RESUMEN
Gene-editing technologies, which include the CRISPR-Cas nucleases1-3 and CRISPR base editors4,5, have the potential to permanently modify disease-causing genes in patients6. The demonstration of durable editing in target organs of nonhuman primates is a key step before in vivo administration of gene editors to patients in clinical trials. Here we demonstrate that CRISPR base editors that are delivered in vivo using lipid nanoparticles can efficiently and precisely modify disease-related genes in living cynomolgus monkeys (Macaca fascicularis). We observed a near-complete knockdown of PCSK9 in the liver after a single infusion of lipid nanoparticles, with concomitant reductions in blood levels of PCSK9 and low-density lipoprotein cholesterol of approximately 90% and about 60%, respectively; all of these changes remained stable for at least 8 months after a single-dose treatment. In addition to supporting a 'once-and-done' approach to the reduction of low-density lipoprotein cholesterol and the treatment of atherosclerotic cardiovascular disease (the leading cause of death worldwide7), our results provide a proof-of-concept for how CRISPR base editors can be productively applied to make precise single-nucleotide changes in therapeutic target genes in the liver, and potentially in other organs.
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Sistemas CRISPR-Cas , LDL-Colesterol/sangre , Edición Génica , Modelos Animales , Proproteína Convertasa 9/genética , Adenina/metabolismo , Animales , Células Cultivadas , Femenino , Hepatocitos/metabolismo , Humanos , Hígado/enzimología , Mutación con Pérdida de Función , Macaca fascicularis/sangre , Macaca fascicularis/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Mutagénesis Sitio-Dirigida , Proproteína Convertasa 9/sangre , Proproteína Convertasa 9/metabolismo , Factores de TiempoRESUMEN
In the brain, neurons, glial cells, vascular endothelial cells (ECs), and mural cells form a functional structure referred to as the neurovascular unit (NVU). The functions of the NVU become impaired with aging. To gain insight into the mechanism underlying the aging-related changes in the NVU, we characterized in the present study the gliovascular interface in transgenic mice expressing a dominant-negative form of the telomeric repeat-binding factor 2 (TERF2) specifically in ECs using the Tie2 promoter. In these transgenic mice, senescence occurred in the cerebral ECs and was accompanied by upregulation of the mRNAs of proinflammatory cell adhesion molecules and cytokines. It is noteworthy that in the deep layers of the cerebral cortex, astrocytes exhibited an increase in the signals for S100ß as well as a decrease in the polarization of the water channel aquaporin-4 (AQP4) to the perivascular endfeet of the astrocytes. Mechanistically, the perivascular localization of dystroglycan and its ligand, laminin α2, was decreased, and their localization correlated well with the perivascular localization of AQP4, which supports the notion that their interaction regulates the perivascular localization of AQP4. The diminished perivascular localization of laminin α2 may be attributed to its proteolytic degradation by the matrix metalloproteinase-2 released by senescent ECs. Pericyte coverage was increased and negatively correlated with the decrease in the perivascular localization of AQP4. We propose that aging-related changes in ECs induce a mild morphological alteration of astrocytes and affect the localization of AQP4 at the gliovascular interface.
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Senescencia Celular , Células Endoteliales , Laminina , Metaloproteinasa 2 de la Matriz , Neuroglía , Animales , Ratones , Acuaporina 4/genética , Acuaporina 4/metabolismo , Astrocitos/metabolismo , Células Endoteliales/metabolismo , Laminina/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , Ratones Transgénicos , Neuroglía/metabolismoRESUMEN
[Figure: see text].
Asunto(s)
Enfermedad de la Arteria Coronaria/metabolismo , Proteína ADAMTS7/química , Proteína ADAMTS7/genética , Proteína ADAMTS7/metabolismo , Animales , Dominio Catalítico , Células Cultivadas , Enfermedad de la Arteria Coronaria/genética , Células HEK293 , Humanos , Ratones , Ratones Endogámicos C57BL , Músculo Liso Vascular/metabolismo , Mutación , Polimorfismo de Nucleótido SimpleRESUMEN
BACKGROUND: It is increasingly recognized that gut microbiota play a pivotal role in the development of atherosclerotic cardiovascular disease. Previously, we have reported that the abundance of genus Bacteroides is lower in patients with coronary artery disease (CAD) than in patients without CAD with coronary risk factors or in healthy volunteers. However, it remains unclear which and how specific gut bacteria contribute to the progression of atherosclerosis. METHODS: We recruited patients with CAD patients and controls without CAD with coronary risk factors. We then compared gut microbial composition using 16S ribosomal RNA gene sequencing in fecal samples to detect species with differential abundance between 2 groups. Subsequently, we used atherosclerosis-prone mice to study the mechanisms underlying the relationship between such species and atherosclerosis. RESULTS: Human fecal 16S ribosomal RNA gene sequencing revealed a significantly lower abundance of Bacteroides vulgatus and Bacteroides dorei in patients with CAD. This significant differential abundance was confirmed by quantitative polymerase chain reaction. Gavage with live B. vulgatus and B. dorei attenuated atherosclerotic lesion formation in atherosclerosis-prone mice, markedly ameliorating endotoxemia followed by decreasing gut microbial lipopolysaccharide production, effectively suppressing proinflammatory immune responses. Furthermore, fecal lipopolysaccharide levels in patients with CAD were significantly higher and negatively correlated with the abundance of B. vulgatus and B. dorei. CONCLUSIONS: Our translational research findings identify a previously unknown link between specific gut bacteria and atherosclerosis. Treatment with live B. vulgatus and B. dorei may help prevent CAD. CLINICAL TRIAL REGISTRATION: URL: https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000018051 . Unique identifier: UMIN000015703.
Asunto(s)
Aterosclerosis/patología , Bacteroides/aislamiento & purificación , Microbioma Gastrointestinal , Lipopolisacáridos/sangre , Anciano , Animales , Aterosclerosis/epidemiología , Aterosclerosis/inmunología , Aterosclerosis/veterinaria , Bacteroides/genética , Heces/microbiología , Femenino , Humanos , Inmunidad Mucosa , Intestinos/inmunología , Lipopolisacáridos/análisis , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , ARN Ribosómico 16S/genética , ARN Ribosómico 16S/metabolismo , Factores de Riesgo , Análisis de Secuencia de ARN , Uniones Estrechas/metabolismo , Uniones Estrechas/microbiologíaRESUMEN
OBJECTIVE: UVB irradiation is an established treatment for immunoinflammatory cutaneous disorders and has been shown to suppress cutaneous and systemic inflammatory diseases through modulation of the adaptive immune response. However, it remains unknown whether UVB irradiation prevents an immunoinflammatory disease of arteries such as atherosclerosis. APPROACH AND RESULTS: Here, we show that UVB exposure inhibits the development and progression of atherosclerosis in atherosclerosis-prone mice by expanding and enhancing the functional capacity of CD4+ forkhead box P3+ regulatory T cells and regulating proatherogenic T-cell responses. Experimental studies in Langerhans cell-depleted mice revealed that epidermal Langerhans cells play a critical role in UVB-dependent induction of CD4+ forkhead box P3+ regulatory T cells, suppression of proatherogenic T-cell responses, and prevention of atherosclerotic plaque development. CONCLUSIONS: Our findings suggest the skin immune system as a novel therapeutic target for atherosclerosis and provide a novel strategy for the treatment and prevention of atherosclerosis.
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Aorta/efectos de la radiación , Enfermedades de la Aorta/prevención & control , Aterosclerosis/prevención & control , Inflamación/prevención & control , Piel/efectos de la radiación , Linfocitos T Reguladores/efectos de la radiación , Rayos Ultravioleta , Animales , Aorta/inmunología , Aorta/metabolismo , Aorta/patología , Enfermedades de la Aorta/inmunología , Enfermedades de la Aorta/metabolismo , Enfermedades de la Aorta/patología , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Aterosclerosis/inmunología , Aterosclerosis/metabolismo , Aterosclerosis/patología , Células Cultivadas , Modelos Animales de Enfermedad , Factores de Transcripción Forkhead/inmunología , Factores de Transcripción Forkhead/metabolismo , Predisposición Genética a la Enfermedad , Inflamación/inmunología , Inflamación/metabolismo , Inflamación/patología , Células de Langerhans/inmunología , Células de Langerhans/metabolismo , Células de Langerhans/efectos de la radiación , Activación de Linfocitos/efectos de la radiación , Ratones Noqueados , Fenotipo , Placa Aterosclerótica , Transducción de Señal/efectos de la radiación , Piel/inmunología , Piel/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismoRESUMEN
BACKGROUND: Gut microbiome composition or circulating microbiome-related metabolites in patients with heart failure (HF) have not been investigated at different time points (i.e., in the decompensated (Decomp) and compensated (Comp) phases). MethodsâandâResults: We prospectively enrolled 22 patients admitted for HF and 11 age-, sex-, and comorbidity-matched hospitalized control subjects without a history of HF. Gut flora and plasma microbiome-related metabolites were evaluated by amplicon sequencing of the bacterial 16S ribosomal RNA gene and capillary electrophoresis time-of-flight mass spectrometry, respectively. HF patients were evaluated in both the Decomp and Comp phases during hospitalization. The phylum Actinobacteria was enriched in HF patients compared with control subjects. At the genus level, Bifiodobacterium was abundant while Megamonas was depleted in HF patients. Meanwhile, plasma concentration of trimethylamine N-oxide (TMAO), a gut microbiome-derived metabolite, was increased in HF patients (Decomp HF vs. control, P=0.003; Comp HF vs. control, P=0.004). A correlation analysis revealed positive correlations between the abundance of the genus Escherichia/Shigella and levels of TMAO and indoxyl sulfate (IS, a microbe-dependent uremic toxin) in Comp HF (TMAO: r=0.62, P=0.002; IS: r=0.63, P=0.002). Escherichia/Shigella was more abundant in Decomp than in Comp HF (P=0.030). CONCLUSIONS: Our results suggest that gut microbiome composition and microbiome-related metabolites are altered in HF patients.
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Bifidobacterium , Escherichia coli , Microbioma Gastrointestinal , Insuficiencia Cardíaca , Shigella , Anciano , Anciano de 80 o más Años , Bifidobacterium/clasificación , Bifidobacterium/aislamiento & purificación , Escherichia coli/clasificación , Escherichia coli/aislamiento & purificación , Femenino , Insuficiencia Cardíaca/sangre , Insuficiencia Cardíaca/microbiología , Insuficiencia Cardíaca/terapia , Humanos , Masculino , Persona de Mediana Edad , Shigella/clasificación , Shigella/aislamiento & purificaciónRESUMEN
The gut microbiota were shown to play critical roles in the development of atherosclerosis, but the detailed mechanism is limited. The purpose of this study is to clarify the influence of gut microbiota on atherogenesis via lipid metabolism and systemic inflammation. Germ-free or conventionally raised (Conv) ApoE-deficient (ApoE-/-) mice were fed chow diet and euthanized at 20 weeks of age. We found that the lack of gut microbiota in ApoE-/- mice caused a significant increase in the plasma and hepatic cholesterol levels compared with Conv ApoE-/- mice. The absence of gut microbiota changed the bile acid composition in the ileum, which was associated with activation of the enterohepatic fibroblast growth factor 15, fibroblast growth factor receptor 4 axis, and reduction of cholesterol 7α-hydroxylase and hepatic bile acid synthesis, resulting in the accumulation of liver cholesterol content. However, we found that the lack of microbiota caused a significant reduction in atherosclerotic lesion formation compared with Conv ApoE-/- mice, which might be associated with the attenuation of lipopolysaccharide-mediated inflammatory responses. Our findings indicated that the gut microbiota affected both hypercholesterolemia and atherogenesis in mice.
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Apolipoproteínas E/genética , Aterosclerosis/microbiología , Colesterol/metabolismo , Inflamación/microbiología , Animales , Aterosclerosis/genética , Aterosclerosis/metabolismo , Aterosclerosis/patología , Bacterias/metabolismo , Bacterias/patogenicidad , Colesterol/genética , Colesterol 7-alfa-Hidroxilasa/genética , Dieta , Modelos Animales de Enfermedad , Factores de Crecimiento de Fibroblastos/genética , Microbioma Gastrointestinal/genética , Homeostasis , Humanos , Íleon/metabolismo , Íleon/microbiología , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Metabolismo de los Lípidos/genética , Ratones , Receptor Tipo 4 de Factor de Crecimiento de Fibroblastos/genéticaRESUMEN
BACKGROUND: Previously, we have reported that daily glucose fluctuations could affect coronary plaque vulnerability, but the underlying mechanisms remained unclear. This study sought to investigate the impact of CD14++CD16+ monocytes on plaque vulnerability, as assessed by virtual histology intravascular ultrasound (VH-IVUS), as well as their relationship to fluctuating glucose levels in patients with asymptomatic coronary artery disease (CAD). METHODS: Fifty-one patients with asymptomatic CAD, who were undergoing lipid-lowering therapy and underwent VH-IVUS evaluation for angiographically mild to moderate lesions, were enrolled in the study. Standard VH-IVUS parameters, including the percentage volume of the necrotic core (%NC) within the plaque and the presence of a virtual histology thin-cap fibroatheroma (VH-TCFA), were then evaluated. Additionally, monocyte subsets were assessed by flow cytometry, and daily glucose fluctuations were analyzed by measuring the mean amplitude of glycemic excursion (MAGE). RESULTS: Among 82 plaques from 22 diabetes mellitus (DM) patients and 29 non-DM patients, 15 VH-TCFAs were identified. CD14++CD16+ monocyte counts significantly correlated with both %NC and the presence of VH-TCFA (%NC: r = 0.339, p = 0.002; VH-TCFA: p = 0.003). Multivariate logistic regression analysis revealed that CD14++CD16+ monocyte counts were independently associated with VH-TCFA (odds ratio = 1.029, p = 0.004). Furthermore, CD14++CD16+ monocyte counts were significantly correlated with the MAGE score in the non-DM patients (r = 0.544, p = 0.005). CONCLUSIONS: CD14++CD16+ monocyte levels are associated with coronary plaque vulnerability and can serve as a biomarker for VH-TCFA in patients with CAD undergoing lipid-lowering therapy. In patients without DM, glucose fluctuations may alter the balance of monocyte subsets. Trial registration UMIN Registry number: UMIN000021228.
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Enfermedad de la Arteria Coronaria/patología , Vasos Coronarios/diagnóstico por imagen , Monocitos/patología , Placa Aterosclerótica/patología , Anciano , Anciano de 80 o más Años , Angiografía Coronaria/métodos , Estudios Transversales , Diabetes Mellitus/inmunología , Diabetes Mellitus/patología , Femenino , Humanos , Receptores de Lipopolisacáridos/inmunología , Masculino , Persona de Mediana Edad , Receptores de IgG/inmunología , Factores de Riesgo , Ultrasonografía IntervencionalRESUMEN
OBJECTIVE: Although T-cell-mediated chronic inflammation contributes to atherosclerosis development, the role of a negative regulatory molecule cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) in atherosclerosis is poorly understood. We investigated the effects of CTLA-4 overexpression on atherosclerosis in apolipoprotein E-deficient (Apoe(-/-)) mice. APPROACH AND RESULTS: We generated CTLA-4 transgenic (CTLA-4-Tg)/Apoe(-/-) mice that display constitutive cell surface and intracellular expression of CTLA-4 in T cells and assessed atherosclerosis at age 16 weeks. CTLA-4 overexpression significantly reduced atherosclerotic lesion formation and intraplaque accumulation of macrophage and CD4(+) T cells in the aortic root compared with controls. CTLA-4-Tg/Apoe(-/-) mice showed decreased numbers of effector CD4(+) T cells and decreased expression of costimulatory molecules CD80 and CD86, ligands for CTLA-4, and a costimulatory molecule CD28, on CD11c(+) dendritic cells compared with controls. Consistent with in vivo findings, in vitro experiments revealed that CD4(+) T cells from CTLA-4-Tg/Apoe(-/-) mice showed decreased proliferative capacity and proinflammatory cytokine production, downregulated CD80 expression on CD11c(+) dendritic cells, and suppressed the proliferation of other T cells by limiting the costimulatory pathway. Moreover, CD11c(+) dendritic cells from CTLA-4-Tg/Apoe(-/-) mice showed reduced proliferative activity of T cells in vitro, suggesting the suppression of dendritic cell maturation in vivo. CONCLUSIONS: CTLA-4 regulates atherosclerosis by suppressing proatherogenic immune responses and could be an attractive therapeutic target for atherosclerosis.
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Aorta/metabolismo , Enfermedades de la Aorta/prevención & control , Aterosclerosis/prevención & control , Antígeno CTLA-4/metabolismo , Linfocitos T/metabolismo , Animales , Aorta/inmunología , Aorta/patología , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/inmunología , Enfermedades de la Aorta/metabolismo , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Aterosclerosis/genética , Aterosclerosis/inmunología , Aterosclerosis/metabolismo , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Antígeno CD11c/metabolismo , Antígenos CD28/metabolismo , Antígeno CTLA-4/genética , Proliferación Celular , Células Cultivadas , Células Dendríticas/metabolismo , Femenino , Genotipo , Activación de Linfocitos , Macrófagos/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Fenotipo , Placa Aterosclerótica , Transducción de Señal , Linfocitos T/inmunología , Regulación hacia ArribaRESUMEN
The association between atherosclerosis and gut microbiota has been attracting increased attention. We previously demonstrated a possible link between gut microbiota and coronary artery disease. Our aim of this study was to clarify the gut microbiota profiles in coronary artery disease patients using data mining analysis of terminal restriction fragment length polymorphism (T-RFLP). This study included 39 coronary artery disease (CAD) patients and 30 age- and sex- matched no-CAD controls (Ctrls) with coronary risk factors. Bacterial DNA was extracted from their fecal samples and analyzed by T-RFLP and data mining analysis using the classification and regression algorithm. Five additional CAD patients were newly recruited to confirm the reliability of this analysis. Data mining analysis could divide the composition of gut microbiota into 2 characteristic nodes. The CAD group was classified into 4 CAD pattern nodes (35/39 = 90 %), while the Ctrl group was classified into 3 Ctrl pattern nodes (28/30 = 93 %). Five additional CAD samples were applied to the same dividing model, which could validate the accuracy to predict the risk of CAD by data mining analysis. We could demonstrate that operational taxonomic unit 853 (OTU853), OTU657, and OTU990 were determined important both by the data mining method and by the usual statistical comparison. We classified the gut microbiota profiles in coronary artery disease patients using data mining analysis of T-RFLP data and demonstrated the possibility that gut microbiota is a diagnostic marker of suffering from CAD.
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Enfermedad de la Arteria Coronaria/diagnóstico , Enfermedad de la Arteria Coronaria/microbiología , ADN Bacteriano/genética , Microbioma Gastrointestinal , Polimorfismo de Longitud del Fragmento de Restricción , Anciano , Biomarcadores , Estudios de Casos y Controles , Minería de Datos , Heces/microbiología , Femenino , Humanos , Japón , Masculino , Persona de Mediana EdadRESUMEN
The intestinal microbiota appears to play an important role in the development of atherosclerosis. We investigated the effect of the probiotic lactic acid bacterium Pediococcus acidilactici R037 on atherosclerosis using apolipoprotein E-deficient (ApoE -/-) mice. Six-week-old ApoE -/- mice were orally administered R037 six times a week. Mice treated with R037 for 12 weeks exhibited markedly attenuated atherosclerotic lesions in the aortic root (2.3 ± 0.15 × 105 µm2 vs. 3.3 ± 0.29 × 105 µm2, respectively; P < 0.01; n = 15-17 each group). The expression of Ki-67 in CD4+ T cells, the population of interferon γ-producing CD4+ T cells in the spleen, and pro-inflammatory cytokine production from splenic lymphocytes were significantly decreased in R037-treated mice. Interestingly, splenic dendritic cells (DCs) isolated from R037-treated mice suppressed CD4+ T-cell proliferation and pro-inflammatory cytokine production ex vivo, suggesting that R037 treatment induced tolerogenic DCs. Programmed cell death ligand 1 expression in DCs was significantly enhanced in R037-treated mice, which might explain the immunosuppressive effect of DCs at least in part. These results indicate that R037 attenuates atherosclerosis by inducing tolerogenic DCs, which suppress Th1-driven inflammation and the proliferative activity of CD4+ T cells. Our findings may provide a novel therapeutic approach for the prevention of atherosclerosis based on dietary supplementation with probiotics.
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Aterosclerosis/prevención & control , Linfocitos T CD4-Positivos/inmunología , Células Dendríticas/inmunología , Pediococcus acidilactici , Probióticos/administración & dosificación , Administración Oral , Animales , Aorta/patología , Femenino , Ratones , Ratones Noqueados para ApoERESUMEN
Atherosclerosis is considered a chronic inflammatory disease and an intervention targeting the inflammatory process could be a new therapeutic strategy for preventing atherosclerotic cardiovascular diseases (CVD). We hypothesized that the intestine, which is considered the biggest immune organ in the human body, could be a therapeutic target for preventing CVD. We demonstrated that oral administration of anti-CD3 antibody or an active form of vitamin D3 reduced atherosclerosis in mice via induction of regulatory T cells and tolerogenic dendritic cells in the gut-associated lymphoid tissues. Similar to regulatory immune responses achieved by oral tolerance, our method had systemic effects that ultimately contributed towards atherosclerosis reduction. Recently, we have been interested in the gut microbiota, which have been reported as highly associated with intestinal immunity and systemic metabolic disorders, including obesity and diabetes. Notably, the guts of obese individuals are predominantly colonized by Firmicutes over Bacteroidetes. The association between atherosclerosis and microbiota has been attracting increased attention, and gut microbiota have been shown to participate in the metabolism of a proatherogenic compound called trimethylamine-N-oxide (TMAO) and aggravate CVD. Our investigation of the relationship between susceptibility to CVD and the gut microbiota revealed a characteristic flora type. Here, we discuss the evidence for the relationship between the gut microbiota and cardiometabolic diseases, and consider the gut microbiota as new potential therapeutic targets for treating CVD. (Circ J 2015; 79: 1882-1890).
Asunto(s)
Aterosclerosis , Bacteroidetes/inmunología , Firmicutes/inmunología , Microbioma Gastrointestinal/inmunología , Inmunidad Mucosa , Intestinos , Animales , Aterosclerosis/inmunología , Aterosclerosis/microbiología , Humanos , Intestinos/inmunología , Intestinos/microbiología , Metilaminas/inmunología , RatonesRESUMEN
BACKGROUND: The protective function of regulatory T cells (Treg) has been identified in experimental atherosclerosis, but the contribution of Treg to the pathogenesis of human coronary artery disease (CAD) remains poorly understood. We investigated Treg and regulatory T-cell/effector T-cell (Treg/Teff) ratio in peripheral blood samples from CAD patients using a new strategy for precise identification of Treg. METHODSâANDâRESULTS: Peripheral blood samples were collected from 73 stable CAD patients (55 middle-aged CAD patients and 18 old CAD patients) and 64 controls (47 middle-aged controls and 17 young controls). CD3(+)CD4(+)FoxP3(+)T cells were divided into 3 fractions: CD45RA(+)FoxP3(low)resting Treg(Fr1), CD45RA(-)FoxP3(high)activated Treg(Fr2), and CD45RA(-)FoxP3(low)non-Treg(Fr3). CAD patients had lower percentages of Fr1 and Fr2 and higher percentages of Fr3 and CD45RA(-)Foxp3(-)Teff(Fr4+5) within the CD3(+)CD4(+)T-cell population compared to age-matched controls. Treg/Teff ratio (Fr1+2/Fr3+4+5) in CAD patients was also markedly lower than in controls (middle-aged control, 0.17±0.09 vs. middle-aged CAD, 0.10±0.05; P<0.001). The percentage of CD4(+)CD28(null)T cells within the CD4(+)T-cell population was negatively correlated with Treg/Teff ratio, excluding CD4(+)CD28(null)T cells <0.3% (r=-0.27, P<0.05). High-sensitivity C-reactive protein was also negatively correlated with Treg/Teff ratio (r=-0.22, P<0.05). CONCLUSIONS: CAD patients had reduced Treg and Treg/Teff ratio compared to healthy controls. The present findings may be helpful when developing immunotherapy for the prevention of CAD.
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Angina de Pecho/inmunología , Infarto del Miocardio/inmunología , Subgrupos de Linfocitos T/inmunología , Linfocitos T Reguladores/inmunología , Adulto , Anciano , Envejecimiento/inmunología , Angina de Pecho/sangre , Antígenos de Diferenciación de Linfocitos T/análisis , Aterosclerosis/sangre , Aterosclerosis/inmunología , Glucemia/análisis , Femenino , Humanos , Lípidos/sangre , Activación de Linfocitos , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Infarto del Miocardio/sangreRESUMEN
Angiopoietin-like 3 (ANGPTL3) is a key regulator of lipoprotein metabolism, known for its potent inhibition on intravascular lipoprotein and endothelial lipase activities. Recent studies have shed light on the cellular functions of ANGPTL3. However, the precise mechanism underlying its regulation of cellular lipid metabolism remains elusive. We recently reported that ANGPTL3 interacts with the chromatin regulator SMARCAL1, which plays a pivotal role in maintaining cellular lipid homeostasis. Here, through a combination of in vitro and in vivo functional analyses, we provide evidence that ANGPTL3 indeed influences cellular lipid metabolism. Increased expression of Angptl3 prompted the formation of lipid droplets (LDs) in response to slow growth conditions. Notably, under the conditions, Angptl3 accumulated within cytoplasmic peroxisomes, where it interacts with SmarcAL1, which translocated from nucleus as observed previously. This translocation induced changes in gene expression favoring triglyceride (TG) accumulation. Indeed, ANGPTL3 gene knockout (KO) in human cells increased the expression of key lipid genes, which could be linked to elevated nuclear localization of SMARCAL1, whereas the expression of these genes decreased in SMARCAL1 KO cells. Consistent with these findings, the injection of Angptl3 protein to mice led to hepatic fat accumulation derived from circulating blood, a phenotype likely indicative of its long-term effect on blood TG, linked to SmarcAL1 activities. Thus, our results suggest that the Angptl3-SmarcAL1 pathway may confer the capacity for TG storage in cells in response to varying growth states, which may have broad implications for this pathway in regulating energy storage and trafficking.
RESUMEN
Population-based association studies have identified many genetic risk loci for coronary artery disease (CAD), but it is often unclear how genes within these loci are linked to CAD. Here, we perform interaction proteomics for 11 CAD-risk genes to map their protein-protein interactions (PPIs) in human vascular cells and elucidate their roles in CAD. The resulting PPI networks contain interactions that are outside of known biology in the vasculature and are enriched for genes involved in immunity-related and arterial-wall-specific mechanisms. Several PPI networks derived from smooth muscle cells are significantly enriched for genetic variants associated with CAD and related vascular phenotypes. Furthermore, the networks identify 61 genes that are found in genetic loci associated with risk of CAD, prioritizing them as the causal candidates within these loci. These findings indicate that the PPI networks we have generated are a rich resource for guiding future research into the molecular pathogenesis of CAD.
Asunto(s)
Enfermedad de la Arteria Coronaria , Humanos , Enfermedad de la Arteria Coronaria/genética , Mapas de Interacción de Proteínas , Redes Reguladoras de Genes , Sitios Genéticos , ProteómicaRESUMEN
Biallelic mutations of the chromatin regulator SMARCAL1 cause Schimke Immunoosseous Dysplasia (SIOD), characterized by severe growth defects and premature mortality. Atherosclerosis and hyperlipidemia are common among SIOD patients, yet their onset and progression are poorly understood. Using an integrative approach involving proteomics, mouse models, and population genetics, we investigated SMARCAL1's role. We found that SmarcAL1 interacts with angiopoietin-like 3 (Angptl3), a key regulator of lipoprotein metabolism. In vitro and in vivo analyses demonstrate SmarcAL1's vital role in maintaining cellular lipid homeostasis. The observed translocation of SmarcAL1 to cytoplasmic peroxisomes suggests a potential regulatory role in lipid metabolism through gene expression. SmarcAL1 gene inactivation reduces the expression of key genes in cellular lipid catabolism. Population genetics investigations highlight significant associations between SMARCAL1 genetic variations and body mass index, along with lipid-related traits. This study underscores SMARCAL1's pivotal role in cellular lipid metabolism, likely contributing to the observed lipid phenotypes in SIOD patients.
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Síndromes de Inmunodeficiencia , Animales , Humanos , Ratones , Cromatina , ADN Helicasas/genética , ADN Helicasas/metabolismo , Síndromes de Inmunodeficiencia/genética , Síndromes de Inmunodeficiencia/metabolismo , Metabolismo de los Lípidos/genética , LípidosRESUMEN
AIMS: Genetic studies have implicated the ARHGEF26 locus in the risk of coronary artery disease (CAD). However, the causal pathways by which DNA variants at the ARHGEF26 locus confer risk for CAD are incompletely understood. We sought to elucidate the mechanism responsible for the enhanced risk of CAD associated with the ARHGEF26 locus. METHODS AND RESULTS: In a conditional analysis of the ARHGEF26 locus, we show that the sentinel CAD-risk signal is significantly associated with various non-lipid vascular phenotypes. In human endothelial cell (EC), ARHGEF26 promotes the angiogenic capacity, and interacts with known angiogenic factors and pathways. Quantitative mass spectrometry showed that one CAD-risk coding variant, rs12493885 (p.Val29Leu), resulted in a gain-of-function ARHGEF26 that enhances proangiogenic signalling and displays enhanced interactions with several proteins partially related to the angiogenic pathway. ARHGEF26 is required for endothelial angiogenesis by promoting macropinocytosis of Vascular Endothelial Growth Factor Receptor 2 (VEGFR2) on cell membrane and is crucial to Vascular Endothelial Growth Factor (VEGF)-dependent murine vessel sprouting ex vivo. In vivo, global or tissue-specific deletion of ARHGEF26 in EC, but not in vascular smooth muscle cells, significantly reduced atherosclerosis in mice, with enhanced plaque stability. CONCLUSIONS: Our results demonstrate that ARHGEF26 is involved in angiogenesis signaling, and that DNA variants within ARHGEF26 that are associated with CAD risk could affect angiogenic processes by potentiating VEGF-dependent angiogenesis.
Asunto(s)
Factores de Intercambio de Guanina Nucleótido , Receptor 2 de Factores de Crecimiento Endotelial Vascular , Animales , Humanos , Ratones , Neovascularización Patológica , Neovascularización Fisiológica/fisiología , Transducción de Señal , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Factores de Intercambio de Guanina Nucleótido/genéticaRESUMEN
Human genetics studies have uncovered genetic variants that can be used to guide biological research and prioritize molecular targets for therapeutic intervention for complex diseases. We have identified a missense variant (P746S) in EDEM3 associated with lower blood triglyceride (TG) levels in >300,000 individuals. Functional analyses in cell and mouse models show that EDEM3 deficiency strongly increased the uptake of very-low-density lipoprotein and thereby reduced the plasma TG level, as a result of up-regulated expression of LRP1 receptor. We demonstrate that EDEM3 deletion up-regulated the pathways for RNA and endoplasmic reticulum protein processing and transport, and consequently increased the cell surface mannose-containing glycoproteins, including LRP1. Metabolomics analyses reveal a cellular TG accumulation under EDEM3 deficiency, a profile consistent with individuals carrying EDEM3 P746S. Our study identifies EDEM3 as a regulator of blood TG, and targeted inhibition of EDEM3 may provide a complementary approach for lowering elevated blood TG concentrations.
RESUMEN
Vascular inflammation via T-cell-mediated immune responses has been shown to be critically involved in the pathogenesis of abdominal aortic aneurysm (AAA). T-cell coinhibitory molecule cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) is known to act as a potent negative regulator of immune responses. However, the role of this molecule in the development of AAA remains completely unknown. We determined the effects of CTLA-4 overexpression on experimental AAA. We continuously infused CTLA-4 transgenic (CTLA-4-Tg)/apolipoprotein E-deficient (Apoe-/-) mice or control Apoe-/- mice fed a high-cholesterol diet with angiotensin II by implanting osmotic mini-pumps and evaluated the development of AAA. Ninety percent of angiotensin II-infused mice developed AAA, with 50% mortality because of aneurysm rupture. Overexpression of CTLA-4 significantly reduced the incidence (66%), mortality (26%), and diameter of AAA. These protective effects were associated with a decreased number of effector CD4+ T cells and the downregulated expression of costimulatory molecules CD80 and CD86, ligands for CTLA-4, on CD11c+ dendritic cells in lymphoid tissues. CTLA-4-Tg/Apoe-/- mice had reduced accumulation of macrophages and CD4+ T cells, leading to attenuated aortic inflammation, preserved vessel integrity, and decreased susceptibility to AAA and aortic rupture. Our findings suggest T-cell coinhibitory molecule CTLA-4 as a novel therapeutic target for AAA.
Asunto(s)
Aorta Abdominal/patología , Aneurisma de la Aorta Abdominal/inmunología , Rotura de la Aorta/inmunología , Aterosclerosis/complicaciones , Antígeno CTLA-4/metabolismo , Angiotensina II/toxicidad , Animales , Aorta Abdominal/efectos de los fármacos , Aorta Abdominal/inmunología , Aneurisma de la Aorta Abdominal/inducido químicamente , Aneurisma de la Aorta Abdominal/diagnóstico , Aneurisma de la Aorta Abdominal/patología , Rotura de la Aorta/inducido químicamente , Rotura de la Aorta/patología , Aterosclerosis/inmunología , Antígeno CTLA-4/genética , Antígeno CTLA-4/inmunología , Dieta Aterogénica/efectos adversos , Modelos Animales de Enfermedad , Humanos , Masculino , Ratones , Ratones Noqueados para ApoE , Índice de Severidad de la Enfermedad , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
BACKGROUND: Pathogenic immune responses are known to play an important role in abdominal aortic aneurysm (AAA) development. Ultraviolet B (UVB) irradiation has been demonstrated to have therapeutic potential not only for cutaneous diseases but also for systemic inflammatory diseases in mice by suppressing immunoinflammatory responses. We investigated the effect of UVB irradiation on experimental AAA. METHODS AND RESULTS: We used an angiotensin II-induced AAA model in apolipoprotein E-deficient mice fed a high-cholesterol diet. Mice aged 10 weeks were irradiated with 5 kJ/m2 UVB once weekly for 6 weeks (UVB-irradiated, n=38; nonirradiated, n=42) and were euthanized for evaluation of AAA formation at 16 weeks. Overall, 93% of angiotensin II-infused mice developed AAA, with 60% mortality possibly because of aneurysm rupture. UVB irradiation significantly decreased the incidence (66%) and mortality (29%) of AAA (P=0.004 and P=0.006, respectively). UVB-irradiated mice had significantly smaller diameter AAA (P=0.008) and fewer inflammatory cells in the aortic aneurysm tissue than nonirradiated mice, along with systemic expansion of CD4+Foxp3+ regulatory T cells and decreased effector CD4+CD44highCD62Llow T cells in para-aortic lymph nodes. Genetic depletion of regulatory T cells abrogated these beneficial effects of UVB treatment, demonstrating a critical role of regulatory T cells. CONCLUSIONS: Our data suggest that UVB-dependent expansion of regulatory T cells has beneficial effects on experimental AAA and may provide a novel strategy for the treatment of AAA.