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BACKGROUND AND AIMS: The distinct functions of immune cells in atherosclerosis have been mostly defined by preclinical mouse studies. Contrastingly, the immune cell composition of human atherosclerotic plaques and their contribution to disease progression is only poorly understood. It remains uncertain whether genetic animal models allow for valuable translational approaches. METHODS AND RESULTS: Single cell RNA-sequencing (scRNA-seq) was performed to define the immune cell landscape in human carotid atherosclerotic plaques. The human immune cell repertoire demonstrated an unexpectedly high heterogeneity and was dominated by cells of the T-cell lineage, a finding confirmed by immunohistochemistry. Bioinformatical integration with 7 mouse scRNA-seq data sets from adventitial and atherosclerotic vascular tissue revealed a total of 51 identities of cell types and differentiation states, of which some were only poorly conserved between species and exclusively found in humans. Locations, frequencies, and transcriptional programs of immune cells in mouse models did not resemble the immune cell landscape in human carotid atherosclerosis. In contrast to standard mouse models of atherosclerosis, human plaque leukocytes were dominated by several T-cell phenotypes with transcriptional hallmarks of T-cell activation and memory formation, T-cell receptor-, and pro-inflammatory signaling. Only mice at the age of 22 months partially resembled the activated T-cell phenotype. In a validation cohort of 43 patients undergoing carotid endarterectomy, the abundance of activated immune cell subsets in the plaque defined by multi-color flow cytometry associated with the extend of clinical atherosclerosis. CONCLUSIONS: Integrative scRNA-seq reveals a substantial difference in the immune cell composition of murine and human carotid atherosclerosis - a finding that questions the translational value of standard mouse models for adaptive immune cell studies. Clinical associations suggest a specific role for T-cell driven (auto-) immunity in human plaque formation and -instability.
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Introduction: Myocardial infarction (MI) is a significant contributor to morbidity and mortality worldwide. Many individuals who survive the acute event continue to experience heart failure (HF), with inflammatory and healing processes post-MI playing a pivotal role. Polymorphonuclear neutrophils (PMN) and monocytes infiltrate the infarcted area, where PMN release high amounts of the heme enzyme myeloperoxidase (MPO). MPO has numerous inflammatory properties and MPO plasma levels are correlated with prognosis and severity of MI. While studies have focused on MPO inhibition and controlling PMN infiltration into the infarcted tissue, less is known on MPO's role in monocyte function. Methods and results: Here, we combined human data with mouse and cell studies to examine the role of MPO on monocyte activation and migration. We revealed a correlation between plasma MPO levels and monocyte activation in a patient study. Using a mouse model of MI, we demonstrated that MPO deficiency led to an increase in splenic monocytes and a decrease in cardiac monocytes compared to wildtype mice (WT). In vitro studies further showed that MPO induces monocyte migration, with upregulation of the chemokine receptor CCR2 and upregulation of inflammatory pathways identified as underlying mechanisms. Conclusion: Taken together, we identify MPO as a pro-inflammatory mediator of splenic monocyte recruitment and activation post-MI and provide mechanistic insight for novel therapeutic strategies after ischemic injury.
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Monócitos , Infarto do Miocárdio , Peroxidase , Animais , Infarto do Miocárdio/imunologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/metabolismo , Peroxidase/metabolismo , Monócitos/imunologia , Monócitos/metabolismo , Humanos , Camundongos , Masculino , Movimento Celular , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Feminino , Neutrófilos/imunologia , Neutrófilos/metabolismo , Camundongos Knockout , Receptores CCR2/metabolismo , Pessoa de Meia-IdadeRESUMO
Gravitational changes between micro- and hypergravity cause several adaptations and alterations in the human body. Besides muscular atrophy and immune system impairment, effects on the circulatory system have been described, which can be associated with a wide range of blood biomarker changes. This study examined nine individuals (seven males, two females) during a parabolic flight campaign (PFC). Thirty-one parabolas were performed in one flight day, resulting in ~22 s of microgravity during each parabola. Each participant was subjected to a single flight day with a total of 31 parabolas, totaling 11 min of microgravity during one parabolic flight. Before and after (1 hour (h) and 24 h), the flights blood was sampled to examine potential gravity-induced changes of circulating plasma proteins. Proximity Extension Assay (PEA) offers a proteomic solution, enabling the simultaneous analysis of a wide variety of plasma proteins. From 2925 unique proteins analyzed, 251 (8.58%) proteins demonstrated a differential regulation between baseline, 1 h and 24 h post flight. Pathway analysis indicated that parabolic flights led to altered levels of proteins associated with vesicle organization and apoptosis up to 24 h post microgravity exposure. Varying gravity conditions are associated with poorly understood physiological changes, including stress responses and fluid shifts. We provide a publicly available library of gravity-modulated circulating protein levels illustrating numerous changes in cellular pathways relevant for inter-organ function and communication.
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Macrophages are major players in myocardial infarction (MI) and atherosclerosis, two major cardiovascular diseases (CVD). Atherosclerosis is caused by the buildup of cholesterol-rich lipoproteins in blood vessels, causing inflammation, vascular injury, and plaque formation. Plaque rupture or erosion can cause thrombus formation resulting in inadequate blood flow to the heart muscle and MI. Inflammation, particularly driven by macrophages, plays a central role in both atherosclerosis and MI. Recent integrative approaches of single-cell analysis-based classifications in both murine and human atherosclerosis as well as experimental MI showed overlap in origin, diversity, and function of macrophages in the aorta and the heart. We here discuss differences and communalities between macrophages in the heart and aorta at steady state and in atherosclerosis or upon MI. We focus on markers, mediators, and functional states of macrophage subpopulations. Recent trials testing anti-inflammatory agents show a major benefit in reducing the inflammatory burden of CVD patients, but highlight a necessity for a broader understanding of immune cell ontogeny and heterogeneity in CVD. The novel insights into macrophage biology in CVD represent exciting opportunities for the development of novel treatment strategies against CVD.
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Aterosclerose , Doenças Cardiovasculares , Infarto do Miocárdio , Placa Aterosclerótica , Humanos , Animais , Camundongos , Doenças Cardiovasculares/etiologia , Macrófagos , Aterosclerose/etiologia , Placa Aterosclerótica/etiologia , Infarto do Miocárdio/complicações , InflamaçãoRESUMO
Background: An abdominal aortic aneurysm (AAA) is a life-threatening cardiovascular disease. Although its pathogenesis is still poorly understood, recent evidence suggests that AAA displays autoimmune disease characteristics. Particularly, T cells responding to AAA-related antigens in the aortic wall may contribute to an initial immune response. Single-cell RNA (scRNA) T cell receptor (TCR) and B cell receptor (BCR) sequencing is a powerful tool for investigating clonality. However, difficulties such as limited numbers of isolated cells must be considered during implementation and data analysis, making biological interpretation challenging. Here, we perform a representative single-cell immune repertoire analysis in experimental murine AAA and show a reliable bioinformatic processing pipeline highlighting opportunities and limitations of this approach. Methods: We performed scRNA TCR and BCR sequencing of isolated lymphocytes from the infrarenal aorta of male C57BL/6J mice 3, 7, 14, and 28 days after AAA induction via elastase perfusion of the aorta. Sham-operated mice at days 3 and 28 and non-operated mice served as controls. Results: Comparison of complementarity-determining region (CDR3) length distribution of 179 B cells and 796 T cells revealed neither differences between AAA and control nor between the disease stages. We found no clonal expansion of B cells in AAA. For T cells, we identified several clones in 11 of 16 AAA samples and one of eight control samples. Immune receptor repertoire comparison indicated that only a few clones were shared between the individual AAA samples. The most frequently used V-genes in the TCR beta chain in AAA were TRBV3, TRBV19, and the splicing variant TRBV12-2 + TRBV13-2. Conclusion: We found no clonal expansion of B cells but evidence for clonal expansion of T cells in elastase-induced AAA in mice. Our findings imply that a more precise characterization of TCR and BCR distribution requires a more extensive number of lymphocytes to prevent undersampling and potentially detect rare clones. Thus, further experiments are necessary to confirm our findings. In summary, this paper examines TCR and BCR sequencing results, identifies limitations and pitfalls, and offers guidance for future studies.
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Vascular diseases like atherosclerosis and abdominal aortic aneurysm (AAA) are common pathologies in the western world, promoting various potentially fatal conditions. Here, we evaluate high-resolution (HR) ultrasound in mouse models of atherosclerosis and AAA as a useful tool for noninvasive monitoring of early vascular changes in vivo. We used Apolipoprotein E-deficient (ApoE-/-) mice as an atherosclerosis model and induced AAA development by the implementation of Angiotensin II-releasing osmotic minipumps. HR ultrasound of the carotid artery or the abdominal aorta was performed to monitor vascular remodeling in vivo. Images were analyzed by speckle tracking algorithms and correlated to histological analyses and subsequent automated collagen quantification. Consistent changes were observed via ultrasound in both models: Global radial strain (GRS) was notably reduced in the AAA model (23.8 ± 2.8% vs. 12.5 ± 2.5%, p = 0.01) and in the atherosclerotic mice (20.6 ± 1.3% vs. 15.8 ± 0.9%, p = 0.02). In mice with AAA, vessel distensibility was significantly reduced, whereas intima-media thickness was increased in atherosclerotic mice. The area and collagen content of the tunica media were increased in diseased arteries of both models as measured by automated image analysis of Picrosirius Red-stained aortic sections. Correlation analysis revealed a strong correlation of multiple parameters, predicting early vascular damage in HR ultrasound and histological examinations. In conclusion, our findings underscore the potential of HR ultrasound in effectively tracing early alterations in arterial wall properties in murine models of atherosclerosis and AAA.
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Aneurisma da Aorta Abdominal , Aterosclerose , Camundongos , Animais , Modelos Animais de Doenças , Espessura Intima-Media Carotídea , Aterosclerose/diagnóstico por imagem , Aterosclerose/patologia , Aneurisma da Aorta Abdominal/diagnóstico por imagem , Aneurisma da Aorta Abdominal/patologia , Aorta Abdominal/diagnóstico por imagem , Angiotensina II , Colágeno , Camundongos Endogâmicos C57BLRESUMO
Cardiotoxicity is a major complication of anthracycline therapy that negatively impacts prognosis. Effective pharmacotherapies for prevention of anthracycline-induced cardiomyopathy (AICM) are currently lacking. Increased plasma levels of the neutrophil-derived enzyme myeloperoxidase (MPO) predict occurrence of AICM in humans. We hypothesized that MPO release causally contributes to AICM. Mice intravenously injected with the anthracycline doxorubicin (DOX) exhibited higher neutrophil counts and MPO levels in the circulation and cardiac tissue compared to saline (NaCl)-treated controls. Neutrophil-like HL-60 cells exhibited increased MPO release upon exposition to DOX. DOX induced extensive nitrosative stress in cardiac tissue alongside with increased carbonylation of sarcomeric proteins in wildtype but not in Mpo-/- mice. Accordingly, co-treatment of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with DOX and MPO aggravated loss of hiPSC-CM-contractility compared to DOX treatment alone. DOX-treated animals exhibited pronounced cardiac apoptosis and inflammation, which was attenuated in MPO-deficient animals. Finally, genetic MPO deficiency and pharmacological MPO inhibition protected mice from the development of AICM. The anticancer efficacy of DOX was unaffected by MPO deficiency. Herein we identify MPO as a critical mediator of AICM. We demonstrate that DOX induces cardiac neutrophil infiltration and release of MPO, which directly impairs cardiac contractility through promoting oxidation of sarcomeric proteins, cardiac inflammation and cardiomyocyte apoptosis. MPO thus emerges as a promising pharmacological target for prevention of AICM.
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Cardiomiopatias , Células-Tronco Pluripotentes Induzidas , Peroxidase , Animais , Humanos , Camundongos , Antraciclinas/toxicidade , Cardiomiopatias/induzido quimicamente , Cardiomiopatias/prevenção & controle , Doxorrubicina/toxicidade , Inflamação , Peroxidase/genéticaRESUMO
In atherosclerosis, some regulatory T (Treg) cells become exTreg cells. We crossed inducible Treg and exTreg cell lineage-tracker mice (FoxP3eGFP-Cre-ERT2ROSA26CAG-fl-stop-fl-tdTomato) to atherosclerosis-prone Apoe-/- mice, sorted Treg cells and exTreg cells and determined their transcriptomes by bulk RNA sequencing (RNA-seq). Genes that were differentially expressed between mouse Treg cells and exTreg cells and filtered for their presence in a human single-cell RNA-sequencing (scRNA-seq) panel identified exTreg cell signature genes as CST7, NKG7, GZMA, PRF1, TBX21 and CCL4. Projecting these genes onto the human scRNA-seq with CITE-seq data identified human exTreg cells as CD3+CD4+CD16+CD56+, which was validated by flow cytometry. Bulk RNA-seq of sorted human exTreg cells identified them as inflammatory and cytotoxic CD4+T cells that were significantly distinct from both natural killer and Treg cells. DNA sequencing for T cell receptor-ß showed clonal expansion of Treg cell CDR3 sequences in exTreg cells. Cytotoxicity was functionally demonstrated in cell killing and CD107a degranulation assays, which identifies human exTreg cells as cytotoxic CD4+T cells.
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Aterosclerose , Linfócitos T Reguladores , Humanos , Animais , CamundongosRESUMO
BACKGROUND: "Spaceflight associated neuro-ocular syndrome" (SANS) represents a challenging health condition in modern space medicine. Forty-eight percent of astronauts are diagnosed with SANS after long-term space missions. The pathophysiological mechanism seems to be multifactorial, and yet remains unknown. In this proof-of-concept study we plan to investigate retinal microcirculatory changes in weightlessness and aim to identify their role in the development of SANS. METHODS AND DESIGN: Healthy individuals will take part in a parabolic flight campaign, which recreates fractioned total weightlessness periods. The airplane is specifically equipped, and designed for the execution of parabolic flight maneuvers and scientific research in microgravity. Retinal microcirculation will be assessed with a modified fundus camera, which allows dynamic vessel analysis. We will additionally measure intra-ocular pressure and hemodynamic changes during each phase of the flight. Blood samples will be analyzed at baseline, one hour and 24 hours after exposure to weightlessness. CONCLUSIONS: This pilot study aims to investigate the feasibility of retinal microcirculation assessment during varying gravity. Results of this study may generate insights whether venous stasis in the eye, surrogated by the dilatation of retinal vessels and increase in intraocular pressure as signs of venous insufficiency, may potentially contribute to the development of SANS.
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Voo Espacial , Ausência de Peso , Humanos , Pressão Intracraniana/fisiologia , Microcirculação , Projetos Piloto , Ausência de Peso/efeitos adversosRESUMO
The advent of single-cell biology opens a new chapter for understanding human biological processes and for diagnosing, monitoring, and treating disease. This revolution now reaches the field of cardiovascular disease (CVD). New technologies to interrogate CVD samples at single-cell resolution are allowing the identification of novel cell communities that are important in shaping disease development and direct towards new therapeutic strategies. These approaches have begun to revolutionize atherosclerosis pathology and redraw our understanding of disease development. This review discusses the state-of-the-art of single-cell analysis of atherosclerotic plaques, with a particular focus on human lesions, and presents the current resolution of cellular subpopulations and their heterogeneity and plasticity in relation to clinically relevant features. Opportunities and pitfalls of current technologies as well as the clinical impact of single-cell technologies in CVD patient care are highlighted, advocating for multidisciplinary and international collaborative efforts to join the cellular dots of CVD.
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Aterosclerose , Doenças Cardiovasculares , Placa Aterosclerótica , Humanos , Aterosclerose/patologia , Placa Aterosclerótica/patologiaRESUMO
Atherosclerosis is initiated by subendothelial retention of lipoproteins and cholesterol, which triggers a non-resolving inflammatory process that over time leads to plaque progression in the artery wall. Myeloid cells and in particular macrophages are the primary drivers of the inflammatory response and plaque formation. Several immune cells including macrophages, T cells and B cells secrete the anti-inflammatory cytokine IL-10, known to be essential for the atherosclerosis protection. The cellular source of IL-10 in natural atherosclerosis progression is unknown. This study aimed to determine the main IL10-producing cell type in atherosclerosis. To do so, we crossed VertX mice, in which IRES-green fluorescent protein (eGFP) was placed downstream of exon 5 of the Il10 gene, with atherosclerosis-prone Apoe-/- mice. We found that myeloid cells express high levels of IL-10 in VertX Apoe-/- mice in both chow and western-diet fed mice. By single cell RNA sequencing and flow cytometry analysis, we identified resident and inflammatory macrophages in atherosclerotic plaques as the main IL-10 producers. To address whether IL-10 secreted by myeloid cells is essential for the protection, we utilized LyzMCre+Il10fl/fl mice crossed into the Apoe-/- background and confirmed that macrophages were unable to secrete IL-10. Chow and western diet-fed LyzMCre+Il10fl/fl Apoe-/- mice developed significantly larger atherosclerotic plaques as measured by en face morphometry than LyzMCre-Il10 fl/flApoe-/-. Flow cytometry and cytokine measurements suggest that the depletion of IL-10 in myeloid cells increases Th17 cells with elevated CCL2, and TNFα in blood plasma. We conclude that macrophage-derived IL-10 is critical for limiting atherosclerosis in mice.
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Aterosclerose , Interleucina-10 , Placa Aterosclerótica , Animais , Camundongos , Aterosclerose/genética , Aterosclerose/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Interleucina-10/genética , Interleucina-10/metabolismo , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Placa Aterosclerótica/metabolismo , Camundongos Knockout para ApoERESUMO
BACKGROUND: The SARS-CoV-2 variant B.1.1.529 potentially escapes immunity from vaccination via a heavily mutated Spike protein. Here, we analyzed whether T cell memory towards the B.1.1.529 Spike protein is present in individuals who received two or three doses of vaccines designed against the original Wuhan strain of SARS-CoV-2. METHODS: PBMCs were isolated from two- and three-times vaccinated study participants and incubated in vitro with peptide pools of the Spike protein derived from sequences of the original Wuhan or the B.1.1.529 strains of SARS-CoV-2. Activated antigen-specific T cells were detected by flow cytometry. In silico analyses with NetMHCpan and NetMHCIIpan were used to determine differences in MHC class presentation between the original strain and the B.1.1.529 strain for the most common MHCs in the European-Caucasian population. RESULTS: Here we show, that both CD4 and CD8 responses to the B.1.1.529 Spike protein are marginally reduced compared to the ancestor protein and a robust T cell response is maintained. Epitope analyses reveal minor differences between the two SARS-CoV-2 strains in terms of MHC class presentations for the MHC-alleles being most common in the European-Caucasian population. CONCLUSIONS: The memory T cell response induced via first generation vaccination remains robust and is mostly unaffected by B.1.1.529 mutations. Correspondingly, in silico analyses of MHC presentation of epitopes derived from the B.1.1.529 Spike protein shows marginal differences compared to the ancestral SARS-CoV-2 strain.
Vaccination against SARS-CoV-2 results in the production of proteins called antibodies, that bind and inactivate the virus, and cells that help to eliminate it from the body in a future encounter, such as memory T cells. Both antibodies and memory T cells remain in the body after vaccination with memory T cells being present for longer than antibodies. Here, we determined that even though most of the first generation vaccines were created to prevent infection with the original SARS-CoV-2 virus, the memory T cells generated by this vaccination can also detect the omicron variant.
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Chronic kidney disease's prevalence rises globally. Whereas dialysis treatment replaces the kidney's filtering function and prolongs life, dreaded consequences in remote organs develop inevitably over time. Even milder reductions in kidney function not requiring replacement therapy associate with bacterial infections, cardiovascular and heart valve disease, which markedly limit prognosis in these patients. The array of complications is diverse and engages a wide gamut of cellular and molecular mechanisms. The innate immune system is profoundly and systemically altered in chronic kidney disease and, as a unifying element, partakes in many of the disease's complications. As such, a derailed immune system fuels cardiovascular disease progression but also elevates the propensity for serious bacterial infections. Recent data further point towards a role in developing calcific aortic valve stenosis. Here, we delineate the current state of knowledge on how chronic kidney disease affects innate immunity in cardiovascular organs and on a systemic level. We review the role of circulating myeloid cells, monocytes and neutrophils, resident macrophages, dendritic cells, ligands, and cellular pathways that are activated or suppressed when renal function is chronically impaired. Finally, we discuss myeloid cells' varying responses to uremia from a systems immunology perspective.
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Insuficiência Renal Crônica , Uremia , Humanos , Inflamação , Leucócitos , Macrófagos , Insuficiência Renal Crônica/complicações , Uremia/complicaçõesRESUMO
Atherosclerosis is an inflammatory disease of the artery walls and involves immune cells such as macrophages. Olfactory receptors (OLFRs) are G proteincoupled chemoreceptors that have a central role in detecting odorants and the sense of smell. We found that mouse vascular macrophages express the olfactory receptor Olfr2 and all associated trafficking and signaling molecules. Olfr2 detects the compound octanal, which activates the NLR family pyrin domain containing 3 (NLRP3) inflammasome and induces interleukin-1ß secretion in human and mouse macrophages. We found that human and mouse blood plasma contains octanal, a product of lipid peroxidation, at concentrations sufficient to activate Olfr2 and the human ortholog olfactory receptor 6A2 (OR6A2). Boosting octanal levels exacerbated atherosclerosis, whereas genetic targeting of Olfr2 in mice significantly reduced atherosclerotic plaques. Our findings suggest that inhibiting OR6A2 may provide a promising strategy to prevent and treat atherosclerosis.
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Aldeídos/metabolismo , Aterosclerose/metabolismo , Interleucina-1/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Receptores Odorantes/metabolismo , Adulto , Aldeídos/análise , Aldeídos/sangue , Aldeídos/farmacologia , Animais , Aorta , Aterosclerose/tratamento farmacológico , Humanos , Inflamassomos/metabolismo , Interleucina-1alfa/metabolismo , Peroxidação de Lipídeos , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Estresse Oxidativo , Receptores Odorantes/antagonistas & inibidores , Receptores Odorantes/genética , Transdução de SinaisRESUMO
AIMS: Marfan syndrome (MFS) is a connective tissue disorder caused by mutations in the Fibrillin-1 gene. It is associated with formation of thoracic aortic aneurysms that can potentially be a life-threatening condition due to aortic rupture or dissection. Excessive non-canonical transforming growth factor beta signalling, mediated by activation of extracellular signal-regulated kinases 1/2 (ERK1/2), as well as inducible nitric oxide synthase (NOS2)-dependent nitric oxide production, have been identified to drive aortic pathology in MFS through induction of elastin fragmentation and smooth muscle cell apoptosis. Despite promising results in animal studies, specific pharmacological interventions approved for clinical use in patients with MFS-related aortic disease are rare. Nitro-oleic acid (NO2-OA) is an endogenously generated signalling modulator, which is available as an oral compound and has been shown to inhibit ERK1/2 activation and NOS2 expression in different disease models, thereby exerting promising therapeutic effects. In this study, we investigated whether NO2-OA decreases aortic dilation in MFS. METHODS AND RESULTS: Eight-week-old MFS (Fbn1C1041G/+) mice were treated with NO2-OA or vehicle for 4 weeks via subcutaneously implanted osmotic minipumps. Echocardiography indicated progressive ascending aortic dilation and wall stiffening in MFS mice, which was significantly attenuated by NO2-OA treatment. This protective effect was mediated by inhibition of aortic ERK1/2, Smad2 as well as nuclear factor kappa B overactivation and consequent attenuation of elastin fragmentation by matrix metalloproteinase 2, apoptosis, and collagen deposition. Critically, the therapeutic efficacy of NO2-OA in MFS was further emphasized by demonstrating its capability to reduce lethal aortic complications in Fbn1C1041G/+ mice challenged with Angiotensin II. CONCLUSION: NO2-OA distinctly attenuates progression of aortic dilation in MFS via modulation of well-established disease-mediating pathways, thereby meriting further investigation into its application as a therapeutic agent for the treatment of this condition.
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Aneurisma da Aorta Torácica , Aneurisma Aórtico , Doenças da Aorta , Síndrome de Marfan , Animais , Aneurisma Aórtico/genética , Aneurisma da Aorta Torácica/etiologia , Aneurisma da Aorta Torácica/genética , Doenças da Aorta/patologia , Modelos Animais de Doenças , Elastina/metabolismo , Fibrilina-1/genética , Síndrome de Marfan/complicações , Síndrome de Marfan/tratamento farmacológico , Síndrome de Marfan/genética , Metaloproteinase 2 da Matriz , Camundongos , Nitrocompostos , Ácidos OleicosRESUMO
Atherosclerotic cardiovascular diseases are the major cause of death worldwide. CD4 T cells responding to Apolipoprotein B (ApoB), the core protein of most lipoproteins, have been identified as critical disease modulators. In healthy individuals, ApoB-reactive (ApoB+) CD4 T cells are mostly regulatory T cells (Tregs), which exert anti-inflammatory effects. Yet, they may obtain pro-inflammatory features and thus become proatherogenic. Evidence from animal studies suggests that vaccination against certain major histocompatibility complex (MHC) II-binding ApoB peptides induces an expansion of ApoB+ Tregs and thus confers atheroprotection. To date, in-depth phenotyping of vaccine-expanded ApoB+ T cells has not yet been performed. To this end, we vaccinated C57BL/6J mice with the ApoB-peptide P6 (ApoB978-993 TGAYSNASSTESASY) and performed single-cell RNA sequencing of tetramer-sorted P6+ T cells. P6+ cells were clonally expanded (one major, two minor clones) and formed a transcriptional cluster distinct from clusters mainly containing non-expanded P6+ and P6- cells. Transcriptomic profiling revealed that most expanded P6+ cells had a strong Treg signature and highly expressed genes mediating suppressive functions. Yet, some expanded P6+ cells only had a residual Treg signature and expressed genes related to T helper 1 (TH1) cells, which are proatherogenic. Modeling the T cell receptor (TCR) and P6:MHC-II interaction showed that only three amino acid residues in the α and ß chain contact the P6 peptide in the MHC-II groove and thus determine the specificity of this TCR to P6. Our data begin to reveal the vaccination-induced response to an ApoB epitope.
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Myocardial infarction (MI) is a leading cause of morbidity and mortality worldwide. Improved survival has led to an increasing incidence of ischemic cardiomyopathy, making it a major reason for hospitalization in the western world. The inflammatory response in the ischemic myocardium determines the extent of structural remodeling and functional deterioration, with neutrophils (PMN) being a key modulator of the propagation and resolution of inflammation. The heme enzyme myeloperoxidase (MPO) is abundantly expressed in PMN and is an important mediator of their inflammatory capacities. Here, we examine the effects of PMN reduction, MPO deficiency and MPO inhibition in two murine models of MI. Reduction in PMN count resulted in less scar formation and improved cardiac function. Similar results were obtained in genetically MPO deficient mice, suggesting that MPO is a critical factor in PMN-mediated cardiac remodeling. To test our findings in a therapeutic approach, we orally administered the MPO inhibitor AZM198 in the context of MI and could demonstrate improved cardiac function and reduced structural remodeling. Therefore, MPO appears to be a favorable pharmacological target for the prevention of long-term morbidity after MI.
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Double-positive CD4+CD8αß+ (DP) cells are thought to reside as T cell progenitors exclusively within the thymus. We recently discovered an unexpected CD4+ and CD8αß+ immune cell population in healthy and atherosclerotic mice by single-cell RNA sequencing. Transcriptomically, these cells resembled thymic DPs. Flow cytometry and three-dimensional whole-mount imaging confirmed DPs in thymus, mediastinal adipose tissue, and aortic adventitia, but nowhere else. Deep transcriptional profiling revealed differences between DP cells isolated from the three locations. All DPs were dependent on RAG2 expression and the presence of the thymus. Mediastinal adipose tissue DPs resided in close vicinity to invariant NKT cells, which they could activate in vitro. Thymus transplantation failed to reconstitute extrathymic DPs, and frequencies of extrathymic DPs were unaltered by pharmacologic inhibition of S1P1, suggesting that their migration may be locally confined. Our results define two new, transcriptionally distinct subsets of extrathymic DPs that may play a role in aortic vascular homeostasis.