Navegando por Autor "Salles, Geisa Rodrigues"
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Item 2D and 3D Models of Alzheimer’s Disease: Investigating Neuron-like Cells in Oxidative Environments(ACS Publications) Salles, Geisa Rodrigues; Giraldi, Luiza de Andrade; Silva, Newton Soares da; Porcionatto, Marimelia Aparecida; Soares, Cristina PachecoAlzheimer’s disease (AD) is a complex and enigmatic neurodegenerative disorder in which amyloid-β (Aβ) aggregates and oxidative stress play crucial roles in neuronal damage. Aβ forms senile plaques, while reactive oxygen species (ROS)-induced oxidative stress causes cellular dysfunction. Elucidating neuronal injury led by mild and severe oxidative stress may provide insight into how neurons respond to toxic environments. In parallel, modeling AD three-dimensionally is in the spotlight, sustainably contributing to reducing animals in research and replicating spatially neuronal mechanisms, such as neurite network and oxidative stress responses. This study evaluates the effects of oxidative stress on neuron-like cells cultured in two-dimensional (2D) and 3D spheroids, strengthening their potential as platforms for AD investigation. For the 2D models, SH-SY5Y (cells from human neuroblastoma) cells were differentiated into the neuronal phenotype and exposed to mild or severe concentrations of oxygen peroxide (H2O2, 100 or 200 μM, respectively). Cytoviability, ROS, Aβ particle analyses, and morphological aspects were assessed. Neuronal cells under severe stress produced elevated levels of intra- and extracellular Aβ aggregates. A range of Aβ particle analyses were performed comparing their properties, and morphologically, neurites were compromised under severe stress. For the 3D models, SH-SY5Y spheroids were self-assembled by 10 days of cultivation on developed nonadhesive hydrogel microwells and differentiated into the neuronal phenotype; their area, circularity, and solidity were measured. Spheroids were exposed or not to 200 μM H2O2, stained for cytoskeleton/nuclei, and imaged by scanning electron microscopy (SEM), and their viability was evaluated. Throughout the cultivation period, spheroids grew and differentiated morphologically. Neurite distribution was observed along the 3D composition; however, under oxidative stress, cytoviability decreased, abnormal nuclear staining was observed surrounding the spheroids, and morphological disorganization was evident by SEM, representing structural disarrangement and nuclear disruption. Briefly, this study provides a basis for exploring oxidative stress and producing robust 3D approaches to unraveling AD mechanisms.Item 3D bioprinted human iPSC-derived neural progenitor cells as a novel platform for studying neurogenic niche(AIP Publishing) Machado, Lucas Simões; Ferreira, Paula Scanavez; Pires, Marina Rodrigues; Bim, Larissa Valdemarin; Oliveira, Natália Heloísa de; Salles, Geisa Rodrigues; Ferreira, Natalia Dall'Agnol; Cruz, Elisa Marozzi; Porcionatto, Marimelia AparecidaAnimal models, especially rodents, used to study neurodevelopment have significantly advanced our comprehension of cellular and molecular mechanisms. Nevertheless, differences in species-specific structures, gestation periods, and interneuronal connections limit animal models’ ability to represent human neurodevelopment accurately. The unique characteristics of primate neural progenitor cells (NPCs) enable cortex expansion with gyrus formation, which does not occur in lissencephalic animals, like rodents. Therefore, there is a need for novel in vitro models using human cells that recapitulate the complexity of human brain development. Along with organoids, 3D bioprinting offers a platform for creating more complex in vitro models. We developed, extensively characterized, and successfully used a GeltrexTM/GelMA hydrogel blend to bioprint human induced pluripotent stem cells-derived NPCs (hNPCs). We show that 3D bioprinted hNPCs can selforganize, revealing key features of a neurogenic niche, including proliferation, differentiation, and migration, remaining viable for over 110 days. Within the first 20 days, bioprinted constructs showed the formation of positive cell clusters for the neurogenic niche cell markers FABP7, NESTIN, and GFAP. Clusters were interconnected by process bundles supporting cell migration. The cells proliferated within the clusters, and over time, NPCs originated TUBB3þ neurons with long axonal tracts, prominent around the clusters. We propose this as a 4D model to study neurogenic niches’ key cellular and molecular features in a 3D bioprinted scaffold, adding time as the fourth dimension. Neuronal maturation in this dynamic model recapitulates key neurogenic niche properties, making it suitable for neurodevelopmental disease modeling and drug screening.Item 3D-bioprinted model of adult neural stem cell microenvironment in Alzheimer’s disease(ACCSCIENCE Publishing) Ferreira, Natalia Dall’Agnol; Ferreira, Paula Scanavez; Soares, Cristina Pacheco; Porcionatto, Marimelia Aparecida; Salles, Geisa RodriguesNeurogenesis plays a major role in neuroplasticity and memory. In adult human and mouse brains, neural stem cells (NSCs) are mainly distributed in two extensively characterized neurogenic niches: the subgranular zone (SGZ) of the hippocampus and the subventricular zone (SVZ) of the lateral ventricles. Impaired neurogenesis is one of the consequences of Alzheimer’s disease (AD), contributing to cognitive decline and progressive memory loss. Developing new in vitro models that resemble this three-dimensional (3D) structure is fundamental for enhancing our understanding of the SVZ neurogenic niche dynamics in AD. Herein, we produced and characterized a 3D-bioprinted model of the adult SVZ neurogenic niche containing amyloid β (Aβ) oligomers, mimicking the NSC microenvironment in AD. In this model, Aβ oligomers induce oxidative stress and reduce the proliferative potential of NSCs, while stimulating neuronal differentiation. We hypothesize that these events are an early attempt of adult NSCs to compensate for neuronal death in AD pathogenesis. Our 3D model simulates the NSC niche physiology, reproducing an early response of NSCs in AD, strengthening the importance of studying the potential of neurogenesis in neurodegeneration.Item Photobiomodulation by LED 660 nm and Taurine against H2O2 oxidative stress in SH-SY5Y cells(Springer Nature Link) Rossato, Rafaella Carvalho; Salles, Geisa Rodrigues; Albuquerque, Amanda Lira; Porcionatto, Marimélia Aparecida; Granato, Alessandro Eustáquio Campos; Ulrich, Henning; Santos, Mariela Inês Batista dos; Soares, Cristina PachecoAlzheimer's Disease (AD) is a progressive uncurable neurodegenerative pathology affecting millions worldwide. Photo- biomodulation and Taurine are promising alternatives for preventing and reducing the rapid progression of neurodegenera- tion, stimulating the reconstructing of neural tissue structures, especially improving mitochondrial activity, which is highly impaired in AD. In this study, the mitochondrial effects of Taurine combined with light emitting diode (LED) irradiation were evaluated on human neuroblastoma cells (SH-SY5Y), under oxidative stress condition by hydrogen peroxide (H2O2) exposure, a considerable modulator in AD. We evaluated LED irradiation at the wavelength of 660 nm and Taurine under different concentrations before and together with exposing SH-SY5Y cells to different concentrations of H2O2, assessing mitochondrial activity by the MTT colorimetric test and labeling live cells mitochondria by the fluorescent probe MitoTracker. Cell viability was also evaluated by the trypan blue exclusion assay, and cellular morphological structures were imaged by scanning electron microscopy (SEM). Neuroprotective effects were achieved by both LED irradiation and LED irradia- tion + Taurine when cells were exposed to them before H2O2-induced stress. Comparing both agents, LED irradiation at 660 nm is sufficient to improve mitochondrial activity, however, healthy mitochondrial morphology was only observed when cells were treated with Taurine together with LED irradiation, representing affordable candidates that act in synergy against oxidative stress, one of the main contributors to neurodegeneration.Item Self-assembly and 3D Bioprinting of Neurospheres and Evaluation of Caffeine and Photobiomodulation Effects in an Alzheimer's Disease In Vitro Model(Springer Nature Link) Salles, Geisa Rodrigues; Granato, Alessandro Eustáquio Campos; Viero, Fernanda Tibolla; Soares, Cristina Pacheco; Ferreira, Sergio Teixeira; Porcionatto, Marimélia Aparecida; Ulrich, HenningSeveral in vitro models of Alzheimer’s disease (AD) rely on 2D cell culture, and, more recently, 3D cultures represented by free-floating neurospheres have been used as models for the disease. The advantage of 3D over 2D cell culture is that cell-extracellular matrix and cell-cell interactions can be assessed, better representing the molecular and cellular hallmarks of the disease. In the current study, we developed two complementary 3D neurosphere models using SH-SY5Y human neuroblastoma cells to investigate AD pathology and evaluate potential therapies. First, self-assembled neurospheres were exposed to hydrogen peroxide (H2O2) and amyloid-beta oligomers (AβOs), inducing AD-like features such as increased production of reactive oxygen species (ROS), amyloid aggregation, and apoptosis. Treatment with caffeine or photo- biomodulation (PBM) using LED irradiation significantly reduced Aβ1−42 accumulation, ROS generation, and decreased apoptosis markers. Second, 3D bioprinting of SH-SY5Y cells resulted in neurospheres with enhanced cellular organization and differentiation. These findings emphasize the advantages of 3D models for studying neurodegeneration and evaluating therapeutic strategies, bridging the gap between traditional 2D cultures and complex in vitro systems.