3.02 Facultad de Medicina
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Browsing 3.02 Facultad de Medicina by Author "Andía Kohnenkampf, Marcelo Edgardo"
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- ItemNanoparticle use for the study of exosome transport to the brain through the lymphatic pathway(2022) Ramos Zaldívar, Héctor M.; Andía Kohnenkampf, Marcelo Edgardo; Pontificia Universidad Católica de Chile. Escuela de MedicinaIntroduction: Exosomes are extracellular vesicles with a size of 50-150 nm that have been associated with the transportation of various biological contents and with intercellular communication. Given their role in metastasis, understanding exosome tissue distribution is critical to cancer pathophysiology. The exact routes and mechanisms of exosome distribution from peripheral organs to the central nervous system (CNS) remain unknown. A possible route is through the recently discovered brain lymphatic system, due to its connection with the deep cervical lymph nodes and its morphological characteristics. Hypothesis: Metastatic cell-derived exosomes are transported from the deep cervical lymph nodes to the central nervous system through the meningeal lymphatic vessels. Objective: To develop nanoparticle-loaded exosomes derived from a metastatic cell line and administer these exosomes via the cervical and meningeal lymphatic system to evaluate their arrival to the central nervous system. Methodology: Superparamagnetic iron oxide nanoparticles (SPIONs) (mean size 8.3 ± 2.9 nm and Zeta potential 36.8 ± 5.44 mV) were prepared by chemical coprecipitation of ferric and ferrous chlorides. Exosomes (41.77 ± 1.64 nm and -10.8 ± 2.49 mV) were isolated from the melanoma B16F10 cell line through the Exo-Spin column protocol and loaded with SPIONs through electroporation. Gold nanorods (11.25 ± 0.57 nm and 45.4 ± 7.62 mV) were prepared and functionalized with polyethylene glycol. Chinese ink nanoparticles (61.62 ± 4.84 nm and -6.34 ± 0.63 mV) were also used. C57BL/6 mice were used to evaluate the anterograde and retrograde route of the lymphatic meningeal system with post-mortem and in-vivo procedures. All animal procedures were approved by the Ethical Animal Committee of our institution. Mice were anesthetized with isoflurane. To evaluate the anterograde nanoparticle flow we injected 10 µL of each nanoparticle solution in the cisterna magna (3 animals per condition). To evaluate the retrograde nanoparticle flow we injected 10 µl of each nanoparticle solution (SPIONs 3200 μg/mL; exosomes + SPIONs 1.67 x 1011 particles/mL; gold nanorods 1.71 x 1014 particles/mL; Chinese ink 10%) in the deep cervical lymph node (3 animals per condition). The animals were euthanized after 30 min post injection. The head and neck were fixed with 4% paraformaldehyde for histological analysis and post-mortem MRI imaging. Results: Anterograde pathway: Both SPIONs and SPION-loaded exosomes showed hypointense signals of cervical lymphatic structures after intracerebroventricular injections through the cisterna magna in the T2w and T2* MRI images. Gold Enhancement technique confirmed anterograde flow of both gold nanorods and Chinese ink nanoparticles by cervical lymphatic staining. Macroscopically, cisterna magna injections showed staining of deep cervical lymph nodes within the first minute after the administration of Evans Blue dye and Chinese ink. Retrograde pathway: Both SPIONs and SPION-loaded exosomes revealed hypointense signals in the brain ventricles and parenchyma in MRI T2w image and T2* map, after 30 min of deep cervical lymphatic injection. Gold Enhancement staining showed histological confirmation of the arrival of gold nanorods and Chinese ink nanoparticles to the brain parenchyma from the cervical injections. Macroscopically, deep cervical lymph node injections with Evans Blue and Chinese ink showed staining of the meninges and brain parenchyma. Nanoparticles colocalized with the stain of meningeal lymphatic vessels using anti-LYVE-1. Discussion: The cervical and meningeal lymphatic system can transport nanoparticles not only in the classically described lymphatic drainage towards the thorax but can also serve as an access gate to the brain. This newly discovered mechanism for the meningeal lymphatic pathway could be exploited in the theranostic field of nanomedicine to deliver drugs for the treatment of various neurological diseases and the developing of diagnostic contrast media. The understanding of cancer exosome distribution through the cervical and meningeal lymphatic system will aid in a more profound comprehension of brain metastasis pathophysiology.
- ItemSex differences in the relationship between body composition and metabolic dysfunction-associated steatosis liver disease progression in a murine model of metabolic syndrome(2025) Manjarrés Madrid, Laura; Andía Kohnenkampf, Marcelo Edgardo; Pontificia Universidad Católica de Chile. Escuela de MedicinaLa progresión de la esteatosis hepática asociada a disfunción metabólica (MASLD) varía significativamente entre sexos, y los hombres generalmente presentan una enfermedad más avanzada. Utilizando un modelo murino knock out de óxido nítrico sintasa (eNOS KO), analizamos las diferencias específicas de sexo en la progresión de MASLD durante una intervención de dieta occidental. Se empleó la resonancia magnética (MRI) 3T para evaluar la composición corporal y la fracción de grasa hepática, revelando mayor grasa visceral, volumen hepático y proporciones de grasa hepática a muscular en los hombres. La reducción de la dimensionalidad y los análisis de agrupamiento destacaron distintos fenotipos y patrones de progresión de MASLD específicos de sexo. Las evaluaciones histológicas confirmaron un mayor daño hepático en los hombres, como lo indican los puntajes de actividad MASLD más altos. Estos hallazgos demuestran la importancia del sexo como variable biológica en la patología MASLD, enfatizando el papel de la composición corporal y la distribución de la grasa en la progresión de la enfermedad. El estudio destaca el potencial de los métodos analíticos y de imágenes avanzados para refinar los diagnósticos no invasivos e informar las intervenciones específicas según el sexo para la MASLD, contribuyendo al desarrollo de estrategias de tratamiento personalizadas.Con esta tesis se está optando al magister en Investigación en Ciencias de la Salud.
- ItemSex-dependent differences on the impact of anti-inflammatory treatment in the progression of coronary artery disease in a murine model of lethal ischemic heart disease induced by diet(2023) Parra Núñez, Laura Macarena; Andía Kohnenkampf, Marcelo Edgardo; González, Leticia; Pontificia Universidad Católica de Chile. Facultad de MedicinaCardiovascular risk differs significantly between adult men and women. Therefore, it is expected that different treatments may affect both groups differently. We aim to compare sexdependent differences on survival and systemic inflammation in response to anti-inflammatory treatment using a diet-induced myocardial infarction mouse model. Method. Male and female SR-B1−/−ApoER61h/h mice, aged 2-3 months, were randomly assigned into two groups: Control (HFD-Control) and minocycline (HFD-MIN). Atherosclerosis was induced by feeding an atherogenic diet (15% fat, 1.25% cholesterol, 0.5% cholate). Minocycline was administered in the drinking water at a dose of 0.05 mg/mL. Female mice had a slightly better survival than male mice when fed an HFD (p=0.12). Minocycline improved survival in male by 35% (p=0.006) and by 33% in female p=0.01), without affecting total cholesterol levels. Male mice fed with HFD tended to have higher IL-6 levels than female mice (p=0.08). Minocycline significantly reduced IL-6 levels (p=0.04) and Ly6Chigh (p=0.006) and increased the Ly6Clow subset (p=0.006). Male and female fed with HFD clustered in different groups by analyzing inflammatory parameters by PCA; however, after minocycline intervention, were indistinguishable. High fat diet decreased survival and caused early death in this animal model, however, females had slightly better survival than male mice. Minocycline treatment improved survival in both groups although it did not affect their cholesterol levels. Males showed higher inflammatory serum biomarkers than females, and minocycline treatment showed a higher impact on systemic anti-inflammation in male mice than female, by reducing plasma IL-6 levels and shifting toward a more "reparative" phenotype on circulating monocyte subsets.