Publication: From Clinical Inflammatory Profiling to Translational Experimental Modeling of Endometriosis : Peritoneal Macrophages as a Therapeutic Target
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Date
2025-10-13
Authors
Ramírez Pavez, Tamara Nadira
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Escuelas::Escuela Internacional de Doctorado
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Martínez-Esparza Alvargonzález, María Concepción ; Machado Linde, Francisco
Publisher
Universidad de Murcia
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DOI
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info:eu-repo/semantics/doctoralThesis
Description
Abstract
Objetivos
Esta tesis tiene como objetivo profundizar en la comprensión del entorno inmunológico y molecular de la endometriosis mediante un enfoque experimental multifacético, que integra modelos clínicos, inmunológicos y preclínicos. Los objetivos específicos incluyeron: optimizar protocolos de recogida de muestras peritoneales; caracterizar el microambiente inmunológico, en particular los macrófagos y mediadores solubles; correlacionar estos hallazgos con parámetros clínicos; e implementar un modelo murino de endometriosis para estudios in vivo.
Metodología
Se reclutaron 322 mujeres entre 2014 y 2025, de las cuales 197 fueron confirmadas con endometriosis mediante histología y 72 eran mujeres sanas. Se diseñó un protocolo estandarizado para la recogida de líquido y lavados peritoneales, junto con biopsias quirúrgicas. Las poblaciones de macrófagos peritoneales se caracterizaron mediante citometría de flujo y marcadores fenotípicos asociados a activación M1/M2, reparación tisular y otras subpoblaciones. Se midieron mediadores inflamatorios solubles humanos y microbianos en líquido peritoneal y plasma mediante ELISA. Por último, se desarrolló un modelo murino homólogo de endometriosis con implante de tejido endometrial GFP en ratones C57BL/6, evaluando la formación de lesiones, cambios de conducta tipo ansiedad y persistencia celular.
Conclusiones
Se optimizaron los protocolos de recogida de muestras, mejorando la recuperación celular en lavados peritoneales, especialmente en mujeres con obesidad.
Se observó una infiltración generalizada de macrófagos, incluso en tejidos sin lesiones visibles, con una expansión significativa de subpoblaciones no-clásicas y un fenotipo complejo de marcadores proinflamatorios, reguladores y reparativos.
A nivel sistémico, se detectaron alteraciones en leucocitos y citocinas, incluyendo elevación de galectina-1 y descenso de sCD163, así como un incremento moderado de β-glucanos.
El índice de masa corporal se asoció con mediadores inflamatorios y mayor severidad de la enfermedad, subrayando el papel del peso en su progresión.
El análisis detallado de subpoblaciones de macrófagos reveló correlaciones con la percepción de salud física y posibles vínculos con síntomas.
Se identificaron asociaciones entre marcadores tumorales (CEA, HE4) y perfiles inmunológicos, lo que resalta su potencial como biomarcadores sensibles.
El modelo murino homólogo aportó información útil aunque con limitaciones en la formación y detección de lesiones, destacando la necesidad de perfeccionar los sistemas preclínicos.
Objectives This thesis aims to deepen the understanding of the immunological and molecular environment of endometriosis through a multifaceted experimental approach integrating clinical, immunological, and preclinical models. The specific objectives included: optimizing protocols for peritoneal sample collection; characterizing the immune microenvironment, particularly macrophages and soluble mediators; correlating these findings with clinical parameters; and implementing a murine model of endometriosis for in vivo studies. Methodology A total of 322 women were recruited between 2014 and 2025, of whom 197 were histologically confirmed with endometriosis and 72 were healthy controls. A standardized protocol was developed for collecting peritoneal fluid and washings, along with surgical biopsies. Peritoneal macrophage populations were characterized by flow cytometry using phenotypic markers associated with M1/M2 activation, tissue repair, and other immune subsets. Human and microbial soluble inflammatory mediators were measured in peritoneal fluid and plasma using ELISA. Finally, a homologous murine model of endometriosis was developed by implanting GFP-labeled endometrial tissue into C57BL/6 mice, evaluating lesion formation, anxiety-like behavioral changes, and cellular persistence. Conclusions Sampling protocols were optimized, improving immune cell recovery in peritoneal washings, particularly in obese women. A generalized infiltration of macrophages was observed, even in tissues without visible lesions, with a significant expansion of non-classical subsets and a complex phenotype including proinflammatory, regulatory, and reparative markers. At the systemic level, alterations in leukocytes and cytokines were detected, including increased galectin-1 and decreased sCD163, as well as a moderate rise in β-glucans. Body mass index was associated with inflammatory mediators and greater disease severity, underscoring the role of weight in disease progression. Detailed analysis of macrophage subsets revealed correlations with perceived physical health and possible links to symptoms. Associations were identified between tumor markers (CEA, HE4) and immune profiles, highlighting their potential as sensitive biomarkers. The homologous murine model provided useful insights but showed limitations in lesion formation and detection, emphasizing the need to refine preclinical systems.
Objectives This thesis aims to deepen the understanding of the immunological and molecular environment of endometriosis through a multifaceted experimental approach integrating clinical, immunological, and preclinical models. The specific objectives included: optimizing protocols for peritoneal sample collection; characterizing the immune microenvironment, particularly macrophages and soluble mediators; correlating these findings with clinical parameters; and implementing a murine model of endometriosis for in vivo studies. Methodology A total of 322 women were recruited between 2014 and 2025, of whom 197 were histologically confirmed with endometriosis and 72 were healthy controls. A standardized protocol was developed for collecting peritoneal fluid and washings, along with surgical biopsies. Peritoneal macrophage populations were characterized by flow cytometry using phenotypic markers associated with M1/M2 activation, tissue repair, and other immune subsets. Human and microbial soluble inflammatory mediators were measured in peritoneal fluid and plasma using ELISA. Finally, a homologous murine model of endometriosis was developed by implanting GFP-labeled endometrial tissue into C57BL/6 mice, evaluating lesion formation, anxiety-like behavioral changes, and cellular persistence. Conclusions Sampling protocols were optimized, improving immune cell recovery in peritoneal washings, particularly in obese women. A generalized infiltration of macrophages was observed, even in tissues without visible lesions, with a significant expansion of non-classical subsets and a complex phenotype including proinflammatory, regulatory, and reparative markers. At the systemic level, alterations in leukocytes and cytokines were detected, including increased galectin-1 and decreased sCD163, as well as a moderate rise in β-glucans. Body mass index was associated with inflammatory mediators and greater disease severity, underscoring the role of weight in disease progression. Detailed analysis of macrophage subsets revealed correlations with perceived physical health and possible links to symptoms. Associations were identified between tumor markers (CEA, HE4) and immune profiles, highlighting their potential as sensitive biomarkers. The homologous murine model provided useful insights but showed limitations in lesion formation and detection, emphasizing the need to refine preclinical systems.
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