Conventional diagnostic imaging is currently the best method for detecting, measuring, and assisting in the treatment of calcification. However, these common imaging modalities are unable to detect early subclinical stages of disease at the level of microcalcifications; therefore, the vast majority of patients are diagnosed when macrocalcifications are already established. In this review, we unravel the current knowledge of how innate and adaptive immunity regulate cardiovascular calcification; and put forward differences and similarities between vascular and valvular disease. Additionally, we highlight potential immunomodulatory drugs with the potential to target calcification and propose avenues in need of further translational inquiry.

Rheumatic Heart Valve Disease Pathophysiology and Underlying Mechanisms.

Rheumatic heart valve disease (RHVD) is a post-infectious sequel of acute rheumatic fever resulting from an abnormal immune response to a streptococcal pharyngitis that triggers valvular damage. RHVD is the leading cause of cardiovascular death in children and young adults, mainly in women from low and middle-income countries. It is known that long-term inflammation and high degree of fibrosis leads to valve dysfunction due to anatomic disruption of the valve apparatus. However, since public and private investments in RHVD studies are practically inexistent the number of publications is scarce. This disease shows different natural history and clinical presentations as compared to other degenerative heart valve diseases. Although more than five decades passed after the pioneering studies on the pathogenesis of RHVD, it is still unclear how self-tolerance mechanisms fail in this disease, and how humoral and cellular inflammatory responses are interconnected. Despite that pathological mechanisms have been already proposed for RHVD, none of them are able to explain the preferential involvement of the mitral valve. This review focuses on pathophysiology and underlying mechanisms of RHVD.

Raising awareness for rheumatic mitral valve disease.

Rheumatic heart disease (RHD) is a major burden in low- to mid-income countries, where each year it accounts for over a million premature deaths associated with severe valve disease. Life-saving valve replacement procedures are not available to the majority of affected RHD patients, contributing to an increased risk of death in young adults and creating a devastating impact. In December 2017, a group of representatives of major cardiothoracic societies and industry, discussed the plight of the millions of patients who suffer from RHD. A comprehensive solution based on this global partnership was outlined in "The Cape Town Declaration on Access to Cardiac Surgery in the Developing World". The key challenge in controlling RHD is related to identification and removal of barriers to the translation of existing knowledge into policy, programs, and practice to provide high-quality care for patients with RHD. This review provides an overview on RHD by emphasizing the disease medical and economic burdens worldwide, risk factors, recent advance for early disease detection, and overall preventive strategies.

Co-infection with distinct Trypanosoma cruzi strains induces an activated immune response in human monocytes.

AIMS: The aim of the study was to evaluate the immune response triggered by the first contact of human monocytes with two T cruzi strains from distinct discrete typing units (DTUs) IV and V, and whether co-infection with these strains leads to changes in monocyte immune profiles, which could in turn influence the subsequent infection outcome. METHODS AND RESULTS: We evaluated the influence of in vitro single- and co-infection with AM64 and 3253 strains on immunological characteristics of human monocytes. Single infection of monocytes with AM64 or 3253 induced opposing anti-inflammatory and inflammatory responses, respectively. Co-infection was observed in over 50% of monocytes after 15 hours of culture, but this percentage dropped ten-fold after 72 hours. Co-infection led to high monocyte activation and an increased percentage of both IL-10 and TNF. The decreased percentage of co-infected cells observed after 72 hours was associated with a decreased frequency of TNF-expressing cells. CONCLUSION: Our results show that the exacerbated response observed in co-infection with immune-polarizing strains is associated with a decreased frequency of co-infected cells, suggesting that the activated response favours parasite control. These findings may have implications for designing new Chagas disease preventive strategies.

CD4+ T cells apoptosis in Plasmodium vivax infection is mediated by activation of both intrinsic and extrinsic pathways.

BACKGROUND: Reduction in the number of circulating blood lymphocytes (lymphocytopaenia) has been reported during clinical episodes of malaria and is normalized after treatment with anti-malaria drugs. While this phenomenon is well established in malaria infection, the underlying mechanisms are still not fully elucidated. In the present study, the occurrence of apoptosis and its pathways in CD4+ T cells was investigated in naturally Plasmodium vivax-infected individuals from a Brazilian endemic area (Porto Velho - RO). METHODS: Blood samples were collected from P. vivax-infected individuals and healthy donors. The apoptosis was characterized by cell staining with Annexin V/FITC and propidium iodide and the apoptosis-associated gene expression profile was carried out using RT2 Profiler PCR Array-Human Apoptosis. The plasma TNF level was determined by ELISA. The unpaired t-test or Mann-Whitney test was applied according to the data distribution. RESULTS: Plasmodium vivax-infected individuals present low number of leukocytes and lymphocytes with a higher percentage of CD4+ T cells in early and/or late apoptosis. Increased gene expression was observed for TNFRSF1B and Bid, associated with a reduction of Bcl-2, in individuals with P. vivax malaria. Furthermore, these individuals showed increased plasma levels of TNF compared to malaria-naive donors. CONCLUSIONS: The results of the present study suggest that P. vivax infection induces apoptosis of CD4+ T cells mediated by two types of signaling: by activation of the TNFR1 death receptor (extrinsic pathway), which is amplified by Bid, and by decreased expression of the anti-apoptotic protein Bcl-2 (intrinsic pathway). The T lymphocytes apoptosis could reflect a strategy of immune evasion triggered by the parasite, enabling their persistence but also limiting the occurrence of immunopathology.