RESUMEN
Optimal vascular structure and function are essential for maintaining the physiological functions of the cardiovascular system. Vascular remodelling involves changes in vessel structure, including its size, shape, cellular and molecular composition. These changes result from multiple risk factors and may be compensatory adaptations to sustain blood vessel function. They occur in diverse cardiovascular pathologies, from hypertension to heart failure and atherosclerosis. Dynamic changes in the endothelium, fibroblasts, smooth muscle cells, pericytes or other vascular wall cells underlie remodelling. In addition, immune cells, including macrophages and lymphocytes, may infiltrate vessels and initiate inflammatory signalling. They contribute to a dynamic interplay between cell proliferation, apoptosis, migration, inflammation, and extracellular matrix reorganisation, all critical mechanisms of vascular remodelling. Molecular pathways underlying these processes include growth factors (e.g., vascular endothelial growth factor and platelet-derived growth factor), inflammatory cytokines (e.g., interleukin-1ß and tumour necrosis factor-α), reactive oxygen species, and signalling pathways, such as Rho/ROCK, MAPK, and TGF-ß/Smad, related to nitric oxide and superoxide biology. MicroRNAs and long noncoding RNAs are crucial epigenetic regulators of gene expression in vascular remodelling. We evaluate these pathways for potential therapeutic targeting from a clinical translational perspective. In summary, vascular remodelling, a coordinated modification of vascular structure and function, is crucial in cardiovascular disease pathology.
Asunto(s)
Enfermedades Cardiovasculares , Hipertensión , Inflamación , Remodelación Vascular , Humanos , Inflamación/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Enfermedades Cardiovasculares/metabolismo , Hipertensión/fisiopatología , Hipertensión/metabolismo , Animales , Estrés Oxidativo , Transducción de Señal , Oxidación-ReducciónRESUMEN
Multiple endocrine neoplasia type 1 (MEN1) is a syndrome characterized by tumors in multiple organs. Although being a dominantly inherited monogenic disease, disease phenotypes are unpredictable and differ even among members of the same family. There is growing evidence for the role of modifier genes in the alteration of the course of this disease. However, genome-wide screening data are still lacking. In our study, we addressed the different outcomes of the disease, focusing on pituitary and adrenocortical tumors. By means of exome sequencing we identified the affected signaling pathways that segregated with those symptoms. Most significantly, we identified damaging alterations in numerous structural genes responsible for cell adhesion and migration. Additionally, in the case of pituitary tumors, genes related to neuronal function, survival, and morphogenesis were repeatedly identified, while in patients with adrenocortical tumors, TLR10, which is involved in the regulation of the innate immunity, was commonly modified. Our data show that using exome screening, it is possible to find signatures which correlate with the given clinical MEN1 outcomes, providing evidence that studies addressing modifier effects in MEN1 are reasonable.
Asunto(s)
Neoplasias de la Corteza Suprarrenal , Neoplasia Endocrina Múltiple Tipo 1 , Humanos , Neoplasia Endocrina Múltiple Tipo 1/diagnóstico , Neoplasia Endocrina Múltiple Tipo 1/genética , Exoma , Adhesión Celular , Transducción de Señal/genéticaRESUMEN
BACKGROUND: The use of doxorubicin is associated with an increased risk of acute and long-term cardiomyopathy. Despite the constantly growing number of cancer survivors, little is known about the transcriptional mechanisms which progress in the time leading to a severe cardiac outcome. It is also unclear whether long-term transcriptomic alterations related to doxorubicin use are similar to transcriptomic patterns present in patients suffering from other cardiomyopathies. METHODS: We have sequenced miRNA from total plasma and extracellular vesicles (EVs) from 66 acute lymphoblastic leukemia (ALL) survivors and 61 healthy controls (254 samples in total). We then analyzed processes regulated by differentially expressed circulating miRNAs and cross-validated results with the data of patients with clinically manifested cardiomyopathies. RESULTS: We found that especially miRNAs contained within EVs may be informative in terms of cardiomyopathy development and may regulate pathways related to neurotrophin signaling, transforming growth factor beta (TGFß) or epidermal growth factor receptors (ErbB). We identified vesicular miR-144-3p and miR-423-3p as the most variable between groups and significantly correlated with echocardiographic parameters and, respectively, for plasma: let-7g-5p and miR-16-2-3p. Moreover, vesicular miR-144-3p correlates with the highest number of echocardiographic parameters and is differentially expressed in the circulation of patients with dilated cardiomyopathy. We also found that distribution of particular miRNAs between of plasma and EVs (proportion between compartments) e.g., miR-184 in ALL, is altered, suggesting changes within secretory and miRNA sorting mechanisms. CONCLUSIONS: Our results show that transcriptomic changes resulting from doxorubicin induced myocardial injury are reflected in circulating miRNA levels and precede development of the late onset cardiomyopathy phenotype. Among miRNAs related to cardiac function, we found vesicular miR-144-3p and miR-423-3p, as well as let-7g-5p and miR-16-2-3p contained in the total plasma. Selection of source for such studies (plasma or EVs) is of critical importance, as distribution of some miRNA between plasma and EVs is altered in ALL survivors, in comparison to healthy people, which suggests that doxorubicin-induced changes include miRNA sorting and export to extracellular space.
Asunto(s)
MicroARN Circulante , Vesículas Extracelulares , MicroARNs , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Cardiotoxicidad/etiología , Cardiotoxicidad/metabolismo , Vesículas Extracelulares/metabolismo , MicroARN Circulante/genética , MicroARN Circulante/metabolismo , Doxorrubicina/efectos adversosRESUMEN
Hyperphagia and obesity, which underlie metabolic syndrome, have been linked to multiple health complications and increased mortality. Here, we investigate the differences in plasma proteome between obese and lean Zucker rats in order to identify circulating proteins involved in obesity-related conditions. Plasma samples of male Zucker fatty (obese) rats carrying fatty fa/fa mutation (-/-) and their lean controls were enriched using ProteoMiner technology and labeled with isobaric tags (iTRAQ) for mass spectrometry-based quantitation. We found elevation in levels of coagulation factors whereas levels of serine protease inhibitors were decreased. Levels of acute phase proteins were also altered, as well as complement components. We also noticed differences in the abundance of apolipoproteins. In summary, quantitative proteomic assessment of plasma protein composition in obese Zucker rats revealed a profound landscape of changes, reflecting altered hemostasis, disturbed metabolic processes involving insulin resistance and lipid metabolism and ongoing low-grade inflammation.
Asunto(s)
Enfermedades Cardiovasculares , Estado Prediabético , Masculino , Animales , Ratas , Ratas Zucker , Proteoma , Proteómica , Factores de Riesgo , Factores de Riesgo de Enfermedad Cardiaca , ObesidadRESUMEN
Cancer-specific isoenzyme of phosphofructokinase II (PFKFB4), as our previous research has shown, may be one of the most important enzymes contributing to the intensification of glycolysis in hypoxic malignant melanoma cells. Although the PFKFB4 gene seems to play a crucial role in the progression of melanoma, so far there are no complete data on the expression of PFKFB4 at the isoform level and the influence of hypoxia on alternative splicing. Using RT-qPCR and semi-quantitative RT-PCR, we presented the PFKFB4 gene expression profile at the level of six isoforms described in the OMIM NCBI database in normoxic and hypoxic melanoma cells. Additionally, using VMD software, we analyzed the structure of isoforms at the protein level, concluding about the catalytic activity of individual isoforms. Our research has shown that five isoforms of PFKFB4 are expressed in melanoma cells, of which the D and F isoforms are highly constitutive, while the canonical B isoform seems to be the main isoform induced in hypoxia. Our results also indicate that the expression profile at the level of the PFKFB4 gene does not reflect the expression at the level of individual isoforms. Our work clearly indicates that the PFKFB4 gene expression profile should be definitely analyzed at the level of individual isoforms. Moreover, the analysis at the protein level allowed the selection of those isoforms whose functional validation should be performed to fully understand the importance of PFKFB4 expression in the metabolic adaptation of malignant melanoma cells.
Asunto(s)
Empalme Alternativo , Biomarcadores de Tumor/metabolismo , Regulación Neoplásica de la Expresión Génica , Hipoxia/fisiopatología , Melanoma/patología , Oxígeno/metabolismo , Fosfofructoquinasa-2/genética , Biomarcadores de Tumor/genética , Glucólisis , Humanos , Melanoma/genética , Melanoma/metabolismo , Células Tumorales CultivadasRESUMEN
Atherosclerosis and nonalcoholic fatty liver disease are leading causes of morbidity and mortality in the Western countries. The renin-angiotensin system (RAS) with its two main opposing effectors, i.e., angiotensin II (Ang II) and Ang-(1-7), is widely recognized as a major regulator of cardiovascular function and body metabolic processes. Angiotensin-converting enzyme 2 (ACE2) by breaking-down Ang II forms Ang-(1-7) and thus favors Ang-(1-7) actions. Therefore, the aim of our study was to comprehensively evaluate the influence of prolonged treatment with ACE2 activator, diminazene aceturate (DIZE) on the development of atherosclerotic lesions and hepatic steatosis in apoE-/- mice fed a high-fat diet (HFD). We have shown that DIZE stabilized atherosclerotic lesions and attenuated hepatic steatosis in apoE-/- mice fed an HFD. Such effects were associated with decreased total macrophages content and increased α-smooth muscle actin levels in atherosclerotic plaques. Moreover, DIZE changed polarization of macrophages towards increased amount of anti-inflammatory M2 macrophages in the atherosclerotic lesions. Interestingly, the anti-steatotic action of DIZE in the liver was related to the elevated levels of HDL in the plasma, decreased levels of triglycerides, and increased biosynthesis and concentration of taurine in the liver of apoE-/- mice. However, exact molecular mechanisms of both anti-atherosclerotic and anti-steatotic actions of DIZE require further investigations.
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Enzima Convertidora de Angiotensina 2/genética , Aterosclerosis/tratamiento farmacológico , Diminazeno/análogos & derivados , Hígado Graso/tratamiento farmacológico , Placa Aterosclerótica/tratamiento farmacológico , Taurina/biosíntesis , Angiotensina I/genética , Angiotensina I/metabolismo , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Aterosclerosis/etiología , Aterosclerosis/genética , Aterosclerosis/patología , Dieta Alta en Grasa , Diminazeno/farmacología , Modelos Animales de Enfermedad , Hígado Graso/etiología , Hígado Graso/genética , Hígado Graso/patología , Femenino , Regulación de la Expresión Génica , Humanos , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Activación de Macrófagos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Macrófagos/patología , Arterias Mesentéricas/efectos de los fármacos , Arterias Mesentéricas/metabolismo , Arterias Mesentéricas/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/metabolismo , Placa Aterosclerótica/etiología , Placa Aterosclerótica/genética , Placa Aterosclerótica/patología , Células THP-1 , Taurina/agonistasRESUMEN
Atherosclerosis and NAFLD are the leading causes of death worldwide. The hallmark of NAFLD is triglyceride accumulation caused by an imbalance between lipogenesis de novo and fatty acid oxidation. Agmatine, an endogenous metabolite of arginine, exerts a protective effect on mitochondria and can modulate fatty acid metabolism. In the present study, we investigate the influence of agmatine on the progression of atherosclerotic lesions and the development of hepatic steatosis in apoE-/- mice fed with a Western high-fat diet, with a particular focus on its effects on the DNL pathway in the liver. We have proved that treatment of agmatine inhibits the progression of atherosclerosis and attenuates hepatic steatosis in apoE-/- mice on a Western diet. Such effects are associated with decreased total macrophage content in atherosclerotic plaque as well as a decrease in the TG levels and the TG/HDL ratio in plasma. Agmatine also reduced TG accumulation in the liver and decreased the expression of hepatic genes and proteins involved in lipogenesis de novo such as SREBP-1c, FASN and SCD1. In conclusion, agmatine may present therapeutic potential for the treatment of atherosclerosis and fatty liver disease. However, an exact understanding of the mechanisms of the advantageous actions of agmatine requires further study.
Asunto(s)
Agmatina/efectos adversos , Aterosclerosis/etiología , Aterosclerosis/metabolismo , Dieta Occidental , Hígado Graso/etiología , Hígado Graso/metabolismo , Lípidos/sangre , Lipogénesis , Animales , Aterosclerosis/sangre , Aterosclerosis/patología , Biomarcadores , HDL-Colesterol/sangre , Dieta Occidental/efectos adversos , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Hígado Graso/sangre , Hígado Graso/patología , Femenino , Inmunohistoquímica , Metabolismo de los Lípidos , Ratones , Ratones Noqueados para ApoE , Triglicéridos/sangreRESUMEN
The reciprocal interactions between cancer cells and the quiescent fibroblasts leading to the activation of cancer-associated fibroblasts (CAFs) serve an important role in cancer progression. Here, we investigated the activation of transcription factors (TFs) in prostate fibroblasts (WPMY cell line) co-cultured with normal prostate or tumorous cells (RWPE1 and RWPE2 cell lines, respectively). After indirect co-cultures, we performed mRNA-seq and predicted TF activity using mRNA expression profiles with the Systems EPigenomics Inference of Regulatory Activity (SEPIRA) package and the GTEx and mRNA-seq data of 483 cultured fibroblasts. The initial differential expression analysis between time points and experimental conditions showed that co-culture with normal epithelial cells mainly promotes an inflammatory response in fibroblasts, whereas with the cancerous epithelial, it stimulates transformation by changing the expression of the genes associated with microfilaments. TF activity analysis revealed only one positively regulated TF in the RWPE1 co-culture alone, while we observed dysregulation of 45 TFs (7 decreased activity and 38 increased activity) uniquely in co-culture with RWPE2. Pathway analysis showed that these 45 dysregulated TFs in fibroblasts co-cultured with RWPE2 cells may be associated with the RUNX1 and PTEN pathways. Moreover, we showed that observed dysregulation could be associated with FER1L4 expression. We conclude that phenotypic changes in fibroblast responses to co-culturing with cancer epithelium result from orchestrated dysregulation of signaling pathways that favor their transformation and motility rather than proinflammatory status. This dysregulation can be observed both at the TF and transcriptome levels.
Asunto(s)
Fibroblastos Asociados al Cáncer/metabolismo , Transformación Celular Neoplásica/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Regulación Neoplásica de la Expresión Génica , Fosfohidrolasa PTEN/genética , Factores de Transcripción/genética , Fibroblastos Asociados al Cáncer/patología , Comunicación Celular , Línea Celular , Línea Celular Tumoral , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Técnicas de Cocultivo , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Células Epiteliales/metabolismo , Células Epiteliales/patología , Fibroblastos/citología , Fibroblastos/metabolismo , Perfilación de la Expresión Génica , Ontología de Genes , Humanos , Masculino , Anotación de Secuencia Molecular , Fosfohidrolasa PTEN/metabolismo , Próstata/metabolismo , Próstata/patología , Transducción de Señal , Factores de Transcripción/clasificación , Factores de Transcripción/metabolismo , TranscriptomaRESUMEN
Anterior segment dysgenesis (ASD) encompasses a spectrum of ocular disorders affecting the structures of the anterior eye chamber. Mutations in several genes, involved in eye development, are implicated in this disorder. ASD is often accompanied by diverse multisystemic symptoms and another genetic cause, such as variants in genes encoding collagen type IV. Thus, a wide spectrum of phenotypes and underlying genetic diversity make fast and proper diagnosis challenging. Here, we used AMELIE, an automatic text mining tool that enriches data with the most up-to-date information from literature, and wANNOVAR, which is based on well-documented databases and incorporates variant filtering strategy to identify genetic variants responsible for severely-manifested ASD in a newborn child. This strategy, applied to trio sequencing data in compliance with ACMG 2015 guidelines, helped us find two compound heterozygous variants of the B3GLCT gene, of which c.660+1G>A (rs80338851) was previously associated with the phenotype of Peters plus syndrome (PPS), while the second, NM_194318.3:c.755delC (p.T252fs), in exon 9 of the same gene was noted for the first time. PPS, a very rare subtype of ASD, is a glycosylation disorder, where the dysfunctional B3GLCT gene product, O-fucose-specific ß-1,3-glucosyltransferase, is ineffective in providing a noncanonical quality control system for proper protein folding in cells. Our study expands the mutation spectrum of the B3GLCT gene related to PPS. We suggest that the implementation of automatic text mining tools in combination with careful variant filtering could help translate sequencing results into diagnosis, thus, considerably accelerating the diagnostic process and, thereby, improving patient management.
Asunto(s)
Labio Leporino/genética , Córnea/anomalías , Galactosiltransferasas/genética , Pruebas Genéticas/métodos , Glucosiltransferasas/genética , Trastornos del Crecimiento/genética , Deformidades Congénitas de las Extremidades/genética , Mutación , Segmento Anterior del Ojo/patología , Labio Leporino/diagnóstico , Minería de Datos/métodos , Trastornos del Crecimiento/diagnóstico , Humanos , Recién Nacido , Deformidades Congénitas de las Extremidades/diagnóstico , Análisis de Secuencia de ADN/métodosRESUMEN
Atherosclerosis and nonalcoholic fatty liver disease (NAFLD) are frequent causes of death in the Western countries. Recently, it has been shown that autophagy dysfunction plays an important role in the pathogenesis of both atherosclerosis and NAFLD; thus, activators of autophagy might be useful for novel therapeutic interventions. Trehalose-a naturally occuring disaccharide present in plants, bacteria, fungi, insects, and certain types of shrimps-is a known inducer of autophagy. However, according to the literature, its anti-atherosclerotic and anti-steatotic potential seem to depend on the experimental setting. The aim of our study was to comprehensively describe the influence of a prolonged treatment with orally administered trehalose on the development of atherosclerotic lesions and hepatic steatosis in apolipoprotein E knockout (apoE-/-) mice in an experimental set up reflecting both moderate and severe proatherogenic conditions: male apoE-/- mice on a chow diet (CD) and female apoE-/- mice fed with a high-fat diet (HFD). We found that exogenous trehalose inhibited atherosclerosis and attenuated hepatic steatosis in apoE-/- mice. Such effects of trehalose were not associated with changes of plasma cholesterol, low-density lipoproteins (LDL), or high-density lipoproteins (HDL). Moreover, the anti-steatotic action of trehalose in the liver was associated with the induction of autophagy. The exact molecular mechanisms of both the anti-atherosclerotic action of trehalose and its inhibitory effect on liver steatosis require further clarification.
Asunto(s)
Apolipoproteínas E/genética , Aterosclerosis/tratamiento farmacológico , Autofagia/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Trehalosa/uso terapéutico , Animales , Aorta/efectos de los fármacos , Aorta/patología , Aterosclerosis/etiología , Aterosclerosis/genética , Aterosclerosis/patología , Dieta Alta en Grasa/efectos adversos , Femenino , Hígado/efectos de los fármacos , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patología , Trehalosa/administración & dosificación , Trehalosa/farmacologíaRESUMEN
It is unknown whether fibrosis-associated microRNAs: miR-21, miR-26, miR-29, miR-30 and miR-133a are linked to cardiovascular (CV) outcome. The study evaluated the levels of extracellular matrix (ECM) fibrosis and the prevalence of particular microRNAs in patients with dilated cardiomyopathy (DCM) to investigate any correlation with CV events. METHODS: Seventy DCM patients (48 ± 12 years, EF 24.4 ± 7.4%) underwent right ventricular biopsy. The control group was comprised of 7 patients with CAD who underwent CABG and intraoperative biopsy. MicroRNAs were measured in blood and myocardial tissue via qPCR. The end-point was a combination of CV death and urgent HF hospitalization at the end of 12 months. There were differential levels of circulating and myocardial miR-26 and miR-29 as well as myocardial miR-133a when the DCM and CABG groups were compared. Corresponding circulating and myocardial microRNAs did not correlate with one another. There was no correlation between microRNA and ECM fibrosis. By the end of the 12-month period of the study, CV death had occurred in 6 patients, and a further 19 patients required urgent HF hospitalization. None of the circulating microRNAs was a predictor of the combined end-point; however, myocardial miR-133a was an independent predictor in unadjusted models (HR 1.53; 95% CI 1.14-2.05; P < .004) and adjusted models (HR 1.57; 95% CI 1.14-2.17; P < .005). The best cut-off value for the miR-133a level for the prediction of the combined end-point was 0.74 ΔCq, with an AUC of 0.67. The absence of a correlation between the corresponding circulating and myocardial microRNAs calls into question their cellular source. This study sheds new light on the role of microRNAs in ECM fibrosis in DCM, which warrants further exploration.
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Cardiomiopatía Dilatada/genética , Fibrosis/genética , Ventrículos Cardíacos/metabolismo , MicroARNs/genética , Biomarcadores/sangre , Biopsia , Cardiomiopatía Dilatada/sangre , Cardiomiopatía Dilatada/fisiopatología , Matriz Extracelular/genética , Femenino , Fibrosis/sangre , Fibrosis/fisiopatología , Ventrículos Cardíacos/patología , Humanos , Masculino , MicroARNs/sangre , Persona de Mediana Edad , Miocardio/metabolismo , Miocardio/patologíaRESUMEN
Atherosclerosis is an inflammatory disease in which dysfunction of mitochondria play an important role, and disorders of lipid management intensify this process. Agmatine, an endogenous polyamine formed by decarboxylation of arginine, exerts a protective effect on mitochondria and modulates fatty acid metabolism. We investigated the effect of exogenous agmatine on the development of atherosclerosis and changes in lipid profile in apolipoprotein E knockout (apoE-/-) mice. Agmatine caused an approximate 40% decrease of atherosclerotic lesions, as estimated by en face and cross-section methods with an influence on macrophage but not on smooth muscle content in the plaques. Agmatine treatment did not changed gelatinase activity within the plaque area. What is more, the action of agmatine was associated with an increase in the number of high density lipoproteins (HDL) in blood. Real-Time PCR analysis showed that agmatine modulates liver mRNA levels of many factors involved in oxidation of fatty acid and cholesterol biosynthesis. Two-dimensional electrophoresis coupled with mass spectrometry identified 27 differentially expressed mitochondrial proteins upon agmatine treatment in the liver of apoE-/- mice, mostly proteins related to metabolism and apoptosis. In conclusion, prolonged administration of agmatine inhibits atherosclerosis in apoE-/- mice; however, the exact mechanisms linking observed changes and elevations of HDL plasma require further investigation.
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Agmatina/uso terapéutico , Apolipoproteínas E/deficiencia , Aterosclerosis/tratamiento farmacológico , Agmatina/farmacología , Animales , Aorta/efectos de los fármacos , Aorta/patología , Apolipoproteínas E/metabolismo , Aterosclerosis/sangre , Peso Corporal/efectos de los fármacos , Electroforesis en Gel Bidimensional , Ácidos Grasos/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias Hepáticas/efectos de los fármacos , Mitocondrias Hepáticas/metabolismo , Proteínas Mitocondriales/metabolismo , Proteoma/metabolismoRESUMEN
Decreased plasma levels of microRNA-223 (miR-223), predominantly of platelet origin, were proposed as a surrogate marker of efficacy of antiplatelet therapy. However, higher on-treatment platelet reactivity was associated with lower plasma miR-223 in patients with coronary artery disease (CAD) on dual antiplatelet therapy (DAPT) including clopidogrel and aspirin. Our aim was to compare plasma miR-223 and platelet reactivity in CAD patients on DAPT with newer P2Y12 antagonists vs. clopidogrel. We studied 21 men with CAD admitted to our centre owing to a non-ST-elevation acute coronary syndrome, and with an uncomplicated hospital course. From the day of admission, the patients were receiving either clopidogrel (n = 11) or prasugrel/ticagrelor (n = 10) in addition to aspirin. Before discharge, miR-223 expression in plasma was estimated by quantitative polymerase chain reaction using the comparative Ct method relative to miR-16 as an endogenous control. Multiple electrode aggregometry was used to assess platelet aggregation in response to adenosine diphosphate (ADP). ADP-induced platelet reactivity was decreased in the patients treated with prasugrel or ticagrelor compared with those on clopidogrel (mean ± SD: 139 ± 71 vs. 313 ± 162 arbitrary units [AU]*min, p = 0.006), due to a more potent antiplatelet activity of the novel P2Y12 antagonists. Consequently, six out of seven patients in the lower tertile of the ADP-induced platelet aggregation were treated with the newer P2Y12 blockers, whereas six out of seven patients in the upper tertile were on clopidogrel. Plasma miR-223 was elevated with decreasing platelet reactivity (Spearman's rho = -0.52; p = 0.015 for trend), being significantly higher in the lower tertile of the ADP-induced platelet aggregation (median [range]: 1.06 [0.25-2.31]) vs. the upper tertile (0.20 [0.13-2.30]) (p = 0.04). In conclusion, our preliminary results argue against the notion of low plasma miR-223 as a marker of platelet responsiveness to DAPT. On the contrary, more potent platelet inhibition associated mainly with newer P2Y12 antagonists appears to coincide with higher miR-223 relative to the subjects with attenuated responsiveness to DAPT.
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Plaquetas/metabolismo , Enfermedad de la Arteria Coronaria/genética , Enfermedad de la Arteria Coronaria/metabolismo , MicroARNs/genética , Activación Plaquetaria , Anciano , Aspirina/uso terapéutico , Biomarcadores , Clopidogrel , Enfermedad de la Arteria Coronaria/diagnóstico , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Femenino , Humanos , Masculino , MicroARNs/sangre , Persona de Mediana Edad , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/uso terapéutico , Ticlopidina/análogos & derivados , Ticlopidina/uso terapéutico , Resultado del TratamientoRESUMEN
INTRODUCTION: Multiple endocrine neoplasia type 1 (MEN1) is a monogenic disease caused by inactivating variants in the MEN1 gene. Although the reason for its development is well-known, disease phenotypes are unpredictable and differ even among carriers of the same pathogenic driver mutation. Genetic, epigenetic, and environmental factors may play a role in driving the individual phenotype. Those factors, however, still mostly remain unidentified. In our work, we focused on the inherited genetic background in pancreatic neuroendocrine neoplasms (pNENs) in MEN1 patients, and the pancreatic tumour subgroup with insulinoma. MATERIAL AND METHODS: Whole exome sequencing was performed in MEN1 patients. The symptoms of interest were pancreatic neuroendocrine tumours in one analysis and insulinoma in the second. The study included families as well as unrelated cases. Genes with variants that are not neutral to the encoded gene product were defined in symptom-positive patients as compared to symptom-negative controls. The interpretation of the results was based on functional annotations and pathways shared between all patients with the given symptom in the course of MEN1. RESULTS: Whole-exome screening of family members and unrelated patients with and without pNENs revealed a number of pathways that are common for all the analysed cases with pNENs. Those included pathways crucial for morphogenesis and development, proper insulin signalling, and structural cellular organization. An additional analysis of insulinoma pNEN patients revealed additional pathways engaged in glucose and lipid homeostasis, and several non-canonical insulin-regulating mechanisms. CONCLUSIONS: Our results show the existence of pathways that are identified in a non-literature-predefined manner, which might have a modifying function in MEN1, differentiating the specific clinical outcomes. Those results, although preliminary, provide evidence of the reasonableness of performing large-scale studies addressing the genetic background of MEN1 patients in determining their individual outcomes.
Asunto(s)
Insulinoma , Neoplasia Endocrina Múltiple Tipo 1 , Tumores Neuroendocrinos , Neoplasias Pancreáticas , Humanos , Insulinoma/genética , Neoplasia Endocrina Múltiple Tipo 1/genética , Secuenciación del Exoma , Tumores Neuroendocrinos/genética , Neoplasias Pancreáticas/genética , Insulina , Antecedentes GenéticosRESUMEN
INTRODUCTION: Familial hypercholesterolemia (FH) is an autosomal dominant monogenic lipid metabolism disorder characterized by a significantly elevated level of lowdensity lipoprotein (LDL) cholesterol and leading to premature ischemic heart disease. FH is caused by mutations in the LDLR, APOB, and PCSK9 genes; however, these mutations account for only about 40% of FH cases. In order to obtain a genetic diagnosis of FH, sequencing of other genes involved in the lipid metabolism might be useful. OBJECTIVES: This study aimed to describe genetic variants in genes associated with FH in a group of patients from the Malopolska province in Southern Poland, using the targeted next generation sequencing (NGS) technology. PATIENTS AND METHODS: The study involved 90 unrelated adults (age range, 18-70 years) with FH diagnosed clinically according to the Simon Broome Register criteria. A customdesigned capture assay and the Illumina MiSeq platform were used. The panel included exons and exon / intron boundaries of known FHcausing genes: LDLR, APOB, and PCSK9, as well as genes previously associated with high cholesterol levels: APOE, ABCG5, ABCG8, LPL, NPC1, LDLRAP1, LIPC, STAP1, and CELSR2. Genetic variants were classified based on in silico predictions and ClinVar reports. RESULTS: We detected 4 patients with variants in the LDLR and APOB genes that had not been previously linked to FH in ClinVar. We also found APOB mutations outside the common LDL receptor-binding region, in exons 26 and 29. Interestingly, we observed a high frequency of pathogenic variants in exon 4 of the APOE gene: rs7412, probably damaging (4 patients) and rs429358, benign (16 patients). CONCLUSIONS: NGS is a useful and reliable method to detect new variants in genes related to FH. In addition, the results enable the detection of FH phenocopies and introduction of appropriate treatment.
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Hiperlipoproteinemia Tipo II , Proproteína Convertasa 9 , Adulto , Humanos , Adolescente , Adulto Joven , Persona de Mediana Edad , Anciano , Proproteína Convertasa 9/genética , Polonia , Hiperlipoproteinemia Tipo II/genética , Hiperlipoproteinemia Tipo II/diagnóstico , Apolipoproteínas B , Apolipoproteínas ERESUMEN
Age represents a major risk factor in heart failure (HF). However, the mechanisms linking ageing and HF are not clear. We aimed to identify the functional, morphological and transcriptomic changes that could be attributed to cardiac ageing in a model of slowly progressing HF in Tgαq*44 mice in reference to the cardiac ageing process in FVB mice. In FVB mice, ageing resulted in the impairment of diastolic cardiac function and in basal coronary flow (CF), perivascular and interstitial fibrosis without changes in the cardiac activity of angiotensin-converting enzyme (ACE) or aldosterone plasma concentration. In Tgαq*44 mice, HF progression was featured by the impairment of systolic and diastolic cardiac function and in basal CF that was associated with a distinct rearrangement of the capillary architecture, pronounced perivascular and interstitial fibrosis, progressive activation of cardiac ACE and systemic angiotensin-aldosterone-dependent pathways. Interestingly, cardiac ageing genes and processes were represented in Tgαq*44 mice not only in late but also in early phases of HF, as evidenced by cardiac transcriptome analysis. Thirty-four genes and 8 biological processes, identified as being ageing related, occurred early and persisted along HF progression in Tgαq*44 mice and were mostly associated with extracellular matrix remodelling and fibrosis compatible with perivascular fibrosis resulting in coronary microvascular dysfunction (CMD) in Tgαq*44 mice. In conclusion, accelerated and persistent cardiac ageing contributes to the pathophysiology of chronic HF in Tgαq*44 mice. In particular, prominent perivascular fibrosis of microcirculation resulting in CMD represents an accelerated cardiac ageing phenotype that requires targeted treatment in chronic HF.
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Aldosterona , Insuficiencia Cardíaca , Ratones , Animales , Ratones Transgénicos , Insuficiencia Cardíaca/metabolismo , Enfermedad Crónica , Ratones Endogámicos , Envejecimiento , Angiotensinas , FibrosisRESUMEN
BACKGROUND: The aim of this study was to investigate whether the 3 different substances that can decrease the development of atherosclerosis--nebivolol, AVE 0991 and doxycycline--could at the same time diminish the level of inflammatory indicators interleukin-6 (IL-6), interleukin-12 (IL-12), serum amyloid A (SAA), and monocyte chemotactic protein-1 (MCP-1). MATERIAL/METHODS: Forty 8-week-old female apoE-knockout mice on the C57BL/6J background were divided into 4 groups and put on chow diet for 4 months. Three experimental groups received the same diet as a control group, mixed with AVE 0991 at a dose 0.58 µmol per kg of body weight per day, nebivolol at a dose 2.0 µmol per kg of body weight per day, and doxycycline at a dose 1.5 mg per kg of body weight per day. At the age of 6 months, the mice were sacrificed. RESULTS: All inflammatory indicators (MCP-1, IL-6, IL-12 and SAA) were diminished by AVE 0991. There was also a tendency to lower MCP-1, IL-6, IL-12 and SAA levels by nebivolol and doxycycline; however, it did not reach statistical significance. CONCLUSIONS: Of the 3 presented substances, only AVE 0991 was able to diminish the rise of inflammatory markers. Therefore, drug manipulations in the renin-angiotensin-aldosterone axis seem to be the most promising in the future treatment of atherogenesis.
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Apolipoproteínas E/deficiencia , Aterosclerosis/sangre , Aterosclerosis/tratamiento farmacológico , Benzopiranos/uso terapéutico , Doxiciclina/uso terapéutico , Etanolaminas/uso terapéutico , Imidazoles/uso terapéutico , Mediadores de Inflamación/sangre , Animales , Apolipoproteínas E/metabolismo , Modelos Animales de Enfermedad , Femenino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , NebivololRESUMEN
Endothelial cells are characterized by intense metabolic activity and control of homeostasis. Exposure to benzo(a)pyrene (BaP) plays an important role in the etiology of atherosclerosis. The study aimed to determine the effect of arachidonic (ARA), and eicosapentaenoic acid (EPA) on pro-inflammatory gene and protein levels in human umbilical vein endothelial cells (HUVEC) exposed to BaP. Cyclooxygenase-2 (COX-2), aryl hydrocarbon receptor (AHR), and glutathione S transferase Mu1 (GSTM1) proteins expression were analyzed by Western blot. Prostaglandin synthase 2 (PTGS2), AHR, GSTM1, phospholipase A2 (PLA2G4A), cytochrome P450 CYP1A1, intercellular adhesion molecule-1 (ICAM-1), endothelial nitric-oxide synthase (NOS3), and vascular adhesion molecule-1 gene expression (VCAM-1) was analyzed in Real time-qPCR. Phospholipase A2 activity was measured using the ELISA technique, and CYP1A1 activity was analyzed in luminescence assay. The highest amount of COX-2, the most increased activity of CYP1A1 and cPLA2, and overexpression of GSTM1, CYP1A1, ICAM-1, and VCAM-1 gene was observed in HUVEC cells treated with BaP. After co-treatment with BaP and ARA or EPA, an increase of GSTM1 level was observed. Incubation of endothelial cells with ARA or EPA and BaP resulted in lower CYP1A1 and cPLA2 activities and lower expression of VCAM-1 and ICAM-1 genes. Significant overexpression of AHR, GSTM1, CYP1A1, PTGS2, PLA2G4A, and NOS3 genes was noted in cells treated with EPA and BaP. Our data suggest a beneficial effect of EPA and ARA on endothelial function. Thus, it justifies further research on the participation of fatty acids in the regulation of physiological and pathological processes in endothelial cells.
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Ácido Araquidónico/farmacología , Benzo(a)pireno/toxicidad , Supervivencia Celular/efectos de los fármacos , Ácido Eicosapentaenoico/farmacología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Supervivencia Celular/fisiología , Citocromo P-450 CYP1A1/metabolismo , Relación Dosis-Respuesta a Droga , HumanosRESUMEN
Introduction: A single measurement of any biomarker may not reflect its full biological meaning. The kinetics of fibrosis-linked microRNAs and their relationship with extracellular matrix (ECM) fibrosis in dilated cardiomyopathy (DCM) have not been explored. Material and methods: We evaluated 70 consecutive DCM patients (48 ±12.1 years, left ventricular ejection fraction 24.4 ±7.4%). All patients underwent right ventricular endomyocardial biopsy in order to quantify ECM fibrosis and measure collagen volume fraction (CVF). Circulating microRNAs (miR-21-5p, miR-29b, miR-30c-5p, and miR-133a-3p) were measured with quantitative polymerase chain reaction (PCR) at baseline and at 3 and 12 months. Results: Based on the biopsy results, two groups of patients were identified: with (n = 24, 34.3%) and without (n = 46, 65.7%) ECM fibrosis. Except for a single measurement of miR-29b at 3 months (DCM with fibrosis: 6.03 ±0.72 vs. DCM without fibrosis: 6.4 ±0.75 ΔCq; p < 0.05), baseline, 3- and 12-month kinetics of microRNAs did not differ between the two groups. Moreover, 12-month microRNA kinetics did not differ in patients with new-onset DCM (duration < 6 months; n = 35) and chronic DCM (> 6 months; n = 35). Only miR-29 at 3 months correlated with CVF (r = -0.31; p < 0.05), whereas other microRNAs did not correlate with CVF either at 3 or at 12 months. Conclusions: Regardless of ECM fibrosis status or duration of the disease, 12-month patterns of circulating microRNAs are similar in DCM. Correlations between microRNAs, measured at 3 and 12 months, are lower than expected. In this study, regardless of the time point, circulating microRNAs were not able to differentiate between DCM patients with versus without fibrosis.
RESUMEN
Previously, we showed that mouse delayed-type hypersensitivity (DTH) can be antigen-specifically downregulated by suppressor T cell-derived miRNA-150 carried by extracellular vesicles (EVs) that target antigen-presenting macrophages. However, the exact mechanism of the suppressive action of miRNA-150-targeted macrophages on effector T cells remained unclear, and our current studies aimed to investigate it. By employing the DTH mouse model, we showed that effector T cells were inhibited by macrophage-released EVs in a miRNA-150-dependent manner. This effect was enhanced by the pre-incubation of EVs with antigen-specific antibodies. Their specific binding to MHC class II-expressing EVs was proved in flow cytometry and ELISA-based experiments. Furthermore, by the use of nanoparticle tracking analysis and transmission electron microscopy, we found that the incubation of macrophage-released EVs with antigen-specific antibodies resulted in EVs' aggregation, which significantly enhanced their suppressive activity in vivo. Nowadays, it is increasingly evident that EVs play an exceptional role in intercellular communication and selective cargo transfer, and thus are considered promising candidates for therapeutic usage. However, EVs appear to be less effective than their parental cells. In this context, our current studies provide evidence that antigen-specific antibodies can be easily used for increasing EVs' biological activity, which has great therapeutic potential.