Publication: Enfoque multiómico para la determinación del riesgo cardiovascular en pacientes con neumonía adquirida en la comunidad
Authors
Carrillo Tornel, Salvador
item.page.secondaryauthor
Escuela Internacional de Doctorado
item.page.director
González-Conejero Hilla, Rocío ; Martínez Gómez, Constantion
Publisher
Universidad de Murcia
publication.page.editor
publication.page.department
DOI
item.page.type
info:eu-repo/semantics/doctoralThesis
Description
Abstract
La neumonía adquirida en la comunidad (NAC) es una infección frecuente y potencialmente mortal con una alta tasa de morbimortalidad. La NAC incrementa el riesgo de enfermedad cardiovascular (ECV) entre un 10-30%. En este escenario, se hace necesaria la búsqueda de biomarcadores capaces de predecir ECV asociadas a la infección.
En esta Tesis, hemos abordado la búsqueda de estos biomarcadores desde una estrategia multiómica, en pacientes con NAC con y sin eventos cardiovasculares mayores (MACE).
Objetivo 1- Determinar la prevalencia de CHIP y analizar su asociación con la aparición de MACE durante el seguimiento de pacientes con NAC. Objetivo 2- Caracterizar el perfil de miRNA plasmáticos de pacientes con NAC y evaluar su asociación con MACE. Objetivo 3- Identificar marcadores plasmáticos de MACE en NAC.
Materiales y Métodos. Se recogieron muestras de plasma y DNA de pacientes con NAC, así como de controles sanos. Para el estudio de CHIP, se realizó una NGS. Los miRNA plasmáticos se cuantificaron mediante miRNA-seq y validación por RT-qPCR. Empleamos técnicas de proteómica, validación mediante ELISA, y herramientas de machine learning (ML). Para los experimentos in vitro, se purificaron neutrófilos y monocitos de donantes sanos. En neutrófilos se evaluó la capacidad NETótica, producción de especies reactivas de oxígeno (ROS), capacidad fagocítica y migratoria. En los monocitos se evaluó la producción de factor tisular (FT).
Resultados. Capítulo 1: TET2 fue el gen más frecuentemente mutado en los pacientes con NAC. Los pacientes con mutaciones CHIP fueron predominantemente mujeres, de mayor edad, y con mayores tasas de mortalidad. La incidencia de CHIP fue mayor en los pacientes con MACE temprano (MACE-30d) respecto a los NO MACE-30d. Finalmente, la presencia de CHIP no tuvo un impacto en la supervivencia global (SG) a 4 años en los pacientes MACE-30d, pero sí en los NO MACE-30d.
Capítulo 2: Identificamos 18 miRNA diferencialmente expresados entre pacientes MACE-30d y NO MACE a 4 años (NO MACE-4a). Se validaron los niveles miR-206, miR-320a-3p y miR-423-5p, sobreexpresados en los MACE-30d. La integración de los niveles de miR-320a-3p junto con dislipemia y antecedentes de cardiopatía en un score de riesgo, ofreció un rendimiento predictivo de MACE a 30 días del 82,4%.
Capítulo 3: Identificamos 7 proteínas diferencialmente expresadas entre pacientes MACE-30d y NO MACE-4a, siendo el FGF-23 la más diferencialmente expresada. La validación por ELISA confirmó este dato. El modelo de ML (edad, hipertensión arterial y niveles de FGF-23) ofreció un poder predictivo de MACE temprano del 85%. El estudio funcional reveló que el FGF-23 en neutrófilos aumentó la NETosis, inhibió la producción de ROS y redujo la capacidad fagocítica y migratoria. En monocitos, aumentó el FT y activó diferentes rutas de señalización.
Conclusiones. Esta memoria aporta nuevos datos sobre diferentes enfoques en la búsqueda de biomarcadores moleculares predictores de MACE en pacientes con NAC.
1. La frecuencia de mutaciones relacionadas con CHIP se encuentra aumentada en pacientes con NAC y MACE-30d, aunque no afecta a la SG en este subgrupo de pacientes de alto riesgo. O bien el ECV tiene implicaciones en la SG de este subgrupo de pacientes, o bien el pequeño tamaño muestral nos impide sacar conclusiones inequívocas en este sentido.
2. La inclusión de los niveles de miR-320a-3p con variables clínicas en un score permite identificar con una precisión del 82,4% pacientes con NAC con riesgo cardiovascular a 30 días.
3. Los niveles plasmáticos de FGF-23 junto con variables clínicas en un modelo de ML ha mostrado un potencial valor pronóstico de MACE a 30 días en estos pacientes. El efecto funcional del FGF-23 en diferentes tipos celulares, podría justificar su papel como biomarcador de MACE en un escenario de trombo-inflamación.
Community-acquired pneumonia (CAP) is a common and potentially fatal infection with a high morbidity and mortality rate. CAP increases the risk of cardiovascular disease (CVD) by 10-30%. In this scenario, it is necessary to search for biomarkers capable of predicting CVD associated with infection. In this thesis, we have approached the search for these biomarkers using a multi-omic strategy in patients with CAP with and without major adverse cardiovascular events (MACE). Objective 1- To determine the prevalence of CHIP and analyze its association with the onset of MACE during follow-up of patients with CAP. Objective 2- To characterize the plasma miRNA profile of patients with CAP and evaluate its association with MACE. Objective 3- To identify plasma markers of MACE in CAP. Materials and Methods. Plasma and DNA samples were collected from patients with CAP, as well as from healthy controls. NGS was performed for the CHIP study. Plasma miRNAs were quantified using miRNA-seq and validated by RT-qPCR. We used proteomics techniques, ELISA validation, and machine learning (ML) tools. For in vitro experiments, neutrophils and monocytes were purified from healthy donors. In neutrophils, NETotic capacity, reactive oxygen species (ROS) production, phagocytic capacity, and migratory capacity were evaluated. In monocytes, tissue factor (TF) production was evaluated. Results. Chapter 1: TET2 was the most frequently mutated gene in patients with CAD. Patients with CHIP mutations were predominantly female, older, and had higher mortality rates. The incidence of CHIP was higher in patients with early MACE (MACE-30d) than in those without MACE-30d. Finally, the presence of CHIP had no impact on 4-year overall survival (OS) in MACE-30d patients, but it did in those without MACE-30d. Chapter 2: We identified 18 differentially expressed miRNAs between MACE-30d and NO MACE patients at 4 years (NO MACE-4a). The levels of miR-206, miR-320a-3p, and miR-423-5p, which were overexpressed in MACE-30d patients, were validated. The integration of miR-320a-3p levels together with dyslipidemia and a history of heart disease in a risk score offered a predictive performance of MACE at 30 days of 82.4%. Chapter 3: We identified seven proteins differentially expressed between MACE-30d and NO MACE-4a patients, with FGF-23 being the most differentially expressed. Validation by ELISA confirmed this finding. The ML model (age, hypertension, and FGF-23 levels) offered an 85% predictive power for early MACE. The functional study revealed that FGF-23 in neutrophils increased NETosis, inhibited ROS production, and reduced phagocytic and migratory capacity. In monocytes, it increased FT and activated different signaling pathways. Conclusions. This report provides new data on different approaches in the search for molecular biomarkers predictive of MACE in patients with NSCLC. 1. The frequency of CHIP-related mutations is increased in patients with NSCLC and MACE-30d, although it does not affect OS in this subgroup of high-risk patients. Either CVE has implications for OS in this subgroup of patients, or the small sample size prevents us from drawing unequivocal conclusions in this regard. 2. The inclusion of miR-320a-3p levels with clinical variables in a score allows the identification of patients with NSCLC at 30-day cardiovascular risk with 82.4% accuracy. 3. Plasma FGF-23 levels together with clinical variables in an ML model have shown a potential prognostic value for 30-day MACE in these patients. The functional effect of FGF-23 in different cell types could justify its role as a biomarker of MACE in a thrombo-inflammatory scenario.
Community-acquired pneumonia (CAP) is a common and potentially fatal infection with a high morbidity and mortality rate. CAP increases the risk of cardiovascular disease (CVD) by 10-30%. In this scenario, it is necessary to search for biomarkers capable of predicting CVD associated with infection. In this thesis, we have approached the search for these biomarkers using a multi-omic strategy in patients with CAP with and without major adverse cardiovascular events (MACE). Objective 1- To determine the prevalence of CHIP and analyze its association with the onset of MACE during follow-up of patients with CAP. Objective 2- To characterize the plasma miRNA profile of patients with CAP and evaluate its association with MACE. Objective 3- To identify plasma markers of MACE in CAP. Materials and Methods. Plasma and DNA samples were collected from patients with CAP, as well as from healthy controls. NGS was performed for the CHIP study. Plasma miRNAs were quantified using miRNA-seq and validated by RT-qPCR. We used proteomics techniques, ELISA validation, and machine learning (ML) tools. For in vitro experiments, neutrophils and monocytes were purified from healthy donors. In neutrophils, NETotic capacity, reactive oxygen species (ROS) production, phagocytic capacity, and migratory capacity were evaluated. In monocytes, tissue factor (TF) production was evaluated. Results. Chapter 1: TET2 was the most frequently mutated gene in patients with CAD. Patients with CHIP mutations were predominantly female, older, and had higher mortality rates. The incidence of CHIP was higher in patients with early MACE (MACE-30d) than in those without MACE-30d. Finally, the presence of CHIP had no impact on 4-year overall survival (OS) in MACE-30d patients, but it did in those without MACE-30d. Chapter 2: We identified 18 differentially expressed miRNAs between MACE-30d and NO MACE patients at 4 years (NO MACE-4a). The levels of miR-206, miR-320a-3p, and miR-423-5p, which were overexpressed in MACE-30d patients, were validated. The integration of miR-320a-3p levels together with dyslipidemia and a history of heart disease in a risk score offered a predictive performance of MACE at 30 days of 82.4%. Chapter 3: We identified seven proteins differentially expressed between MACE-30d and NO MACE-4a patients, with FGF-23 being the most differentially expressed. Validation by ELISA confirmed this finding. The ML model (age, hypertension, and FGF-23 levels) offered an 85% predictive power for early MACE. The functional study revealed that FGF-23 in neutrophils increased NETosis, inhibited ROS production, and reduced phagocytic and migratory capacity. In monocytes, it increased FT and activated different signaling pathways. Conclusions. This report provides new data on different approaches in the search for molecular biomarkers predictive of MACE in patients with NSCLC. 1. The frequency of CHIP-related mutations is increased in patients with NSCLC and MACE-30d, although it does not affect OS in this subgroup of high-risk patients. Either CVE has implications for OS in this subgroup of patients, or the small sample size prevents us from drawing unequivocal conclusions in this regard. 2. The inclusion of miR-320a-3p levels with clinical variables in a score allows the identification of patients with NSCLC at 30-day cardiovascular risk with 82.4% accuracy. 3. Plasma FGF-23 levels together with clinical variables in an ML model have shown a potential prognostic value for 30-day MACE in these patients. The functional effect of FGF-23 in different cell types could justify its role as a biomarker of MACE in a thrombo-inflammatory scenario.
publication.page.subject
Citation
item.page.embargo
Collections
Ir a Estadísticas
Este ítem está sujeto a una licencia Creative Commons. http://creativecommons.org/licenses/by-nc-nd/4.0/