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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.
- ItemSpatial biology of Ising-like synthetic genetic networks(2021) Simpson Alfaro, Kevin Matías; Federici, Fernán; Keymer, Juan E.; Pontificia Universidad Católica de Chile. Facultad de Ciencias BiológicasUnderstanding how spatially-correlated cellular states emerge from the local interaction of gene network dynamics is a fundamental challenge in biology. Short and long-range correlations and anti-correlations in gene expression can be found in spatially-distributed cellular systems such as eukaryotic tissues and microbial communities. However, the study of gene spatial correlations emerging from cell-cell coupling in natural systems is difficult since complex interactions are the norm. An alternative is to generate synthetic genetic networks (SGNs) that capture essential features of cell-cell interactions and reveal their influence in the emergence of cellular state patterns. Here, we combine synthetic biology, theoretical modelling and computational simulations to study the emergence of macroscopic gene correlations and address possible mechanisms for multi-scale self-organization of gene states in bacteria. We applied the Ising model as a theoretical framework to study the self-organization of spatially-correlated gene expression in two-state SGNs that are coupled by short-range chemical signals in E. coli. Inspired by the Ising model, we name these SGNs ferromagnetic or anti-ferromagnetic depending if they stabilize the same or the opposite state in neighboring cells. As predicted by our simulations that combine the two-dimensional Ising model with the Contact Process lattice model of cell population dynamics, these SGNs allowed the self-organization of spatial patterns of short and long-scale cellular state domains in bacterial colonies, where the size of the domains depends on the type of interaction, ferromagnetic or anti-ferromagnetic. The emergence of spatial correlations showed to be independent of the cell shape and the underpinning mechanical forces. The similarity found between ferromagnetic colonies and simulated ferromagnetic populations suggest these colonies are near the critical point of phase transition, implying that far regions in the colony are correlated. This work provides resources and a general scope theoretical framework that explain how both short and long-range correlations (and anti-correlations) are able to self-organize from locally-interacting networks. These results on multi-scale organization of gene network states shed light onto the study of pattern formation in developmental biology and microbial ecology, as well as provide a theoretical framework for the engineering of spatially-arranged cell systems.
- ItemAttentional bias for food cues after sleeve gastrectomy : a behavioral and electrophysiological study(2019) Zumaeta Valenzuela, Arturo Ulises; Aboitiz, Francisco; Pontificia Universidad Católica de Chile. Facultad de MedicinaAttentional bias for food cues (ABFC) has been linked to food craving, the inability to lose weight, and obesity. For this study, we explored the presence of ABFC in a bariatric group, using a food-modified Stroop task and visual task, with EEG and eye-tracking recording, respectively. Our data showed an increased attentional bias for appetizing food, a decreased bias for diet food and a prolonged brain processing of food stimuli in the bariatric group.
- ItemEpisodes of meeting as relevant episodes for the study of change in psychoterapy(2017) Duarte Soto, Javiera; Martínez Guzmán, Claudio; Pontificia Universidad Católica de Chile. Escuela de Psicología; Universidad de Chile. Facultad de MedicinaThe study of the change process in psychotherapy has advanced considerably during the last 30 years, focusing its efforts on the study of relevant intra-session episodes in order to contribute to a better understanding of psychotherapy and how change occurs. Although there are different conceptualizations of relevant episodes, up to the present, the different episodes developed and the conception of change that emerges from each one, have focused mainly on the patient, neglecting the dyadic and relational aspects of the therapeutic process, which emphasize that new experiences of relationship with the therapist would be the essence of change. In this line an approach to relevant episodes, which includes change in psychotherapy as a relational and interactive phenomenon between patient and therapist, is that of Daniel Stern and the Boston Group (1998, 2004) which propose moments of meeting in psychotherapy as central moments for reorganizing the patient’s implicit relational knowing. However, despite its theoretical solidity, this approach lacks empirical systematization, which hinders its study. The aim of this dissertation was to design a procedure for identifying and outlining episodes of meeting between therapist and patient during the psychotherapeutic process and to establish the main characteristics (verbal, non-verbal and relational) of the interaction between patient and therapist during its occurrence. This descriptive-relational exploratory study was based on a qualitative methodological approach, that combined grounded theory with a phenomenological approach called the micro analytic interview. Eight therapeutic dyads, thirteen therapists, nine patients and five experts were interviewed to approach this phenomenon from different perspectives and grasp its central features. This thesis is considered relevant as it is a contribution for the investigation of the psychotherapeutic process and for clinical practice.