Diabetes mellitus y hábito tabáquico como factores pronósticos del tratamiento endodóntico: revisión sistemática y metaanálisis

UNIVERSIDAD DE SEVILLA FACULTAD DE ODONTOLOGÍA DOCTORADO DE CIENCIAS DE LA SALUD DIABETES MELLITUS Y HÁBITO TABÁQUICO COMO FACTORES PRONÓSTICOS DEL TRATAMIENTO ENDODÓNTICO: REVISIÓN SISTEMÁTICA Y METAANÁLISIS DIABETES MELLITUS AND SMOKING AS PROGNOSTIC FACTORS OF ENDODONTIC TREATMENT: SYSTEMATIC REVIEW AND METAANALYSIS Tesis Doctoral DANIEL CABANILLAS BALSERA Sevilla, 2021 Sevilla, julio de 2021 D. JUAN JOSÉ SEGURA EGEA, Catedrático de Patología y Terapéutica Dentales del Departamento de Estomatología de la Facultad de Odontología y Director del Máster de Endodoncia Clínica de la Universidad de Sevilla. D. EUGENIO VELASCO ORTEGA, Profesor Titular de Odontología Integrada de Adultos y Gerodontología del Departamento de Estomatología de la Facultad de Odontología y Director del Máster de Implantología Oral de la Universidad de Sevilla. CERTIFICAN: Que D. DANIEL CABANILLAS BALSEA, Graduado en Odontología por la Universidad de Sevilla e inscrito en el programa de Doctorado de Ciencias de la Salud de la Universidad de Sevilla, ha realizado bajo su tutela y dirección el trabajo titulado Diabetes mellitus y hábito tabáquico como factores pronósticos del tratamiento endodóntico: revisión sistemática y metaanálisis / Diabetes mellitus and smoking as prognostic factors of endodontic treatment: systematic review and metaanalysis, que consideramos satisfactorio para optar al título de Doctor en Odontología. Prof. Segura Egea Prof. Velasco Ortega Dpto. de Estomatología FACULTAD DE ODONTOLOGÍA AGRADECIMIENTOS A los Profesores JUAN JOSÉ SEGURA EGEA y EUGENIO VELASCO ORTEGA de la Facultad de Odontología de la Universidad de Sevilla, cuyos conocimientos y estímulos han sido muy importantes en la planificación y realización del presente estudio de investigación doctoral. A los Profesores e Investigadores compañeros del departamento de Estomatología de la Universidad de Sevilla, con los que durante los últimos años he participado en diversas actividades universitarias y científicas relacionadas con el objeto de la presente tesis doctoral. Al Profesor Luciano Tavares Angelo Cintra y a su equipo, por su apoyo y supervisión durante mi estancia de investigación online en el Departamento de Odontología Preventiva y Restauradora de la Facultad de Odontología de la Universidad Estadual de São Paulo (Unesp) de Brasil. A la Universidad de Sevilla y al Ministerio de Universidades por la financiación para la Formación de Profesorado Universitario (FPU) y para el proyecto de investigación abordado en esta tesis doctoral. ÍNDICE INTRODUCCIÓN ___________________________________________ 8 Periodontitis apical ______________________________________ 9 Tratamiento de conductos. Pronóstico en endodoncia. __________ 12 Medicina endodóntica ___________________________________ 14 Posibles vías de conexión entre la periodontitis apical y las enfermedades sistémicas ________________________________ 15 Patologías sistémicas asociadas a la patología endodóntica _____ 20 OBJETIVOS ______________________________________________ 29 OBJECTIVES _____________________________________________ 32 MATERIAL Y MÉTODOS ____________________________________ 35 REVISIÓN SISTEMÁTICA _________________________________ 36 Pregunta PICO _________________________________________ 38 Criterios de inclusión y exclusión ___________________________ 38 Estrategia de búsqueda __________________________________ 38 Selección de los estudios y extracción de datos _______________ 39 Calidad de la evidencia __________________________________ 40 METAANÁLISIS _________________________________________ 45 Forest plot ____________________________________________ 46 Variables dicotómicas y variables continuas __________________ 47 Modelos de efectos fijos y modelos de efectos aleatorios ________ 48 Heterogeneidad ________________________________________ 49 Sesgo de publicación ____________________________________ 51 5 PRIMERA PARTE: DIABETES MELLITUS Y PRONÓSTICO DEL TRATAMIENTO DE CONDUCTOS CAPÍTULO I Association between diabetes and the prevalence of radiolucent periapical lesions in root-filled teeth: systematic review and metaanalysis _________________________________________________ 54 ABSTRACT _____________________________________________ 55 1. INTRODUCTION _____________________________________ 57 2. AIM OF THE STUDY __________________________________ 58 3. MATERIALS AND METHODS ___________________________ 59 4. RESULTS ___________________________________________ 62 5. DISCUSSION ________________________________________ 69 6. CONCLUSION _______________________________________ 71 REFERENCES __________________________________________ 72 CAPÍTULO II Association between diabetes and nonretention of root filled teeth: a systematic review and meta ‐ analysis _________________________ 79 ABSTRACT _____________________________________________ 80 1. INTRODUCTION _____________________________________ 81 2. REVIEW ____________________________________________ 83 3. DISCUSSION ________________________________________ 91 4. CONCLUSIONS ______________________________________ 94 REFERENCES __________________________________________ 95 6 CAPÍTULO III Endodontics and diabetes: association versus causation _______ 104 ABSTRACT ____________________________________________ 105 1. INTRODUCTION ____________________________________ 106 2. METHODOLOGICAL APPROACH _______________________ 110 3. INTERPRETATION OF THE STUDIES ON ENDODONTIC MEDICINE: CAUSATION OR ASSOCIATION ______________ 110 4. DIABETES MELLITUS – ENDODONTIC DISEASE: CAUSATION OR ASSOCIATION? __________________________________ 114 5. CONCLUSIONS _____________________________________ 126 REFERENCES _________________________________________ 127 DISCUSIÓN Diabetes mellitus y pronóstico del tratamiento de conductos ____ 138 SEGUNDA PARTE: HÁBITO TABÁQUICO Y PRONÓSTICO DEL TRATAMIENTO DE CONDUCTOS CAPÍTULO IV Smoking and Radiolucent Periapical Lesions in Root Filled Teeth: Systematic Review and Meta-Analysis _______________________ 144 ABSTRACT ____________________________________________ 145 1. INTRODUCTION ____________________________________ 147 2. METHODS _________________________________________ 148 3. RESULTS __________________________________________ 153 4. DISCUSSION _______________________________________ 162 5. CONCLUSIONS ______________________________________ 164 7 REFERENCES _________________________________________ 165 CAPÍTULO V Cigarette Smoking and Root Filled Teeth Extraction: Systematic Review and Meta-Analysis _________________________________ 174 ABSTRACT ____________________________________________ 175 1. INTRODUCTION ____________________________________ 176 2. MATERIALS AND METHODS __________________________ 177 3. RESULTS __________________________________________ 181 4. DISCUSSION _______________________________________ 189 5. CONCLUSIONS _____________________________________ 192 REFERENCES _________________________________________ 193 DISCUSIÓN Hábito tabáquico y pronóstico del tratamiento de conductos ____ 201 CONCLUSIONES _________________________________________ 205 CONCLUSIONS __________________________________________ 207 REFERENCIAS___________________________________________ 209 INTRODUCCIÓN Introducción 9 Las implicaciones de las enfermedades sistémicas en la salud o patología de los tejidos periapicales es un tema de gran interés en la investigación en los últimos años. En este sentido, surge el concepto de “Medicina Endodóntica” (Segura-Egea et al. 2015), que se desarrolla para buscar y analizar posibles implicaciones de enfermedades y/o hábitos sistémicos en la pulpa y los tejidos periapicales, así como de las consecuencias sistémicas que puedan tener la pulpitis y la periodontitis apical (Segura-Egea et al. 2015, Cintra et al. 2018). PERIODONTITIS APICAL La periodontitis apical es un proceso de inflamación y destrucción de los tejidos perirradiculares causado por una infección microbiana del sistema de conductos radiculares del diente afectado (Nair 2004, Al-Nazhan et al. 2017). Las bacterias causantes de la infección endodóntica presentes en el espacio pulpar, junto con las toxinas, antígenos y moléculas con efectos biológicos liberadas por las mismas, avanzan a través del foramen apical hacia los tejidos periapicales, desencadenando una defensa del hospedador (Figura 1 y Figura 2). Esto va a generar una reacción inmuno-inflamatoria local, junto con un daño tisular en la estructura ósea periapical, resultando en una reabsorción identificada radiográficamente como una lesión radiolúcida periapical (LRP) (Nair et al. 1999, Estrela et al. 2008, Van der Veken et al. 2017). Introducción 16 a) Propagación de bacterias endodónticas Pueden existir dos formas de propagación de las bacterias de la inflamación periapical: a.1) por contigüidad, mediante la propagación y extensión de la infección localizada en el periápice a los tejidos adyacentes, provocando celulitis cervicofacial, y pudiendo comprometer sistémicamente al paciente mediante un cuadro febril e inflamatorio generalizado si no se instaura el tratamiento adecuado; y a.2.) por bacteriemia, mediante el paso de bacterias endodónticas a la sangre, gracias a la existencia un tejido periapical inflamatorio granulomatoso muy rico en capilares, lo cual posibilita la invasión del torrente sanguíneo por parte de las bacterias presentes en el periápice, así como su propagación metastásica por vía hematógena (Marton & Bergenholtz 2005, Gomes et al. 2013). b) Paso de moléculas liberadas en el tejido periapical a la sangre Las bacterias causantes de la infección endodóntica también liberan moléculas con efectos biológicos. Asimismo, las citoquinas liberadas en la región periapical inflamada pueden pasar al torrente sanguíneo, produciendo efectos en zonas alejadas del organismo (Marton 2004). Los patrones moleculares asociados a patógenos LPS y LTA (ácido lipoteicoico, antígeno de la pared celular de bacterias gram positivas), potentes estimulantes de la inmunidad innata, interaccionan con sus receptores LPS/TLR4 y LTA/TLR2 activando el factor transcripcional NF-, potenciando la síntesis de las citoquinas pro-inflamatorias IL-1, IL-6, IL-8, IL-12 y TNF (Ao et al. 2015). Esto conllevaría a una estimulación de la síntesis y liberación en el hígado de los reactantes de la fase aguda, como la proteína C reactiva (PCR), y el fibrinógeno, que favorecen la trombogénesis y podrían provocar lesiones en órganos alejados de la lesión endodóntica (Garrido et al. 2019, Georgiou et al. 2019). Además, las citoquinas pro-inflamatorias y los reactantes de la fase aguda, junto con los tromboxanos, las prostaglandinas (PGE2) y los leucotrienos que se acumulan en la región periapical inflamada durante el proceso de reabsorción ósea (Stashenko et al. 1998), pueden pasar al torrente sanguíneo Introducción 17 (Li et al. 2000) e inducir un estado sistémico pro-inflamatorio crónico (Caplan et al. 2006, Gomes et al. 2013). Por otro lado, el mimetismo molecular por el que secuencias de péptidos bacterianos son confundidas con tejidos humanos, provocaría una reacción autoimmune por reactividad cruzada, pudiendo constituir otra vía de conexión entre la inflamación periapical y la patología sistémica (Marton 2004). c) Presencia de factores de riesgo comunes a la patología endodóntica y sistémica y el polimorfismo genético Determinados factores de riesgo, extrínsecos e intrínsecos, como por ejemplo la edad, el tabaco, el estilo de vida o el estado socioeconómico pueden estar implicados con la etiología tanto de la patología pulpo-periapical, como de numerosas enfermedades sistémicas. Estos factores comunes deben considerarse como posibles variables de confusión en estudios epidemiológicos que analicen la asociación entre ambas patologías. Ejemplo de ello es el tabaco, factor etiológico clave en la patología cardiovascular, y a la vez, asociado con una mayor prevalencia de periodontitis apical (Segura-Egea et al. 2015, Cintra et al. 2018). Por otro lado, determinados polimorfismos genéticos, existencia de más de un alelo para el locus de un gen, pueden predisponer a patologías tanto sistémicas como endodónticas, como por ejemplo, las variantes polimórficas en genes relacionados con interleuquinas, enzimas, factores transcripcionales o moléculas implicadas en la respuesta inmune y reparativa (Morsani et al. 2011). Así tenemos el polimorfismo de un solo nucleótido en el gen KCNK3, que predispone a la hipertensión y, a la vez, se asocia a mayor prevalencia de periodontitis apical (Messing et al. 2019), o el polimorfismo en el gen IL-1β, que se asocia a mayor prevalencia de periodontitis apical (Salles et al. 2018) y a mayor riesgo de infarto de miocardio y de ictus (Bis et al. 2008). Introducción 18 2. Influencia de los factores sistémicos en la patología periapical El proceso reparativo periapical tras la realización del tratamiento de conductos también puede verse afectado por determinadas condiciones sistémicas que afecten a la respuesta inmune innata, alterando la activación macrofágica y la función fagocítica de los neutrófilos, estimulando un estado proinflamatorio que altera la proliferación celular, y retrasando o impidiendo la reparación tisular periapical pos-endodóntica. Por tanto, determinadas patologías como son la diabetes mellitus, la arteriosclerosis, la hipertensión, la osteoporosis, el hábito tabáquico, problemas hereditarios de la coagulación, o algunos trastornos digestivos pueden disminuir la respuesta reparativa pulpar y periapical, lo que puede manifestarse como un incremento de la probabilidad de sufrir periodontitis apical y tratamiento de conductos, así como un aumento de la prevalencia de periodontitis apical persistente tras el tratamiento endodóntico (Ng et al. 2011a, Segura-Egea et al. 2015). Otras patologías sistémicas producen una fuerte reacción inflamatoria que activa el factor NF-κB en los macrófagos e incrementa el estrés oxidativo, produciendo una alteración del recambio óseo, con incremento de la actividad osteoclástica y la reabsorción ósea, y disfunción de los osteoblastos y fibroblastos con alteración de la síntesis de colágeno. Estas enfermedades, que presentan altos niveles serológicos de PCR, colagenasas, citoquinas proinflamatorias (IL-1b, IL-6, IL-8, IL-10, TNF-) e incremento de radicales libres, suelen generar lesiones periapicales de mayor progresión, a la vez que el proceso de cicatrización periapical se enlentece o detiene (Taylor et al. 2013, Peddis et al. 2019). Por último, alteraciones en la vascularización con reducción del flujo sanguíneo y disminución del aporte de nutrientes y oxígeno a los tejidos, como las descritas en los fumadores crónicos, también pueden retrasar o detener la curación de la periodontitis apical, llevando incluso al fracaso del tratamiento endodóntico (Kinane & Chestnutt 2000, Duncan & Pitt Ford 2006, Segura-Egea et al. 2015). Introducción 19 Figura 3. Los patrones moleculares asociados a patógenos (PAMP), como el lipopolisacárido capsular (LPS) de las bacterias gram-negativas o el ácido lipoteicoico (LTA) de la pared celular de las bacterias gram-positivas, junto con moléculas resultantes de la necrosis celular, denominadas patrones moleculares asociados a daño tisular (DAMP), como la proteína de alta movilidad del grupo 1 (HMGB1), invaden los tejidos periapicales y estimulan los receptores TLR2 y TLR4 de las células inmunes, provocando la reacción inmune e inflamatoria característica de la periodontitis apical. Las posibles interrelaciones de la patología endodóntica y sistémica pueden producirse a través de tres vías: la propagación de las bacterias endodónticas, el paso a la sangre de moléculas que se están liberando en el tejido periapical y la presencia de factores de riesgo comunes a la patología endodóntica y sistémica o el polimorfismo genético (Segura-Egea et al. 2021). Introducción 20 PATOLOGÍAS SISTÉMICAS ASOCIADAS A LA PATOLOGÍA ENDODÓNTICA Las patologías sistémicas cuya posible relación con la periodontitis apical se han investigado son numerosas (Segura-Egea et al. 2015) (Figura 4). En la mayoría de los casos se ha encontrado asociación significativa, lo que no implica que exista relación causal (Segura-Egea et al. 2019). Figura 4. Patologías sistémicas que han sido asociadas a la patología endodóntica y al tratamiento de conductos (Segura-Egea et al. 2021). Diabetes mellitus La diabetes mellitus, enfermedad metabólica caracterizada por niveles elevados crónicos de glucosa en sangre como resultado de la incapacidad de las células beta del páncreas para producir la insulina adecuada o la utilización ineficaz de la insulina por las células del cuerpo (Alberti & Zimmet 1998), tiene una prevalencia estimada del 9,3% (463 millones) de la población global mundial, y se prevé su aumento al 10,2% (578 millones) para el 2030 y al 10,9% (700 millones) para 2045. Además, su prevalencia es mayor en zonas urbanas (10,8%) que en zonas rurales (7,2%) y en zonas de ingresos altos (10,4%) que en países de ingresos bajos (4,0%) (Saeedi et al. 2019) (Figura 5). Estos datos convierten a la diabetes mellitus en uno de los desafíos más importantes de la salud pública del siglo XXI (Zimmet et al. 2016). Introducción 21 Figura 5. Mapa de la prevalencia mundial de diabetes mellitus (Our World in Data 2017). Desde hace décadas, la diabetes mellitus se considera un factor de riesgo para determinadas patologías orales, como los problemas periodontales (Löe 1993) o el desarrollo de infecciones periapicales de mayor tamaño en pacientes con diabetes mellitus mal controlados (Bender et al. 1963). Desde entonces, se han llevado a cabo numerosas investigaciones buscando asociaciones entre la patología periapical y la diabetes mellitus (Riós-Osorio et al. 2020), encontrando entre sus resultados un aumento de la prevalencia de periodontitis apical en pacientes diabéticos comparado con sanos (Segura-Egea et al. 2005, López- López et al. 2011), un aumento de la frecuencia de lesiones perirradiculares en pacientes diabéticos de larga duración frente a corta duración (Mesgarani et al. 2014) o una asociación entre la infección periapical y el grado de control metabólico de la diabetes mellitus (Sánchez-Domínguez et al. 2015, Smadi 2017). Además, un estudio reciente indica que los pacientes con diabetes mellitus tipo 2 tienen cambios de dimensión fractal, una medida de la complejidad estructural de la región ósea evaluada, significativamente menores en el área de Introducción 22 la lesión apical de dientes tras el tratamiento de conductos en comparación con los individuos sanos, indicando una menor complejidad trabecular de forma cuantitativa, así como una menor curación de las lesiones periapicales tras el tratamiento endodóntico (Uğur Aydın et al. 2021). En los pacientes con diabetes mellitus, el deterioro de la inmunidad innata, la hiperglucemia, y el incremento del nivel sérico de productos avanzados de la glicosilación (AGEs), podrían ser los mecanismos biológicos implicados en la mayor prevalencia y el retraso de la curación de la periodontitis apical (Segura- Egea et al. 2012, 2015) (Figura 6). Además, el polimorfismo genético también podría jugar un papel importante, habiéndose encontrado que el polimorfismo de los genes RANK y RANKL se asocia tanto a mayor prevalencia de periodontitis apical persistente (Petean et al. 2019) como a mayor prevalencia de diabetes tipo 2 (Duan et al. 2016). Por otro lado, el posible efecto de la inflamación periapical sobre el control de la diabetes podría explicarse por el incremento de interleuquinas proinflamatorias que la periodontitis apical crónica induciría en el paciente diabético, contribuyendo al desarrollo de resistencia periférica a la insulina y al descontrol metabólico de la diabetes (Segura-Egea et al. 2019) (Figura 6). Figura 6. Posibles mecanismos biológicos que conectan la periodontitis apical con la diabetes tipo 2 (Segura-Egea et al. 2019). Introducción 23 Hábito tabáquico El hábito tabáquico, factor de riesgo ampliamente reconocido de influencia negativa para la salud general y de morbilidad y mortalidad prematuras, presenta una prevalencia de fumadores diarios en mayores de 15 años de entre un 25.0- 31,1% en hombres y del 5,4-6,2% en mujeres, lo que supone una tasa de número de fumadores mundial de alrededor de mil millones de personas (Ng et al. 2014, Reitsma et al. 2017) (Figura 7). Sin embargo, la distribución entre los diferentes países es muy heterogénea, lo que hace encontrar países con tasas del 40% de fumadores, como ocurre en Chile o Grecia (World Health Organization 2020). Además, aunque se espera una tendencia de disminución en la prevalencia del hábito tabáquico, en la actualidad supone una importante amenaza para la salud pública mundial, y en especial en los países de ingresos medios y bajo, puesto que el crecimiento y envejecimiento de la población general se traduce en un aumento de las complicaciones, enfermedades y número total de muertes relacionados con el tabaco (Ritchie & Roser 2013, Chung-Hall et al. 2019). Figura 7. Mapa de la prevalencia mundial de fumadores (Our World in Data 2016). Introducción 24 El hábito tabáquico supone un riesgo bien definido para la salud sistémica y oral, incrementando el riesgo de caries (Fure 2004), de enfermedad periodontal (Walter et al. 2012a) y de patología endodóntica, encontrándose un aumento de la prevalencia de lesiones periapicales (Oginni et al. 2015) y de tratamientos de conductos (Segura-Egea et al. 2008, 2011). Los posibles mecanismos por los cuales el tabaco puede alterar la respuesta inmune y reparadora, influyendo en la inflamación periapical, podrían ser diversos (Segura-Egea et al. 2015). El tabaco induce una fuerte respuesta inflamatoria crónica sistémica, con aumento de IL-1β, TNF-α o proteína C- reactiva, que junto con una disfunción de leucocitos polimorfonucleares, macrófagos y linfocitos T y una reducción de los niveles de anticuerpos (IgA, IgM, IgG), va a generar una alteración de la respuesta inmune frente a la infección pulpo-periapical (Tappia et al. 1995, Fröhlich et al. 2003) (Figura 8). Por otro lado, el tabaco también provoca alteraciones morfológicas y funcionales en la microcirculación, con disminución del suministro de oxígeno y daño a células endoteliales por aumento de radicales libres (Ijzerman et al. 2003). Esto, junto con una disfunción de los fibroblastos y disminución de la síntesis de colágeno, se traduce en un aumento de la destrucción ósea y una alteración de la reparación tisular periapical (Wong et al. 2004, Balto et al. 2019) (Figura 8). Introducción 25 Figura 8. Posibles vías de conexión entre el tabaquismo y la periodontitis apical (Segura-Egea et al. 2015). Otras patologías Las enfermedades cardiovasculares (ECV), trastornos relativos al corazón y los vasos sanguíneos que tienen como factor etiológico común la aterosclerosis, suponen la principal causa de muerte mundial en la actualidad (Margaix-Muñoz et al. 2008). La cardiopatía coronaria o isquémica (producida por una oclusión parcial o total de las arterias coronarias, que resulta en un aporte sanguíneo insuficiente al miocardio, apareciendo hipoxia y acúmulo de metabolitos en una región del músculo cardíaco, y manifestándose como un angor pectoris, un infarto agudo de miocardio o una muerte súbita) y la hipertensión arterial (trastorno consistente en una elevación de los niveles de presión que el corazón ejerce sobre las arterias de forma continua o sostenida) son las ECVs que presentan mayor prevalencia. Pioneros estudios epidemiológicos publicados hace más de 30 años relacionan las ECVs y las infecciones orales como la periodontitis apical (Mattila et al. 1989, 1995, Mattila 1993). Desde entonces se ha descrito mayor prevalencia de infarto de miocardio en pacientes con infecciones periodontales OBJECTIVES Objectives 33 The objective of this doctoral thesis is to investigate the possible association between the systemic state of the patient with diabetes mellitus or smoking, and the prognosis of root canal treatment by conducting systematic reviews and metaanalyses for the following specific objectives: 1. Determine the possible association between diabetes mellitus and endodontic treatment failure by evaluating: - The prevalence of radiolucent periapical lesions in root-filled teeth in diabetic patients and in healthy control subjects. In: CHAPTER I. Association between diabetes and the prevalence of radiolucent periapical lesions in root-filled teeth: systematic review and meta-analysis. - The prevalence of extracted root-filled teeth in diabetic patients and in healthy control subjects. In: CHAPTER II. Association between diabetes and nonretention of root filled teeth: a systematic review and meta ‐ analysis. - The type of association, causal or not, existing between endodontic disease and diabetes mellitus, according to the Bradford-Hill criteria. In: CHAPTER III. Endodontics and diabetes: association versus causation. 2. Determine the possible association between smoking and endodontic treatment failure by evaluating: - The prevalence of radiolucent periapical lesions in root-filled teeth in smoking patients and in non-smoking control subjects. In: CHAPTER IV. Smoking and Radiolucent Periapical Lesions in Root Filled Teeth: Systematic Review and Meta-Analysis. Objectives 34 - The prevalence of extracted root-filled teeth in smoking patients and non-smoking control subjects. In: CHAPTER V. Cigarette Smoking and Root Filled Teeth Extraction: Systematic Review and Meta-Analysis. MATERIAL Y MÉTODOS Material y métodos 36 La metodología empleada para llevar a cabo los objetivos de la presente tesis doctoral incluye la realización de una revisión sistemática y metaanálisis para cada una de las posibles variables de asociación, permitiendo la combinación y el análisis de datos de diferentes estudios realizados sobre temas de investigación similares. REVISIÓN SISTEMÁTICA Una revisión sistemática es una revisión de la literatura planificada y ejecutada de manera meticulosa, incluyendo un plan detallado y completo, y una estrategia de búsqueda bien definida, con el objetivo de reducir el sesgo al identificar, evaluar y sintetizar todos los estudios relevantes sobre un tema en particular (Uman 2011, Nagendrababu et al. 2020b). Habitualmente, las revisiones sistemáticas incluyen un componente de metaanálisis que implica el uso de técnicas estadísticas para analizar y sintetizar los datos de varios estudios, combinándolos en una sola estimación cuantitativa o tamaño del efecto global (Uman 2011). Tradicionalmente, la pirámide de la evidencia establece una jerarquía otorgando un nivel estático para los distintos diseños de estudios, posicionando aquellos con metodología más débil en la parte inferior (informes y series de casos), seguidos de estudios de casos y controles y estudios de cohortes en el centro, posteriormente ensayos controlados aleatorios y, en la parte superior, revisiones sistemáticas y metaanálisis (Figura 10) (Murad et al. 2016). Sin embargo, esta pirámide de la evidencia se cuestiona al considerar que la certeza de la evidencia se basa en la combinación del diseño del estudio junto con otros factores, como la imprecisión, incoherencia o resultados indirectos. De esta manera, en el modelo actual, las revisiones sistemáticas y metaanálisis dejan de ocupar la parte superior de la pirámide, pasando a considerarse los mismos como una “lupa” a través de la cual se ve y se aplica un análisis de la Material y métodos 37 evidencia, siendo la certeza de la evidencia variable y dinámica para distintos estudios con un mismo diseño (Figura 10) (Murad et al. 2016). Figura 10. Nueva pirámide de la evidencia propuesta por Murad et al. (Murad et al. 2016). Material y métodos 38 Una revisión sistemática debe considerar el análisis de los siguientes factores (Uman 2011, Ahn & Kang 2018, Higgins et al. 2019, Nagendrababu et al. 2020b): Pregunta PICO El primer paso de una revisión sistemática es definir la pregunta de revisión con metodología PICO: population, intervention, comparison, outcome. La formulación de esta pregunta permite establecer los diferentes componentes claves que se van a tener en cuenta antes de comenzar la revisión, definiendo la población (P): problema o pacientes con una condición en particular; intervención (I): actuación o tratamiento que queremos analizar; comparación (C): intervención alternativa con la cual comparar; y resultado (O, de outcome): desenlace esperado de la intervención. Criterios de inclusión y exclusión Los investigadores deben definir a priori las características de los estudios que se van a incluir, precisando el tipo de estudios (ensayos controlados aleatorios, estudios de cohortes, de casos y controles…), el período de investigación, el tamaño muestral mínimo de cada grupo, el rango de edad de la población, las condiciones, resultados y tipos de intervenciones y grupos de control, o las restricciones de idioma o acceso de los trabajos (Uman 2011). Asimismo, también es necesario determinar los criterios o características de los estudios que conllevarán a su exclusión de la revisión sistemática. Estrategia de búsqueda Esta etapa es esencial para garantizar una búsqueda que incluya todos los estudios que reporten información acerca de la pregunta planteada. Material y métodos 39 Para ello, es conveniente elaborar una lista completa de palabras clave o términos MeSH relacionados con cada componente de la pregunta PICO, desarrollando una estrategia de búsqueda óptima y equilibrada entre sensibilidad (identificar una alta proporción de estudios relevantes) y especificidad (identificar una baja proporción de estudios irrelevantes) (Ahn & Kang 2018). Las búsquedas deben realizarse incluyendo varias bases de datos electrónicas relevantes, así como, búsquedas manuales en las principales revistas de investigación y referencias de artículos y revisiones destacados sobre el tema abordado. Además, también puede considerarse la denominada literatura gris, incluyendo búsquedas en actas de congresos, tesis doctorales, artículos no publicados o informes o registros de investigaciones (Kang 2016). Selección de los estudios y extracción de datos Una vez identificados todos los estudios de las búsquedas, los investigadores deben eliminar los artículos duplicados y seleccionar los estudios que cumplen con los criterios de inclusión / exclusión según su título y resumen. Posteriormente, se lleva a cabo la selección final de los estudios mediante la revisión del texto completo de los mismos. Para mantener la objetividad y eliminar posibles sesgos en la selección de los estudios, este proceso debe realizarse de forma independiente por al menos dos investigadores, recurriendo a la intervención de un revisor adicional en caso de inconsistencia entre las opiniones. A continuación, se lleva a cabo la extracción de datos de los diferentes estudios seleccionados, siendo interesante la sintetización de la información aportada en los mismos en una tabla resumen. Esta tabla puede incluir los autores, año de publicación, diseño del estudio, número de participantes, variables de evaluación o principales resultados (Ahn & Kang 2018). Material y métodos 40 Calidad de la evidencia Finalmente, la evaluación de la calidad o certeza de la evidencia permite determinar la solidez de las conclusiones o recomendaciones de una revisión sistemática y/o metaanálisis, basada en la evaluación de los artículos que se incluyen. Existen multitud de métodos que describen un procedimiento para evaluar la calidad de la evidencia. Entre ellos encontramos los niveles de evidencia propuestos por el Centro de Medicina Basada en Evidencia (CEBM) de Oxford. Se trata de una escala o jerarquía para estratificar la evidencia de la más fuerte a la más débil sobre la base de la susceptibilidad al sesgo y la calidad del diseño del estudio, esto es, dependiendo de las características metodológicas propias de la investigación. De esta manera, los ensayos clínicos aleatorizados reciben el nivel más alto porque están diseñados para ser imparciales y tienen menos riesgo de errores sistemáticos, mientras que una serie de casos o la opinión de un experto a menudo está sesgada por la experiencia u opiniones del autor y no hay control de los factores de confusión (Tabla 1). Además, en base a estos niveles de evidencia se emite un grado de recomendación o clasificación del juicio de la sugerencia de llevar a cabo la intervención según rigor científico (Tabla 2) (‘Oxford Centre for Evidence-based Medicine - Levels of Evidence - CEBM’ 2009). Tabla 1. Niveles de evidencia propuestos por el Centro de Medicina Basada en la Evidencia (CEBM) de Oxford (‘Oxford Centre for Evidence-based Medicine - Levels of Evidence - CEBM’ 2009). Level Type of evidence 1a Systematic review with homogeneity of randomized control trials 1b Individual randomized control trials with narrow confidence interval Material y métodos 41 1c All or none related outcome 2a Systematic review with homogeneity of cohort studies 2b Individual cohort study (including low quality randomized control trials; e.g., <80% follow-up) 2c "Outcomes" Research; Ecological studies 3a Systematic review with homogeneity of case-control studies 3b Individual case-control study 4 Case-series (and poor quality cohort and case-control studies) 5 Expert opinion without explicit critical appraisal, or based on physiology, bench research or "first principles" Tabla 2. Grados de recomendación propuestos por el Centro de Medicina Basada en la Evidencia (CEBM) de Oxford (‘Oxford Centre for Evidence-based Medicine - Levels of Evidence - CEBM’ 2009). A Consistent level 1 studies B Consistent level 2 or 3 studies or extrapolations from level 1 studies C Level 4 studies or extrapolations from level 2 or 3 studies D Level 5 evidence or troublingly inconsistent or inconclusive studies of any level Otro procedimiento para la evaluación de la certeza de la evidencia es la metodología GRADE (Grading of Recommendations Assessment, Development and Evaluation), que ofrece un proceso estructurado para desarrollar y presentar Material y métodos 48 Como antes se ha explicado, cuando se combinan datos para variables dicotómicas se puede utilizar la odds ratio (OR), la razón de riesgo o riesgo relativo (RR) o la diferencia de riesgo (DR). La RR y la DR se deben emplear en la evaluación en estudios de cohortes prospectivos, mientras que la OR se utiliza para estudios retrospectivos, casos-control, o transversales (Ranganathan et al. 2015). Modelos de efectos fijos y modelos de efectos aleatorios Para analizar el tamaño del efecto se pueden emplear modelos de efectos fijos o de efectos aleatorios. Un modelo de efectos fijos supone que el efecto del tratamiento es el mismo y que la variación entre los resultados en diferentes estudios se debe a un error aleatorio, por lo que se utiliza cuando se considera que los estudios tienen el mismo diseño y metodología, con escasa variabilidad en los diferentes resultados. Por tanto, los modelos fijos asumen homogeneidad del efecto en los diferentes estudios que se combinan, incluyendo como únicos determinantes en el peso ponderado en el metaanálisis el tamaño del estudio y su propia varianza. El método de Mantel-Haenszel (MH) es uno de los más utilizados en la estimación ponderada con efectos fijos en medidas de resultado dicotómicas. El estimador global o combinado de Mantel-Haenszel es consistente incluso si los estudios individuales tienen pocos sujetos, siendo la media ponderada de los efectos de los estudios individuales con factores de ponderación peso MH. Por otro lado, un modelo de efectos aleatorios supone heterogeneidad entre los estudios que se combinan y, en consecuencia, se utilizan cuando se considera que los estudios son diferentes, incluso si una prueba de heterogeneidad no muestra un resultado significativo. El modelo de efectos aleatorios supone que el tamaño del efecto del tratamiento difiere entre los estudios y, por tanto, se cree que las diferencias en la variación entre los estudios Material y métodos 49 se deben no solo al error aleatorio, sino también a la variabilidad de los resultados entre las investigaciones. Por ello, la ponderación de los estudios tiene en cuenta, además de la varianza o tamaño de los mismos, la varianza entre los diferentes estudios. El modelo de efectos aleatorios de la varianza inversa es un método de agregar dos o más variables aleatorias para minimizar la varianza del promedio ponderado, de manera que cada variable aleatoria se pondera en proporción inversa a su varianza, es decir, proporcional a su precisión. Una modificación del método de la varianza inversa es el modelo de efectos aleatorios DerSimonian y Laird (DerSimonian & Laird 1986), que incorpora la variabilidad interestudio en el estimador combinado, de manera que el peso de cada estudio se calcula como el inverso de la suma de la varianza del estudio individual más la varianza interestudio. En la práctica, el método de DerSimonian y Laird incorpora una prueba de heterogeneidad en la estimación de la varianza interestudio. Cuando el valor p de la prueba de heterogeneidad es no significativo, el resultado del modelo de efectos aleatorios es igual al de efectos fijos. Sin embargo, si la prueba de heterogeneidad es significativa, la varianza interestudio aumenta progresivamente cuanto menor es el valor p de dicha prueba, de forma que los pesos para cada estudio en el modelo de efectos aleatorios tienden a igualarse (Hasselblad et al. 1995). Heterogeneidad La heterogeneidad es una medida de la varianza entre los estudios, esto es, un método para determinar si el grado de heterogeneidad es mayor de lo que se esperaría que ocurriera naturalmente cuando el tamaño del efecto calculado a partir de varios estudios es mayor que el error de muestreo. Una superposición deficiente de los intervalos de confianza para los resultados de los estudios individuales, representados gráficamente mediante Material y métodos 50 líneas horizontales en el forest plot (Lewis & Clarke 2001), generalmente indica la presencia de heterogeneidad estadística. El gráfico L’Abbé plot (Figura 13) (L’Abbé et al. 1987) permite mostrar visualmente las variaciones en los resultados observados en los estudios individuales incluidos en el metaanálisis, esto es, permite explorar la heterogeneidad entre los estudios. Figura 13. Gráfico de L'Abbé (L’Abbé et al. 1987). La línea continua representa la línea sin efecto. La línea discontinua representa el efecto combinado de todos estudios como OR o RR. Los círculos representan los resultados de estudios individuales, y su tamaño representa el peso del estudio. Por otro lado, cuando el valor p de la prueba de chi2 es bajo (inferior a 0.1) se considera que existe heterogeneidad estadística en los efectos de la intervención (variación en las estimaciones del efecto más allá del azar). Finalmente, el estadístico I2 con un valor de entre 0 y 40% sugiere que no es importante la heterogeneidad, 30-60% indica heterogeneidad moderada, 50- Material y métodos 51 90% heterogeneidad sustancial, mientras que un valor 75-100% indica una heterogeneidad considerable (Deeks et al. 2019). Sesgo de publicación El sesgo de publicación es el tipo más común de sesgo en los metaanálisis, generando distorsión en el resultado debido a la mayor probabilidad de publicación de estudios estadísticamente significativos en lugar de estudios no significativos. Un funnel plot o gráfico de embudo (Figura 14) permite probar la presencia o ausencia de sesgo de publicación. En él, los estudios se representan en un diagrama de dispersión con el tamaño del efecto en el eje x y la precisión o el tamaño total de la muestra en el eje y. Cuando los puntos forman un embudo invertido (base amplia que se estrecha hacia la parte superior de la gráfica), indica ausencia de un sesgo de publicación. Por otro lado, una forma asimétrica del gráfico (sin puntos en un lado del gráfico) hace sospechar un sesgo de publicación (Sterne et al. 2011). Material y métodos 52 Figura 14. Ejemplo de gráfico de embudo simétrico (a), compatible con una menor probabilidad de sesgo de publicación; y asimétrico (b), que indica un posible sesgo de publicación (Rao et al. 2017). (a) (b) PRIMERA PARTE DIABETES MELLITUS Y PRONÓSTICO DEL TRATAMIENTO DE CONDUCTOS CAPÍTULO I Association between diabetes and the prevalence of radiolucent periapical lesions in root-filled teeth: systematic review and meta-analysis Capítulo publicado en: Segura-Egea JJ, Martín-González J, Cabanillas-Balsera D, Fouad AF, Velasco-Ortega E, López-López J (2016) Association between diabetes and the prevalence of radiolucent periapical lesions in root-filled teeth: systematic review and meta-analysis. Clinical Oral Investigations 20, 1133–41. Capítulo I 55 ABSTRACT Introduction. The question of whether diabetes mellitus can influence the outcome of root canal treatment (RCT) remains unclear. The aim of this systematic review and meta-analysis was to analyze scientific available evidence on the association between diabetes and the presence of radiolucent periapical lesions (RPLs) in root-filled teeth (RFT). Methods. The review question was as follows: in adult patients who had endodontically treated teeth, does the absence or presence of diabetes result in an increase in the prevalence of RPL associated to RFT? A systematic MEDLINE/PubMed, Wiley Online Database, Web of Science, and Scopus search was conducted using the following MeSH and keywords: Diabetes Mellitus OR Diabetes OR Diabetic OR Hyperglycemia, AND Endodontics, Periapical Periodontitis, Periapical Diseases, Apical Periodontitis, Periradicular Lesion, Periapical Radiolucency, Radiolucent Periapical Lesion, Root Canal Treatment, Root Canal Preparation, Root Canal Therapy, Root Filled Teeth, Endodontically Treated Teeth. Seven studies reporting data on the prevalence of RPL associated to RFT both in diabetic and control subjects were included. Results. After the study selection, seven epidemiological studies fulfilled the inclusion criteria, representing data from 1593 root canal treatments, 1011 in nondiabetic control subjects, and 582 in diabetic patients. The calculated pooled odds ratio (OR = 1.42; 95 % CL = 1.11–1.80; p = 0.0058) indicates that diabetic patients have higher prevalence of RFT with RPLs than controls. Conclusion. Available scientific evidence indicates that diabetes is significantly associated to higher prevalence of periapical radiolucencies in endodontically treated teeth, being an important putative pre-operative prognostic factor in RCT. Clinical relevance. Taking into account that diabetes is the third most prevalent chronic medical condition among dental patients, endodontic providers should be aware of the relationship between the outcome of endodontic treatment and diabetes. Capítulo I 56 Keywords. Diabetes mellitus. Meta-analysis. Periapical inflammation. Persistent apical periodontitis. Root canal treatment outcome. Root-filled teeth. Capítulo I 57 1. INTRODUCTION Apical periodontitis (AP) is an inflammatory process around the apex of a tooth root, following the bacterial infection of the pulp space of the tooth [1]. The bone lesion associated with apical periodontitis is characterized radiographically by the presence of radiolucent periapical lesion (RPL), i.e., a radiolucent image surrounding the root apex of the affected tooth [2]. AP is an extraordinarily prevalent problem [3]. In the USA, radiographic signs of periapical disease are evident in 4.1–5.1 % of all teeth [4, 5]. The incidence of new cases of apical periodontitis over a 24-year period in the USA ranges from 27 to 41 % depending on age [6]. In Europe, the prevalence of AP is as high as 34–61 % of individuals and 2.8–4.2 % of the teeth [7, 8], increasing with patient’s age [9]. The treatment for teeth with AP is root canal treatment (RCT) [10]. In the USA, 4.8–5.5 % of teeth have been endodontically treated [4, 5] and 10 % of young military recruits were shown to have existing RCT [11]. In Europe, the prevalence of endodontic treatment is estimated around 41–59 % of individuals and 2–6.4 % of teeth [7, 8]. When RCT fails, resolution of the periapical lesion and complete healing of periapical tissues do not occur, persisting AP [12, 13]. Persistent apical periodontitis (PAP) is characterized radiographically by a RPL associated with the root-filled tooth (RFT). The prevalence of radiographic evidence of persistent AP is 31–36 % in the USA [4, 5] and 24–65 % in European countries [7, 8, 14]. Periapical granulomas and cysts are the most common periapical lesions of endodontic origin associated with PAP. However, some of the RPL associated with RFT may not represent PAP, but incomplete healed lesions after root canal treatment, periapical connective scars [15], or non-endodontic pathosis [16]. Factors implicated in persistent AP are not only intra-operatives, such as inadequate aseptic control, missed canals, insufficient instrumentation, and leaking temporary or permanent restorations [17], but also systemic factors, such as pro-inflammatory status and impaired immune response associated with systemic diseases [14, 18]. One of the systemic diseases whose possible association with AP has been investigated is diabetes mellitus (DM) [14, 19], a heterogeneous group of Capítulo I 64 Study characteristics In the final analysis, the following seven studies were included: (1) Falk et al. [44]) [44]; (2) Fouad and Burleson [45]) [45]; (3) Britto et al. [46]) [46]; (4) Segura- Egea et al. [47]) [47]; (5) López-López et al. [48]) [48]; (6) Marotta et al. [49]) [49]; and (7) Marques-Ferreira et al. [50]) [50]. Table 2 summarizes the study design, subjects and sample size, diagnosis of RPLs, main results, and evidence level [31]. Radiographic criteria for the diagnosis of apical periodontitis, when are provided, are shown; two studies [46, 49] used the Strindberg’s criteria [51], one study had longitudinal data and used clinical and radiographic analysis by supervising endodontists [45] and three others [47, 48, 50] the PAI system score [52]. Table 2 Studies included in the systematic review. Study design, subjects and sample size, diagnosis of radiolucent periapical lesions, and main results on association between diabetes and RFT with RPL and evidence level Capítulo I 65 Meta-analysis For each selected article, the results were extracted and compiled into a table of evidence, and descriptive statistics and odds ratios calculated (Table 3). When the OR is greater than 1, it indicates that diabetic patients show higher prevalence of RFT with RPLs than control subjects. The BDT was non-significant (Breslow-Day = 4.63; df = 6; p = 0.59), indicating homogeneity among the ORs of the included studies (Fig. 2, L’Abbé plot). Moreover, the proportion of variation through studies due to heterogeneity was very low (I 2 = 0 %; 95 % CI = 0 to 59 %). Mantel-Haenszel method and the Robins, Breslow, and Greenland variance formula, with fixed effects, provide a pooled OR = 1.42 (95 % CI = 1.11– 1.80; chi2 = 7.60; p = 0.0058), indicating that the calculated pooled OR differs significantly from 1. Forest plot shows the ORs for each study and the overall OR calculated from the meta-analysis (Fig. 3). These results indicate that diabetic patients have significantly higher prevalence of RFT with RPLs than control subject. Table 3 Results extracted and compiled, descriptive statistics, and odds ratios calculated Capítulo I 66 Fig. 2. L’Abbé plot showing the percentage of root-filled teeth (RFT) with radiolucent periapical lesions (RPLs) in the seven studies for the comparison of diabetic and controls. Size of circle is proportional to size of study. Study designations: (1) Falk et al. [44]); (2) Fouad and Burleson [45]); (3) Britto et al. [46]); (4) Segura-Egea et al. [47]); (5) López-López et al. [48]); (6) Marotta et al. [49]); and (7) Marques-Ferreira et al. [50]) Capítulo I 67 Fig. 3. Forest plot of odds ratios and 95 % confidence limits (CL) based on data from seven studies for the comparison of diabetic patients and control subjects with regard to the prevalence of RFT (root-filled teeth) with RPLs (radiolucent periapical lesions). The size of each rectangle is proportional to the total sample size for the diabetic/control comparison in that study. Overall estimate based on combined data from the seven studies. The size of the diamond is proportional to the percent weight of each study, i.e., the combined sample size for the diabetic/control comparison. The solid line indicates an odds ratio of 1.0. The dashed lineindicates the overall odds ratio. OR odds ratio, LCL lower confidence level, UCL upper confidence level Interpretation and assessment of the included studies The time frame of publication of the seven studies was 1989 and 2014; however, six of them were published between 2003 and 2014 (Table 2). One was a longitudinal study with two or more years of follow-up, in which successful versus uncertain/failed treatments were compared [45], and the other six were cross-sectional studies [44, 46–50]. The included studies represent data from 1368 subjects, 730 controls, and 319 diabetic patients. Capítulo I 68 In the study of Falk et al. [44], long-duration diabetics showed higher frequency of RFT with RPLs (26 %) compared to non-diabetic patients (21 %) (OR = 1.31; 95 % CL = 0.85–2.01; p = 0.20). However, diabetic women had significantly more RFT with RPLs than control women (p < 0.01). Fouad and Burleson [45] investigated 531 RCT, 72 in diabetic patients, finding increased likelihood of RPLs diabetics, but without statistical significance (OR = 1.24; 95 % CL = 0.70–2.13; p = 0.20). Nevertheless, the frequency of RPLs in RFT of diabetic patients with preoperative periradicular lesions was significant compared to controls (p = 0.007) and when controlling for a number of confounding variables [45]. The study of Britto et al. [46] assessed the periapical status of 99 subjects (56 diabetics) using periapical and panoramic radiographs. Strindberg’s criteria [51] were used to diagnose RPLs. The results did not find significant difference in the percentage of RFT with RPLs between controls [44 %] and diabetics [46] (OR = 1.09; 95 % CL = 0.46–2.63; p = 0.82). However, type 2 diabetic men were more likely to have residual RPLs in their RFT (p < 0.05). The study sample in this investigation showed a striking prevalence of RPLs, finding one or more teeth with RPLs in 97 and 87 % of diabetic patients and control subjects, respectively. Segura-Egea et al. [47] included in their study 38 control subjects and 32 diabetic patients, using periapical radiographs and PAI score system [52] to assess the periapical status. RPLs were found in 83 % of RFT in the diabetic group, whereas only 60 % of RFT in the control group had periapical lesions (OR = 3.33; 95 % CL = 0.48–37.93; p = 0.17). The study of López-López et al. [48] compared the prevalence of RFT with RPLs in well-controlled diabetic patients and control subjects. In this study, patients and controls were age- and sex-matched, and diabetic patients had glycated hemoglobin levels (HbA1c) ≤ 6.5 %. Periapical status of RFT was assessed using panoramic digital radiographs and the PAI index [52]. The results showed that the percentage of RFT with RPLs was almost twice higher in diabetic patients (46 %) than in control subjects (24 %), but the difference was not statically significant (OR = 2.67; 95 % CL = 0.76–10.06; p = 0.09). Marotta et al. [49], in another cross-sectional study, used periapical and panoramic radiographs and Strindberg’s criteria [51] for the diagnostic of RPLs in RFT of diabetic and control subjects. They found that RPLs were significantly Capítulo I 69 more common in untreated teeth from diabetics (10 %) than in nondiabetic controls (7 %) (p = 0.03). However, there was not significant difference in the prevalence of RPLs associated with RFT in diabetics (46 %) and control subjects (38 %) (OR = 1.39; 95 % CL = 0.81–2.39; p = 0.21). Finally, the study conducted by Marques-Ferreira et al. [50] compared the success rate of RFT in two groups of 23 patients, healthy control group and diabetic group. Periapical status was assessed radiographically using the PAI score system [52]. The results demonstrated no significant differences between both groups in the prevalence of RFT with RPLs (OR = 3.05; 95 % CL = 0.84–12.48; p = 0.06). 5. DISCUSSION Since the mid-twentieth century to today, numerous animal [53–60] and human studies [2, 26, 27, 40, 45–50, 61] have investigated the possible relationship between endodontic infections and DM. The endodontic variables analyzed in human studies have been the prevalence of RPLs, the prevalence of RCT, and the outcome of RCT, assessed as the percentage of RFT with or without RPLs, or as the prevalence of tooth extraction after nonsurgical RCT (NSRCT) [14]. Even though the results of these studies are not conclusive, available scientific evidence suggest an association between DM and a higher prevalence of RPLs, greater size of RPLs, and frequency of odontogenic infections [14, 19]. On the contrary, the existing data about the association of diabetes with the prevalence of RCT are sparse and inconclusive [14]. Finally, several studies have investigated the potential relationship between diabetes mellitus and the survival of root canal-treated teeth analyzing the prevalence of tooth extraction after NSRCT [41–43]. Three of these studies [17, 42, 43] provide a very significant OR (p < 0.01) for the contribution of diabetes to decreased retention of RFT. Four studies have provided longitudinal evaluation of the success of root canal treatment longitudinally [17, 18, 41, 45]. The Marending et al. [18] paper showed that diabetes was one of a number of medical problems that significantly influenced the outcomes. The three other studies [17, 41, 45] agreed that when the treatment of all teeth is considered, diabetes did not affect Capítulo I 70 the outcome. The Fouad and Burleson study [45] showed that when only teeth with preoperative lesions are considered, and when controlling for a number of important confounding variables, teeth from diabetics were more significantly classified as uncertain or failing, at two or longer years after treatment. The objective of this systematic review and meta-analysis has been to analyze the potential association between diabetes mellitus and the percentage of RFT with or without RPLs. The observational epidemiological studies involved were “outcomes” research, including one longitudinal study with two or more years of follow-up [45], level of evidence 2, and six cross-sectional studies [44, 46–50], level of evidence 3 [31]. The homogeneity of the seven studies (Breslow-Day = 4.63; df = 6; p = 0.59; and I 2 = 0 %; 95 % CI = 0 to 59 %) was high. Thus, the variations across studies were casual rather than due to heterogeneity. The reasonable time frame of publication of the studies included in this review (1989 to 2014) reinforces the possibility of comparison, discarding important changes in dental concepts, materials, and/or treatments over time [62, 63]. The analysis of the study designs is also very important in a systematic review like this. However, in the present review, most of the included studies were cross-sectional studies. Cross-sectional studies demonstrate differences in the prevalence of PAP, but longitudinal studies could show differences between diabetic and control subjects regarding the healing process of the periapical pathosis. Individually, none of the studies provides significant OR regarding the association of diabetes with the prevalence of RFT with periapical lesions. However, pooled OR provided by MH method, with fixed effects, was significant (OR = 1.42; 95 % CI = 1.11–1.80; p = 0.006) indicating that diabetes is associated to the prevalence of RFT with RPLs. It can be concluded that available scientific evidence supports the association between diabetes and persistent apical periodontitis. This result is in agreement with the studies showing that diabetic patients have delayed periapical repair and greater likelihood of RFT loss [17, 42, 43, 45]. Capítulo I 71 The biological mechanisms linking periapical status of RFT and diabetes mellitus could be the following: (1) diabetes predisposes to chronic inflammation, (2) diabetes reduces tissue repair capacity, (3) diabetes impaired the immune response enhancing the susceptibility to infections, and (4) diabetes impaired bone turnover and delayed wound healing [14, 23, 64, 65]. In inflamed periapical tissues of endodontically treated teeth, diabetes could compromise immune response, upregulating periapical inflammation and altering bone turnover and wound healing, increasing the prevalence of apical periodontitis in RFT [14]. Considering that diabetes is the third most prevalent chronic medical condition among dental patients [66], endodontic providers should be aware of the relationship between the outcome of endodontic treatment and diabetes, should keep current data on the diabetic status of their patients, and should inform diabetic patients of the risks involved in endodontic therapy for them. 6. CONCLUSION Available scientific evidence indicates that diabetes is significantly associated with higher prevalence of periapical radiolucencies in endodontically treated teeth. Well-designed prospective studies are required to further investigate the association between diabetes and RCT outcome and to definitively determine the precise increased risk of treatment failure in diabetic patients. However, at this time, diabetes should be recognized as an important putative pre-operative prognostic factor in endodontic treatment. Compliance with ethical standards Conflict of interest: The authors declare that they have no conflict of interest. Capítulo I 72 Funding: The work was supported by the Department of Stomatology, Endodontics Section, University of Sevilla, Seville, Spain. Ethical approval: This article does not contain any studies with human participants or animals performed by any of the authors. Informed consent: For this type of study, formal consent is not required. REFERENCES 1. Siqueira JRJF, Rôças IN (2014) Present status and future directions in endodontic microbiology. Endod Topics 30:3–22 2. Bender IB, Seltzer S (2003) Roentgenographic and direct observation of experimental lesions in bone: I. 1961. J Endod 29:702–706 3. Figdor D (2002) Apical periodontitis: a very prevalent problem. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 94:651–652 4. Buckley M, Spångberg LS (1995) The prevalence and technical quality of endodontic treatment in an American subpopulation. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 79:92–100 5. Chen CY, Hasselgren G, Serman N, Elkind MS, Desvarieux M, Engebretson SP (2007) Prevalence and quality of endodontic treatment in the Northern Manhattan elderly. J Endod 33:230–234 6. Caplan DJ, Chasen JB, Krall EA, et al. (2006) Lesions of endodontic origin and risk of coronary heart disease. J Dent Res 85:996–1000 7. Jiménez-Pinzón A, Segura-Egea JJ, Poyato M, Velasco E, Ríos JV (2004) Prevalence of apical periodontitis and frequency of root-filled teeth in an adult Spanish population. Int Endod J 37:167–173 8. López-López J, Jané-Salas E, Estrugo-Devesa A, et al. (2012) Frequency and distribution of root filled teeth and apical periodontitis in an adult population of Barcelona, Spain. Int Dental J 62:40–46 Capítulo I 73 9. Eriksen HM (1998) Epidemiology of apical periodontitis. In: Ørstavik D, Pitt Ford TR (eds) Essential Endodontology. Prevention and treatment of apical periodontitis. Blackwell Science, London, pp. pp.179–pp.191 10. Ørstavik D, Pitt Ford T (2007) Apical periodontitis: microbial infection and host responses. In: Ørstavik D, Pitt Ford TR (eds) Essential Endodontology. Prevention and treatment of apical periodontitis, 2nd edn. Wiley-Blackwell, London, UK, pp. pp:179–pp:191 11. Winward BJ, Yaccino JM, Kirkpatrick TC (2014) A panoramic survey of air force basic trainees: how research translates into clinical practice. J Endod 40:1332–1337 12. Sundqvist G, Figdor D (1998) Endodontic treatment of apical periodontitis. In: Ørstavik D, Pitt Ford TR (eds) Essential Endodontology. Blackwell, Oxford, pp. pp:242–pp:277 13. Nair PNR (2006) On the causes of persistent apical periodontitis: a review. Int Endod J 39:249–281 14. Segura-Egea JJ, Martín-González J, Castellanos-Cosano L (2015) Endodontic medicine: connections between apical periodontitis and systemic diseases. Int Endod J 48:933–951 15. Love RM, Firth N (2009) Histopathological profile of surgically removed persistent periapical radiolucent lesions of endodontic origin. Int Endod J 42:198–202 16. Koivisto T, Bowles WR, Rohrer M (2012) Frequency and distribution of radiolucent jaw lesions: a retrospective analysis of 9723 cases. J Endod 38:729–732 17. Ng YL, Mann V, Gulabivala K (2011) A prospective study of the factors affecting outcomes of non-surgical root canal treatment: part 2: tooth survival. Int Endod J 44:610–625 18. Marending M, Peters OA, Zehnder M (2005) Factors affecting the outcome of orthograde root canal therapy in a general dentistry hospital practice. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 99:119–124 19. Segura-Egea JJ, Castellanos-Cosano L, Machuca G, et al. (2012) Diabetes mellitus, periapical inflammation and endodontic treatment outcome. Med Oral Patol Oral Cir Bucal 17:e356–e361 Capítulo II 80 ABSTRACT Previous studies have found an association between the outcome of root canal treatment (RCT) and diabetic status. This systematic review and metaanalysis aimed to analyse the potential relationship between diabetes and the occurrence of extracted root filled teeth (RFT). The clinical PICO question was as follows: in adult patients with RFT, does the absence or presence of diabetes influence the prevalence of RFT extraction? The key words used in the systematic search were as follows: (Diabetes OR Diabetes Mellitus OR Hyperglycaemia OR Diabetic) AND (Endodontic OR Endodontics OR Endodontic Treatment OR Root Canal Treatment OR Root Canal Preparation OR Root Canal Therapy OR Root Filled Teeth OR Endodontically Treated Teeth) AND (Extraction OR Retention OR Survival OR Success OR Failure OR Outcome). The primary outcome variable was odds ratio (OR) for the frequency of extracted RFT in diabetics and healthy subjects. The method of DerSimonian–Laird with random effects was used to calculate the overall OR. Three hundred titles were identified, and three studies achieved the inclusion criteria. Data from 54 936 root canal treatments, 50 301 in nondiabetic control subjects and 4635 in diabetic patients, were analysed. The calculated overall odds ratio (OR = 2.44; 95% CI = 1.54–3.88; P = 0.0001) implies that diabetics had a significantly higher prevalence of extracted RFT than healthy nondiabetic subjects. The results of available studies indicate a significant relationship between DM and increased frequency of nonretained root filled teeth. Diabetes mellitus should be considered an important preoperative prognostic factor in root canal treatment. Keywords: diabetes mellitus, meta-analysis, nonretention, root canal treatment outcome, root filled teeth, tooth extraction. Capítulo II 81 1. INTRODUCTION Apical periodontitis (AP) is an inflammatory process surrounding the apex of a root, subsequent to bacterial infection of the root canal system (Siqueira & Rôças 2014). Apical periodontitis is associated with a bone lesion characterized radiographically by a radiolucent image around the root apex of the affected tooth (Bender & Seltzer 2003). AP is a remarkable health problem, being one of the most prevalent diseases in the world (Figdor 2002). The prevalence of AP increases with age and ranges between 34–61% of individuals (Buckley & Spångberg 1995, Chen et al. 2007). Radiographic signs of periapical disease are found in 2.8–5.1% of teeth (Eriksen 1998, Jiménez‐Pinzón et al. 2004, Caplan et al. 2006, López‐López et al. 2012). The incidence of new cases of AP over a 24‐year period is as high as 27–41%, depending on age (Buckley & Spångberg 1995). Root canal treatment (RCT) is the treatment of choice for teeth with AP (Ørstavik & Pitt Ford 2007), with the prevalence of RCT being 41–59% of individuals and 2–6.4% of teeth (Eriksen 1998, Jiménez‐Pinzón et al. 2004, Caplan et al. 2006, López‐López et al. 2012). Ten per cent of young military recruits had at least one root filled tooth (RFT) (Winward et al. 2014). The outcome of RCT has traditionally been studied based on clinical parameters, radiographic assessment of periapical status and histopathological evaluation of extirpated tissue (Lazarski et al. 2001). According to clinical and radiographic evaluation criteria, follow‐up studies have reported success rates of 53–95% (Jokinen et al. 1978, Ingle 1985, Ng et al. 2011). Preoperative factors (systemic conditions, impaired immune response, root fractures, anatomic irregularities, periodontal diseases, etc.) (Marending et al. 2005, Segura‐ Egea et al. 2015), intra‐operative factors (poor initial therapy, incomplete aseptic control, inadequate instrumentation or root filling) and postoperative factors (crown fractures or improper coronal restoration) determine the prognosis of RCT (Vire 1991, Ng et al. 2011). Persistent apical periodontitis indicates RCT failure, and the subsequent untoward events include root canal retreatment, apical surgery and extraction. The assessment of the outcome of RCT can be made by quantifying the Capítulo II 82 incidence of these events (Lazarski et al. 2001). The analysis of more than 100.000 cases of RCT revealed an overall incidence 6.4% of untoward events, 3.6% extractions, 1.9% root canal retreatments and 1% periradicular surgery (Lazarski et al. 2001). Another study conducted in Taiwan found untoward events in 9.7% of RFT after a 5‐year follow‐up, with the most common untoward event being tooth extraction (71.1%) followed by root canal retreatment (24.1%) and apical surgery (4.8%) (Chen et al. 2008). The percentage of retention of RFT is an indicator that allows the outcome of RCT to be assessed. A study conducted in the Netherlands in 1983 evaluated the outcome of RCT in servicemen and reported that 55% of root filled teeth had been retained after 17 years (Meeuwissen & Eschen 1983). Tooth loss after RCT correlated with the number of proximal contacts, age, history of facial injury, number of missing teeth and abutment status (Caplan & Weintraub 1997). Extractions after RCT may be due to prosthetic (59%), periodontal (32%) and endodontic causes (9%) (Vire 1991). Diabetes mellitus (DM) is a systemic disease with a possible association with AP (Segura‐Egea et al. 2012, 2015). DM is a metabolic disorder due to pancreatic β‐cells dysfunction, with a deficiency in insulin secretion, and/or peripheral insulin resistance, resulting in hyperglycaemia as the main feature (ECDCDM 2000, Mealey & Oates 2006). The function of leucocytes is altered in diabetes, with increased release of pro‐inflammatory cytokines and decreased secretion of macrophage growth factors, facilitating the development of chronic inflammatory processes and reducing tissue repair ability (Iacopino 2001). Additionally, the levels of advanced glycation end‐products (AGEs) are elevated in diabetic patients, increasing tissue oxidative stress and upregulating inflammatory responses (Fouad & Huang 2015). Moreover, poorly controlled diabetics have further immunological alterations in leucocyte function and wound healing (Delamaire et al. 1997, Iacopino 2001, Salvi et al. 2008). A recent meta‐analysis has demonstrated a significant association between diabetes and the frequency of RFT with radiological signs of persistent apical periodontitis, in agreement with studies showing a delayed periapical repair in Capítulo II 83 diabetic patients (Segura‐Egea et al. 2016). Consequently, increased failure of root canal treatment with greater likelihood of RFT loss would be expected in diabetic patients. In this study, a systematic review and meta‐analysis were carried out to investigate the potential relationship between diabetes and the occurrence of extracted RFT. 2. REVIEW The clinical PICO question to be answered was as follows: in adult patients with RFT, does the absence or presence of diabetes influence the prevalence of RFT extraction? Literature search strategy The systematic review was carried out following conventional methods (Stroup et al. 2000, Bader 2004). After formulating the PICO question, the search plan was designed and the articles found in the search were selected according to the previously established criteria for inclusion and exclusion. Subsequently, the quality of the articles was evaluated and their results collected and analysed (Pak et al. 2012). The search was carried out in MEDLINE/PubMed, Web of Science, Scopus and the Wiley Online Database. The groupings of Mesh terms and key words used were as follows: (Diabetes OR Diabetes Mellitus OR Hyperglycaemia OR Diabetic) AND (Endodontic OR Endodontics OR Endodontic Treatment OR Root Canal Treatment OR Root Canal Preparation OR Root Canal Therapy OR Root Filled Teeth OR Endodontically Treated Teeth) AND (Extraction OR Retention OR Survival OR Success OR Failure OR Outcome) (Table 1). Capítulo II 84 Table 1. List of the MeSH and key word combinations used for the search strategy. ((‘diabetes mellitus’[MeSH Terms] OR (‘diabetes’[All Fields] AND ‘mellitus’[All Fields]) OR ‘diabetes mellitus’[All Fields]) OR (‘diabetes mellitus’[MeSH Terms] OR (‘diabetes’[All Fields] AND ‘mellitus’[All Fields]) OR ‘diabetes mellitus’[All Fields] OR ‘diabetes’[All Fields] OR ‘diabetes insipidus’[MeSH Terms] OR (‘diabetes’[All Fields] AND ‘insipidus’[All Fields]) OR ‘diabetes insipidus’[All Fields]) OR Diabetic[All Fields] OR (‘hyperglycaemia’[All Fields] OR ‘hyperglycaemia’[MeSH Terms] OR ‘hyperglycaemia’[All Fields])) AND (Endodontic[All Fields] OR (‘endodontics’[MeSH Terms] OR ‘endodontics’[All Fields]) OR (Endodontic[All Fields] AND (‘therapy’[Subheading] OR ‘therapy’[All Fields] OR ‘treatment’[All Fields] OR ‘therapeutics’[MeSH Terms] OR ‘therapeutics’[All Fields])) OR ((‘dental pulp cavity’[MeSH Terms] OR (‘dental’[All Fields] AND ‘pulp’[All Fields] AND ‘cavity’[All Fields]) OR ‘dental pulp cavity’[All Fields] OR (‘root’[All Fields] AND ‘canal’[All Fields]) OR ‘root canal’[All Fields]) AND (‘therapy’[Subheading] OR ‘therapy’[All Fields] OR ‘treatment’[All Fields] OR ‘therapeutics’[MeSH Terms] OR ‘therapeutics’[All Fields])) OR (‘root canal preparation’[MeSH Terms] OR (‘root’[All Fields] AND ‘canal’[All Fields] AND ‘preparation’[All Fields]) OR ‘root canal preparation’[All Fields]) OR (‘root canal therapy’[MeSH Terms] OR (‘root’[All Fields] AND ‘canal’[All Fields] AND ‘therapy’[All Fields]) OR ‘root canal therapy’[All Fields]) OR ((‘plant roots’[MeSH Terms] OR (‘plant’[All Fields] AND ‘roots’[All Fields]) OR ‘plant roots’[All Fields] OR ‘root’[All Fields]) AND Filled[All Fields] AND (‘tooth’[MeSH Terms] OR ‘tooth’[All Fields] OR ‘teeth’[All Fields])) OR (‘tooth, nonvital’[MeSH Terms] OR (‘tooth’[All Fields] AND ‘nonvital’[All Fields]) OR ‘nonvital tooth’[All Fields] OR (‘endodontically’[All Fields] AND ‘treated’[All Fields] AND ‘teeth’[All Fields]) OR ‘endodontically treated teeth’[All Fields])) AND (extraction[All Fields] OR (‘retention (psychology)’[MeSH Terms] OR (‘retention’[All Fields] AND ‘(psychology)’[All Fields]) OR ‘retention (psychology)’[All Fields] OR ‘retention’[All Fields]) OR (‘mortality’[Subheading] OR ‘mortality’[All Fields] OR ‘survival’[All Fields] OR ‘survival’[MeSH Terms]) OR success[All Fields] OR failure[All Fields] OR outcome[All Fields]) Capítulo II 85 Several endodontic journals and the references of significant papers and reviews were hand‐searched. The last search was made on 23 March 2018. Study selection and inclusion and exclusion criteria The titles and abstracts of the published studies collected in the search were screened by three investigators (D.C‐B., J.M‐G. and J.J.S‐E.). Articles that did not meet the inclusion criteria were excluded. All remaining articles were obtained, and full text reviewed independently by four reviewers (D.C‐B., J.M‐G., P.M‐M. and J.J.S‐E) according to the established inclusion criteria: (i) the type of study: epidemiological studies published from January 1988 to February 2018; (ii) studies comparing diabetic and nondiabetic patients; and (iii) studies establishing the outcome of RCTs and recording data on the frequency of RFT retention in diabetic patients and control subjects. The following were established as exclusion criteria: (i) experimental studies (laboratory and animals) and (ii) studies including only diabetic patients. When there was disagreement among the reviewers, the basis for the decision was consensus. Evidence quality evaluation and data extraction The quality of the methodology used in each study was assessed, and the results were read and analysed carefully. From each of the selected studies, the following data were recorded: name of the authors, date of publication, type of study design, sample size, quantitative results and odds ratio values, and quality level according to Oxford CEBM guidelines (OCEBM 2009). Capítulo II 86 Outcome variables and statistical analysis As a measure of the effect and as a primary outcome variable, the odds ratio (OR) calculated in each study for the relationship between the retained RFT and the diabetic state was collected. To calculate the pooled OR, the DerSimonian– Laird method with random effects was used, calculating 95% confidence intervals for the OR. The Breslow‐Day test (BDT) and the I2 test were used to assess heterogeneity among the calculated ORs (Higgins & Thompson 2002). L'Abbé plots (L'Abbé et al. 1987) were used to test homogeneity. To show the OR results, a Forest plot (Lewis & Clarke 2001) was used, along with the DerSimonian–Laird pooled estimate. StatsDirect software was used to carry out the meta‐analysis (Freemantle 2000). Significance level of P < 0.05 was considered. Search results Figure 1 presents the search strategy, which provided three hundred titles. Removing duplicates for both searches (204 items) and articles published before 1980 (3 items), 93 articles were obtained. Subsequently, the titles and the summary of the obtained studies were analysed, selecting 15 for full‐text reading. Then, 12 studies were disallowed: two of them studied loss of teeth in periodontal patients (Faggion et al. 2007, Dannewitz et al. 2016), six studies only provided data regarding periapical lesions (Britto et al. 2003, Segura‐ Egea et al. 2012, 2016, Rudranaik et al. 2016, Arya et al. 2017, Smadi 2017), and four others did not present results on the frequency of tooth extraction after NSRCT in diabetics and controls (Fouad & Burleson 2003, Doyle et al. 2007, Lin et al. 2014, Marques‐Ferreira et al. 2014). Capítulo II 87 Figure 1. Flow diagram showing the process by which the studies were selected. Capítulo II 88 Study characteristics Three studies were included in the final analysis: (i) Mindiola et al. (2006), (ii) Wang et al. (2011) and (iii) Ng et al. (2011). Table 2 summarizes the study design, sample size, main data and level of evidence (OCEBM 2009). Table 2. Studies included in the systematic review. Study design, subjects and sample size, main results on association between diabetes and extracted RFT and evidence level Meta‐analysis Table 3 shows the compilation of the results of the selected studies, with the calculated descriptive statistics and odds ratios (Table 3). The ORs of the included studies were nonhomogeneous (Breslow‐Day test = 7.03; df = 2; P = 0.03) (Fig. 2, L'Abbé plot), with high heterogeneity (I2 = 70.8%; 95% CI = 0% to 89.3%), so the random effects model was used to calculate the weights. DerSimonian–Laird method with random effects was performed, providing a pooled OR = 2.44 (95% CI = 1.54–3.88; χ2 = 14.40; P = 0.0001), as shown in the Forest plot (Fig. 3), indicating that the probability that root filled teeth are extracted is more than twice as high in diabetics compared to healthy patients. Capítulo II 89 Table 3. Results extracted and compiled, descriptive statistics and odds ratios calculated Figure 2. 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Journal of Oral Pathology & Medicine 22, 168– 74. van Nice E (2006) Management of multiple dental infections in a dog with diabetes mellitus. Journal of Veterinary Dentistry 23, 18– 25. Vire DE (1991) Failure of endodontically treated teeth: classification and evaluation. Journal of Endodontics 17, 338– 42. Wang CH, Chueh LH, Chen SC, Feng YC, Hsiao CK, Chiang CP (2011) Impact of diabetes mellitus, hypertension, and coronary artery disease on tooth extraction after nonsurgical endodontic treatment. Journal of Endodontics 37, 1– 5. Winward BJ, Yaccino JM, Kirkpatrick TC (2014) A panoramic survey of air force basic trainees: how research translates into clinical practice. Journal of Endodontics 40, 1332– 7. Wolle CF, Zollmann LA, Bairros PO, Etges A, Leite CE, Morrone FB, et al. (2013) Outcome of periapical lesions in a rat model of type 2 diabetes: Capítulo II 103 refractoriness to systemic antioxidant therapy. Journal of Endodontics 39, 643– 7. CAPÍTULO III Endodontics and diabetes: association versus causation Capítulo publicado en: Segura-Egea JJ, Cabanillas‐Balsera D, Jiménez‐Sánchez MC, Martín‐ González J (2019) Endodontics and diabetes: association versus causation. International Endodontic Journal 52, 790–802. Capítulo III 105 ABSTRACT Endodontic Medicine has gained more attention and is becoming a more important issue in Endodontics. As an example, more than one hundred articles on this topic have been published in the last eight years. Several of these studies have found an association between endodontic variables, that is the prevalence of apical periodontitis, the prevalence of root canal treatment (RCT) and the outcome of RCT assessed as root filled teeth (RFT) with radiolucent periapical lesions (RPL) or non‐retained RFT, and several systemic diseases, such as diabetes, cardiovascular disease, smoking habits, osteoporosis, inherited coagulopaties, biological medications, low birth weight or physical fitness. However, the demonstration of association does not prove by itself the existence of a cause–effect relationship. Two variables can be related statistically to each other without either variable directly affecting the values of the other thus resulting in a non‐causal relationship. Causality is assumed when one variable is shown to contribute to the development of the other, and its removal is shown to reduce the frequency of disease. Therefore, once a significant statistical association has been found between two variables, it is necessary to exclude the presence of bias, which would imply that the association is artefactual, and to analyse if the causation criteria defined by Hill (Proceedings of the Royal Society of Medicine 1965; 58: 295‐300) are fulfilled to establish a causal relationship. Only if they are satisfied, can it be concluded that the association is causal. The aim of this study was to analyse the difference between association and causation, applying the criteria of causality to the specific case of the association between endodontic disease and diabetes mellitus. Keywords: apical periodontitis, causation criteria, diabetes mellitus, endodontic medicine, root canal treatment, systematic review. Capítulo III 112 Figure 1. Search results in PubMed of articles published in the last three decades about Endodontic Medicine. The keywords used in the search were as follows: (endodontic OR apical periodontitis OR endodontic disease) AND (systemic disease OR systemic health). Interpretation of the statistical results of epidemiological studies The application of statistical methods to the results of epidemiological studies determines if there is an association between two variables. But the demonstration of an association, by itself, is not proof that there is a cause–effect relationship. Two variables can be statistically related to each other without either variable directly affecting the values of the other setting up a non‐causal relationship. Causality is assumed when one variable (be it diabetes or oral infections) is shown to contribute to the development of the other, and its removal is shown to reduce the frequency of disease (Hill 1965). Therefore, once a significant association has been found between two variables, it is necessary to exclude the presence of bias, which would imply that the association is artefactual, and to analyse if the causation criteria defined by Capítulo III 113 Hill (1965) are fulfilled to establish a causal relationship. If they are satisfied, it could be concluded that the association is causal (Fig. 2). Figure 2. Causation versus association. Hill's causation criteria The connection between cigarette smoking and lung cancer was demonstrated by Doll & Hill (1950). Sir Austin Bradford Hill, trying to answer the question ‘What aspects of the association should especially be considered before deciding that the most likely interpretation of it is causation?’, defined a list of causation criteria to provide epidemiologic evidence of the existence of causal relationship (Hill 1965) (Table 1). Although not explicitly, the discussion of scientific papers reporting results on the possible association of endodontic pathosis with some systemic disease, should evaluate and analyse the fulfilment of these criteria. However, unfortunately, the discussions of many of the articles published on endodontic medicine do not describe these criteria. Capítulo III 114 Table 1. Hill causation criteria (Hill 1965) 4. DIABETES MELLITUS – ENDODONTIC DISEASE: CAUSATION OR ASSOCIATION? Taking as an example the possible association between endodontic disease and diabetes mellitus, the achievement of Hill's criteria (strength of the Capítulo III 115 association, temporal relationship, dose–response gradient, consistency of the association, coherence and plausibility of the association) will be analysed. Firstly, it is necessary to review briefly the scientific evidence and, secondly, consider if the Hill criteria of causation are fulfilled. Scientific evidence on the association between endodontic disease and diabetes The endodontic variables analysed in these epidemiological studies are as follows: a) the prevalence of apical periodontitis, b) the prevalence of root canal treatment and c) the outcome of root canal treatment, assessed as c.1) the prevalence of root filled teeth with or without periapical lesions or c.2) the prevalence of tooth extraction after root canal treatment. (A) PREVALENCE OF APICAL PERIODONTITIS IN DIABETIC AND CONTROL SUBJECTS The first question to be answered is whether there is an association between the prevalence of apical periodontitis and diabetes. This association was proposed by Bender et al. (1963). They observed a high percentage of diabetics amongst patients with odontogenic infections, together with greater periapical inflammatory reactions and delayed periapical healing in poorly controlled diabetic patients. They proposed that the increased local inflammation as a consequence of apical periodontitis causes an intensification of diabetes with a rise in the blood glucose, placing the patient in an uncontrolled diabetic status and developing a vicious circle. Since then, six studies have found a significantly greater prevalence of periapical lesions in diabetics compared to healthy subjects (Bender et al. 1963, Falk et al. 1989, Ueta et al. 1993, Segura‐Egea et al. 2005, López‐López et al. 2011, Marotta et al. 2012), one study reported that the frequency of periradicular lesions was higher in long‐term diabetic patients than in short‐term diabetic patients (Mesgarani et al. 2014), and another study Capítulo III 116 reported the size of periapical lesions in diabetic patients was larger (Falk et al. 1989). On the contrary, other studies (Britto et al. 2003, Correia‐Sousa et al. 2015, An et al. 2016, Smadi 2017) have not found a significant association. So, prospective studies must be conducted to confirm whether there is or not a relationship between apical periodontitis and diabetes. (B) PREVALENCE OF ROOT CANAL TREATMENT IN DIABETIC AND CONTROLS SUBJECTS The second question can be formulated as whether there is association between the prevalence of root canal treatment and diabetes. In addition to studying the prevalence of apical periodontitis, six studies have also investigated the frequency of root canal treatment in diabetic and control subjects. Three studies did not find a greater prevalence of RCT in diabetic patients compared to controls (Falk et al. 1989, Segura‐Egea et al. 2005, Marotta et al. 2012), but three studies did (López‐López et al. 2011, Correia‐Sousa et al. 2015, Smadi 2017). Therefore, there is no conclusive evidence about the association of diabetes with the prevalence of RCT. (C) OUTCOME OF RCT IN DIABETIC AND CONTROL SUBJECTS Finally, the third question that can be formulated is whether an association exists between DM and the outcome of RCT. This possible relationship can be investigated analysing the prevalence of RFT with persistent AP, or investigating the prevalence of tooth extraction after root canal treatment. Capítulo III 117 (c.1) Prevalence of RFT with apical periodontitis Studies investigating the prevalence of RFT with apical periodontitis in diabetic and healthy control subjects, have demonstrated delayed periapical healing in diabetic subjects, with a lower rate of repair associated with root filled teeth (Bender et al. 1963, Arya et al. 2017), and slower reduction of periapical lesions in poorly controlled diabetic patients (Cheraskin & Ringsdorf 1968). Other studies have found a greater percentage of RFT with AP in diabetics, compared to control subjects, but without a significant difference (Falk et al. 1989, Britto et al. 2003, Fouad & Burleson 2003, Segura‐Egea et al. 2005, López‐López et al. 2011, Marotta et al. 2012, Marques‐Ferreira et al. 2014). However, some of these studies have found significant differences only in diabetic women (Falk et al. 1989), or only in diabetic men (Britto et al. 2003) or in cases with preoperative periapical lesions (Fouad & Burleson 2003). Another study found a significantly higher prevalence of RFT with AP in all diabetics compared to control subjects (Smadi 2017). (c.2) Retention of RFT The possible relationship between diabetes and outcome following root canal treatment can also be investigated comparing the prevalence of tooth extraction after treatment in diabetic patients and normal healthy subjects. Six studies have analysed this topic demonstrating a greater likelihood of RFT loss in diabetic patients (Mindiola et al. 2006, Ng et al. 2011, Wang et al. 2011, Lin et al. 2014) and a marginal association between RFT loss and diabetes (Doyle et al. 2007). In summary, it can be concluded that the prognosis of root filled teeth is worse in diabetic patients, who are more likely to lose their root filled teeth. Compliance with Hill's criteria when analysing the association between diabetes and endodontic disease It has been demonstrated that several endodontic variables, such as the prevalence of apical periodontitis, the prevalence of root canal treatment and the Capítulo III 118 prevalence of root filled teeth with radiolucent periapical lesions, are associated with diabetic status. As a consequence, it is essential to assess whether Hill's causation criteria are met to determine whether or not the association is causal. (A) STRENGTH OF THE ASSOCIATION The first criteria refer to the size of the relative risk found. Relative risk can be determined in cohort studies and clinical trials from the incidence data. However, the odds ratio calculated in cross‐sectional and case–control studies is a good approximation to the relative risk, especially when the outcome is rare (Bland & Altman 2000). The greater the strength of the association the more likely that it is to be causal (Hill 1965). Most epidemiological studies evaluating the relationship between Endodontics and diabetes are cross‐sectional, nevertheless only some of them have determined the strength of the association by calculating the odds ratio values. A value of OR equal to 1 implies that the first variable (generally the exposure) does not affect the probabilities of the second variable (the outcome) (Bland & Altman 2000). When the OR is >1, it implies that the exposure is associated with higher odds of the outcome. When the OR is <1, the exposure is associated with lower odds of the outcome. The confidence interval (CI) is used to estimate the precision of the OR. A 95% CI means that when sampling the same population on several occasions, the calculated interval would comprise the right population parameter in approximately 95% of the cases, always assuming the absence of bias or confounding factors (Morris & Gardner 1988). A large CI indicates a low level of precision of the OR, whereas a small CI indicates a higher precision of the OR. However, the 95% CI does not determine the statistical significance, as it is inappropriate to interpret an OR with 95% CI that spans the null value (e. g. OR = 1) as indicating evidence for lack of association between the exposure and outcome. Capítulo III 119 In the cross‐sectional studies conducted to analyse the relationship between diabetes and Endodontics, the ORs calculated are not constant and are not always significant (Table 2). Moreover, the estimated 95% CI overlap, in many cases, the null value, indicating that the OR could be 1 or less. However, there are six studies analysing the outcome of RCT in diabetics and control subjects in which the P value is significant, the OR values ranged from 1.3 to 5.3, and their lower limits of the 95% CI ranged from 1.2 to 2.0 (Table 2). At a 10% disease rate in the control group, the reference OR values reflecting a ‘weak association’, a ‘moderate association’ and a ‘strong association’ are 1.5, 2.5 and 4.1 (Chen et al. 2010). Table 2. Strength of the association ‘endodontics/diabetes’ An important point to keep in mind is the age of the patients included in studies. Since incidence of both diabetes and AP and RCT increases with age, a spurious association could be found if the age of the patients is not taken into account in the design of the study. In addition, long‐term diabetics are expected to be older than short‐term diabetics. Therefore, it is important to indicate which studies made adjustments for the age in the analysis of their results and in the calculation of the OR (Table 2). As it can be seen, most studies calculated ORs without adjusting for age. Therefore, the age of the patients is also an important point to take into account in the design of new studies on the association between diabetes and endodontics. Capítulo III 120 In conclusion, the OR values do not support the existence of association between Endodontics and diabetes. Only the outcome of RCT could be considered moderately associated with the diabetic state. (B) TEMPORAL RELATIONSHIP OF THE ASSOCIATION This second criterion means that one of the variables (the cause) must precede the other variable (the effect). Taking into account that cross‐sectional studies measure both variables, the exposure and the outcome, at the same time, they cannot establish a temporal relationship. On the contrary, follow‐up studies, especially longitudinal studies, allow to establish a temporal relationship. Regarding the temporal relationship between Endodontics and diabetes, no longitudinal study is available. The published studies are cross‐sectional, focusing on the prevalence of AP, RCT and AP in teeth with RCT. But prevalence is not very useful to establish causality (Hernan 2004, Rothman & Greenland 2005). A high prevalence of periapical radiolucent lesions may reflect a high incidence of apical periodontitis, but also a delay in the healing of these lesions. However, two cross‐sectional studies (Falk et al. 1989, Mesgarani et al. 2014) provide some information about the possible temporal relationship. Both studies found worse periapical status in long‐term diabetic patients compared to short‐term diabetics. The longer the duration of diabetes, the greater the prevalence of AP and the extension of periapical lesions. However, prospective studies are needed, despite being more difficult to develop and more expensive, to definitively establish the temporal relationship. Capítulo III 121 (C) DOSE–RESPONSE GRADIENT OF THE ASSOCIATION The third criterion refers to the change in effect provoked by differing amount, intensity or duration of the cause. If a dose–response gradient can be demonstrated, the likelihood that the association is causal increases. Two studies have analysed the association of glycated haemoglobin (HbA1c) levels with the prevalence of endodontic variables, and found a significant association between higher HbA1c levels and the prevalence of apical periodontitis (Sánchez‐Domínguez et al. 2015), the prevalence of RFT (Smadi 2017) and the prevalence of RFT with AP (Smadi 2017). However, no studies have been carried out comparing the prevalence levels of AP or RCT with the prevalence of diabetes. (D) CONSISTENCY OF THE ASSOCIATION The fourth causation criterion implies that different studies, carried out by different investigators, and conducted on different populations, and sometimes in different countries, resulted in the same association. Therefore, the greater the number of studies finding the association, the more consistency and more likely that the association is causal. Seven studies have found a higher prevalence of teeth with radiolucent periapical lesions in diabetic patients compared to healthy controls (Table 3). Nevertheless, only three of them (Segura‐Egea et al. 2005, López‐López et al. 2011, Marotta et al. 2012) were significant. Four other studies found no significant differences (Falk et al. 1989, Britto et al. 2003, Correia‐Sousa et al. 2015, Smadi 2017). Therefore, the association between diabetes and apical periodontitis is not consistent. A systematic review with meta‐analysis is needed to reach a definitive conclusion. 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CAPÍTULO IV Smoking and Radiolucent Periapical Lesions in Root Filled Teeth: Systematic Review and Meta-Analysis Capítulo publicado en: Cabanillas-Balsera D; Segura-Egea JJ; Bermudo-Fuenmayor M; Martín- González J; Jiménez-Sánchez MC; Areal-Quecuty V; Sánchez-Domínguez B; Montero-Miralles P; Velasco-Ortega E (2020) Smoking and Radiolucent Periapical Lesions in Root Filled Teeth: Systematic Review and Meta-Analysis. Journal of Clinical Medicine 9, 3506. Capítulo IV 145 ABSTRACT Aim: This systematic review and meta-analysis aimed to investigate the association between smoking habits and the prevalence of radiolucent periapical lesions (RPLs) in root-filled teeth (RFT). Methods: The Population, Intervention, Comparison, and Outcome (PICO) question was: in adult patients who have RFT, does the absence or presence of a smoking habit affect the prevalence of RPLs associated with RFT? Systematic MEDLINE/PubMed, Wiley Online Database, Web of Science, Scopus, and PRISMA protocol were used to evaluate and present the results. Studies comparing smokers with control non-smoker subjects, including RFT, and providing data on the prevalence of RFT with RPLs, were included. The Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) system was used for certainty in the evidence. The risk of bias was assessed according to Cochrane Collaboration common scheme for bias and ROBINS-I tool. Cumulative meta-analysis was performed with a random effects model. PROSPERO registration code: CRD42020165279. Results: Four studies reported data on inclusion criteria, representing data from 9257 root-filled teeth—4465 from non-smokers and 4792 from smoker patients. The meta-analysis provided an odds ratio indicating a significant association between smoking and higher prevalence of root filled teeth with radiolucent periapical lesions (OR = 1.16; 95% CI = 1.07–1.26; p = 0.0004). The certainty of the literature assessment was moderate per GRADE. The ROBINS-I tool classified three studies as low risk of bias, and the fourth as moderate risk of bias. Conclusions: Moderate, quality scientific evidence indicates a weak but significant relationship between smoking and the prevalence of RPLs in RFT. Smoking can be considered a negative prognostic factor for the outcome of root canal treatment. Endodontic providers should be aware of the relationship between smoking and persistent apical periodontitis, assessed as RPLs, in RFT. Capítulo IV 146 Keywords: endodontic medicine; persistent apical periodontitis; radiolucent periapical lesion; root canal treatment outcome; root-filled teeth; smoking habits Capítulo IV 147 1. INTRODUCTION Apical periodontitis (AP) is an inflammatory reaction in the periradicular tissues, induced and maintained by bacterial infection of the root canal system [1]. The prevalence of AP is 0.6–20% for teeth [2,3]. AP is radiographically diagnosed by a disruption of the lamina dura and a radiolucent area encircling the root apex, namely, a radiolucent periapical lesion (RPL) [4]. Teeth with AP, when restorable, should be treated with root canal treatment (RCT) [5]. A key goal of RCT is to seal the apical third of the root canal, interrupting the passage of bacterial antigens from the pulp space to the periapical tissues. If this is not achieved, the root-filled tooth continues to show a radiolucent image around its apex, suffers from apical periodontitis [6,7,8], and presents—to some extent and severity—periapical inflammation [9]. RFT with AP in asymptomatic patients exhibited less pronounced and relatively smaller areas of inflammation [9]. Although radiographic signs of AP are found in 25 to 61% of asymptomatic RFT [3], not in all cases imply the failure of RCT. The healing after RCT may result in the formation of fibrous tissue composed of dense collagen fibers, few cells, and little or no inflammation, which may be regarded as scar tissue [10]. Apical periodontitis is not always the result of inadequate endodontic technique (including deficient aseptic control, missed canals, inadequate instrumentation, etc.) [11,12]. Sometimes, the systemic status of the patient, such as pro-inflammatory status or impaired immune response, can restrict periapical healing [13,14]. This could explain the association between diabetes and the failure of endodontic treatment, as recently demonstrated [15,16,17]. In short, the factors involved in the development of PAP and failure of RCT are many—it is difficult to assess the role that each of them plays. Knowing the possible influence of each one on the outcome of RCT can help to improve the information given to patients regarding the prognosis of RCT. In addition, it could help explain cases of patients in whom the failure of RCT is more frequent. This could be the case with tobacco smoking. Habitual smoking, a systemic condition characterized by a pro-inflammatory status and impaired immune response and wound healing, has been associated Capítulo IV 148 with poor prognosis of periodontal disease, oral cancer, oral mucosa lesions, caries, and high failure rate of dental treatments [18,19]. Defensive and reparative responses of dental pulp are decreased in smokers [20], and tobacco smoking is a risk factor for periapical disease—AP being more prevalent in smokers [21,22,23,24,25,26], probably because of impaired bone healing [27]. A recent systematic review and meta-analysis concluded that tobacco smokers have a prevalence of periapical periodontitis and root canal treatments greater than 2.5 times the prevalence of non-smokers [28]. In addition, another study indicates that RCT is almost two times more prevalent in smokers, with a dose– response relationship [23]. Nevertheless, other studies have found no significant differences in the prevalence of AP and RCT between smokers and non-smoking subjects [13,29,30]. The possible effect of smoking on the outcome of endodontic treatment has been investigated in several epidemiological studies, with contradictory conclusions [31,32,33]. The primary objective of this study was to carry out a systematic review and meta-analysis investigating the possible association between smoking habits and the failure of RCT, the primary outcome measure being the prevalence of RPLs in RFT. 2. METHODS The protocol of this systematic review has been developed and registered in the PROSPERO database (PROSPERO 2020 CRD42020165279). The systematic review has been developed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) Guidelines [33]. 2.1. Review Question The clinical Population, Intervention, Comparison, and Outcome (PICO) question to be answered was as follows: in adult patients who have root filled teeth, does the presence or the absence of smoking habits affect the prevalence Capítulo IV 149 of RFT with RPLs? PICO (Population, Intervention, Comparison, and Outcome) schema for all the included studies to elaborate upon this research question were used to establish the eligibility criteria as follows: Population: adult patients having root-filled teeth. Intervention: presence of smoking habits; smoker. Comparison: absence of smoking habits; non-smoker. Outcome: prevalence of RFT with RPLs. 2.2. Inclusion and Exclusion Criteria The inclusion criteria established were: (a) epidemiological studies published from January 1980 to June 2020; (b) studies comparing smoking patients with non-smoking subjects; (c) studies including RFT; (d) studies providing data on the prevalence of RFT with RPLs, both in smoking patients and in control nonsmoking subjects. Exclusion criteria were defined as: (a) studies carried out in animals or in cell culture, and (b) studies reporting data only from smoking subjects. When there was no initial agreement among the reviewers, consensus was reached through dialogue. 2.3. Literature Search Once the PICO question was established, the search strategy was designed [34,35]. Studies located in the search were selected according inclusion and exclusion criteria, quality evaluation, and data extraction and analysis. A literature search in MEDLINE/PubMed, Scopus, Web of Science, and Wiley Online Database was achieved, using the following Mesh terms and keywords: (tobacco OR smoking OR smoker) AND (endodontics OR periapical periodontitis OR periapical diseases OR apical periodontitis OR periradicular lesion OR periapical Capítulo IV 150 radiolucency OR radiolucent periapical lesion OR root canal treatment OR root canal preparation OR root canal therapy OR root filled teeth OR endodontically treated teeth) (Box 1). Box 1. MeSH and key words combinations used for the search strategy for the Population, Intervention, Comparison, and Outcome (PICO) question: In adult patients who have root filled teeth, does the absence or presence of smoking habits affect the prevalence of root filled teeth with radiolucent periapical lesions? Capítulo IV 151 ((“tobacco”[MeSH Terms] OR “tobacco”[All Fields] OR “tobacco products”[MeSH Terms] OR (“tobacco”[All Fields] AND “products”[All Fields]) OR “tobacco products”[All Fields]) OR (“smoking”[MeSH Terms] OR “smoking”[All Fields]) OR (“smokers”[MeSH Terms] OR “smokers”[All Fields] OR “smoker”[All Fields])) AND ((“endodontics”[MeSH Terms] OR “endodontics”[All Fields]) OR (“periapical periodontitis”[MeSH Terms] OR (“periapical”[All Fields] AND “periodontitis”[All Fields]) OR “periapical periodontitis”[All Fields]) OR (“periapical diseases”[MeSH Terms] OR (“periapical”[All Fields] AND “diseases”[All Fields]) OR “periapical diseases”[All Fields]) OR (“periapical periodontitis”[MeSH Terms] OR (“periapical”[All Fields] AND “periodontitis”[All Fields]) OR “periapical periodontitis”[All Fields] OR (“apical”[All Fields] AND “periodontitis”[All Fields]) OR “apical periodontitis”[All Fields]) OR (Periradicular[All Fields] AND Lesion[All Fields]) OR (Periapical[All Fields] AND Radiolucency[All Fields]) OR (Radiolucent[All Fields] AND Periapical[All Fields] AND Lesion[All Fields]) OR ((“dental pulp cavity”[MeSH Terms] OR (“dental”[All Fields] AND “pulp”[All Fields] AND “cavity”[All Fields]) OR “dental pulp cavity”[All Fields] OR (“root”[All Fields] AND “canal”[All Fields]) OR “root canal”[All Fields]) AND (“therapy”[Subheading] OR “therapy”[All Fields] OR “treatment”[All Fields] OR “therapeutics”[MeSH Terms] OR “therapeutics”[All Fields])) OR (“root canal preparation”[MeSH Terms] OR (“root”[All Fields] AND “canal”[All Fields] AND “preparation”[All Fields]) OR “root canal preparation”[All Fields]) OR (“root canal therapy”[MeSH Terms] OR (“root”[All Fields] AND “canal”[All Fields] AND “therapy”[All Fields]) OR “root canal therapy”[All Fields]) OR ((“plant roots”[MeSH Terms] OR (“plant”[All Fields] AND “roots”[All Fields]) OR “plant roots”[All Fields] OR “root”[All Fields]) AND Filled[All Fields] AND (“tooth”[MeSH Terms] OR “tooth”[All Fields] OR “teeth”[All Fields])) OR (“tooth, nonvital”[MeSH Terms] OR (“tooth”[All Fields] AND “nonvital”[All Fields]) OR “nonvital tooth”[All Fields] OR (“endodontically”[All Fields] AND “treated”[All Fields] AND “teeth”[All Fields]) OR “endodontically treated teeth”[All Fields])) Capítulo IV 152 A hand-search was also carried out in main endodontic journals (International Endodontic Journal, Journal of Endodontics, and Australian Endodontic Journal) and in the references of significant papers and reviews. The last search was made in June of 2020. Electronic and manual searches provided the titles and abstracts of articles related to the aims of the studies, which were categorized by three independent researchers (D.C.-B., M.C.J.-S., and J.J.S.-E.) according to the inclusion and exclusion criteria. Articles selected were reviewed in full by five investigators (D.C.-B., J.M.-G., E.V.O., M.C.J.-S., and J.J.S.-E.). 2.4. Data Extraction The methodology of selected studies was examined and main features were extracted and compiled, including: authors, date of publication, study design, subjects and sample size, main quantitative results and odds ratio values, and diagnoses of RPLs. Data extraction was performed by seven investigators (D.C.- B., J.M.-G., M.C.J.-S., E.V.O., B.S.-D., P.M.-M., and J.J.S.-E.). Disagreements were resolved by discussion among the six and reaching an agreement by majority. 2.5. Outcome Variables and Statistical Analysis The primary outcome measure was the prevalence of RFT with RPL. Odds ratio (OR), with its 95% confidence interval (CI), was calculated in every selected study trying to measure the effect of the relationship between smoking habits and the outcome of RCT. A random-effect model meta-analysis, on the basis of the DerSimonian–Laird method, was performed to determine the pooled OR and its 95% CI. To determine the heterogeneity amongst trials, the Breslow–Day test (BDT) and the Higgins I2 test were employed, taking into account that substantial heterogeneity is considered if I2 test is higher than 50% [36]. To illustrate the homogeneity, L’Abbé plots [37] were used. To show the OR results, a forest plot Capítulo IV 153 [38] was used, along with the DerSimonian–Laird pooled estimate. Finally, a level of p = 0.05 was considered significant. The meta-analyses were calculated with the StatsDirects software (London, UK) [39]. 2.6. Quality Evidence Assessment and Risk of Bias in Individual Studies Quality evidence assessment and risk of bias in individual studies. The quality of evidence of the included studies was analyzed according to the guidelines provided by the Centre for Evidence-Based Medicine at Oxford [40]. The certainty in the evidence was assessed using the GRADE tool (GRADEpro GDT: GRADEpro Guideline Development Tool (Software)) available from gradepro.org: https://gdt.gradepro.org/app/handbook/handbook.html#h.rkkjpmwb6m6z [41]. The GRADE tool has five domains: risk of bias, inconsistency, imprecision, indirectness, and publication bias, which can be downgraded and reduce the quality of the evidence [42]. Articles were assessed independently by 5 reviewers (J.J.S.E., J.M.G., D.C.B., E.V.O., and M.C.J.S.) and cases of disagreements in the risk of bias were discussed until a consensus was achieved. The risk of bias of the included studies was assessed according to Cochrane Collaboration common scheme for bias and ROBINS-I tool [43], initially described to assess nonrandomized studies of interventions, but currently also available for observational designs (https://methods.cochrane.org/robins-i-tool). 3. RESULTS The search strategy is presented in Figure 1. After searching databases and hand-searching relevant bibliographies/papers, 1075 articles were recovered. Excluding duplicates articles (n = 733) and publications before 1980 (n = 3), 339 articles were checked to satisfy the selection criteria by title and abstract, declaring 14 articles for full text review. Among these, ten articles were excluded for the following reasons: four did not deal with the specific topic [25,26,44,45],