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| Author(s): |
Monique Culturato Padilha Mendonça
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Médicas |
| Defense date: | 2016-06-23 |
| Examining board members: |
Maria Alice da Cruz Hofling;
Frank Denis Torres Huaco;
Alexandre Leite Rodrigues de Oliveira;
Patricia da Silva Melo;
Leonardo Fernandes Fraceto
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| Advisor: | Maria Alice da Cruz Hofling |
| Abstract | |
The blood brain barrier (BBB), located on the blood-brain interface, is a dynamic structure destined to maintain the homeostasis of central nervous system (CNS) through the highly restrictive control afforded by physical and molecular barriers from paracellular and transcellular pathway. However, BBB also restricts the entry of important therapeutic agents for the treatment of CNS disorders. Among the approaches used to overcome this problem, the use of nanomaterials is particularly promising. Carbon-based nanomaterials arouse great interest due to their extraordinary physicochemical properties and morphological and functional similarities shared between the (sub)cellular organisation of the CNS and carbon-based nanomaterials. In the first part of the project, we analyze (i) the ability of reduced graphene oxide (rGO) to modulate the permeability of BBB in adults Wistar rats, (ii) the possible mechanisms involved in this process, (iii) the toxicity of this nanomaterial 15 minutes, 1 hour, 3 hours and 7 days after a single intravenous administration (7 mg/kg). rGO were mainly concentrated in thalamus and hippocampus, with peak of distribution occurring at 3 hours, as detected by mass spectroscopy imaging. Systemic presence of rGO induced transient opening of the BBB in the hippocampus of animals as shown at anatomical (Evans blue dye infusion), subcellular (transmission electron microscopy) and molecular (expression of paracellular ans transcellular pathway proteins) levels. The rGO-treated rats were clinically indistinguishable from controls animals injected with vehicle (distilled water). Hematological, histopathological (neurons and astrocytes markers), biochemical (nephrotoxicity and hepatotoxicity assessment) and genotoxicological based tests showed that systemic rGO injection seemed to produce minimal toxicological effects at the time points assessed. Regarding the control group, rGO-treated animals exhibited reduction in blood urea nitrogen level 3 hours post-treatment and increases in superoxide dismutase activity 1 hour and 7 days post-treatment. Intra-treated groups alterations involved leukocytosis (rGO-1 hour vs. rGO-3 hours) and leukopenia (rGO-3 hours vs. rGO-7 days), nevertheless, no inflammatory response was induced in the serum and hippocampus, the permeabilized region of BBB. In the second part of the project, in vivo and in vitro tests were carried out using rGO functionalized with polyethylene glycol (PEG 6,000). The PEGylation has been used to improve the physicochemical characteristics of nanomaterials. Using the highest concentration (100 µg/ml), non-functionalized rGO showed low toxicity whereas rGO-PEG induced deleterious effects and death in primary culture of astrocytes and rat brain endothelial cells. Corroborating the in vitro data, there was a progressive decrease in the expression of astrocytic markers (GFAP and connexin-43), tight and adherens junctions and basal lamina (occludin, ?-catenin and laminin) markers after rGO-PEG injection in vivo, indicating BBB disruption. The formation of intracellular ROS in vitro and the increase in the enzymatic antioxidant system in vivo induced by PEGylated rGO indicated oxidative stress-mediated damage. We concluded that within the experimental conditions used rGO, but not rGO-PEG, could be a promising tool to be tested as a carrier for delivery of drugs that have difficult access to the CNS (AU) | |
| FAPESP's process: | 12/24782-5 - Graphene oxide and central nervous system: evaluation of effects on blood brain barrier and nanotoxicological profile |
| Grantee: | Monique Culturato Padilha Mendonça |
| Support Opportunities: | Scholarships in Brazil - Doctorate |