Evaluation of the role of C-terminal binding proteins (CtBP) in the neuroimmune response

• Autor(a) principal: Massano, Mariana Gomes
• Data de Publicação: 2023
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.6/13879
• topic: Ácido Retinóico; Lipopolissacarídeo; Mtob; Neuroinflamação; Proteínas de Ligação C-Terminal; Domínio/Área Científica::Engenharia e Tecnologia::Bioquímica

Neuroinflammation is an inflammatory response that occurs at the level of the central nervous system (CNS). Various stimuli or injuries can trigger this response, such as ischemia, trauma, infections or exposure to toxins. Microglial cells, considered innate immune cells residing in the CNS, contribute to the neuroinflammatory response. These cells detect changes in brain homeostasis, migrate to the injured site and release inflammatory mediators capable of impacting neuronal survival. Lipopolysaccharide (LPS), one of the components of the outer membrane of gram-negative bacteria, is widely used as a model to understand the pathological mechanisms involved in neurodegeneration induced by neuroinflammation, as well as to identify potential therapeutic molecules. C-terminal binding proteins (CtBPs) are transcriptional coregulators that interact with transcription factors and repress transcription. These proteins are highly expressed in the CNS during embryonic development and regulate neuronal development, survival and function. However, knowledge about the regulation of the expression and function of CtBPs in conditions of neuroinflammation induced by LPS is still scarce. The first objective of this work was to evaluate the expression of CtBPs in in vivo and ex vivo experimental models of neuroinflammation induced by LPS and, subsequently, to evaluate their function in an ex vivo model. The expression of CtBPs was evaluated by Western-Blot analysis of hippocampi from adult C57BL/6J mice and in organotypic hippocampal slice cultures from 6- to 9-day-old C57BL/6J mice. It was concluded that the concentration of 500 µg/kg of LPS was the most effective in inducing an increase in the expression of CtBP1 and CtBP2 when the animals were exposed for 4 days. In mice exposed to the same concentration of LPS for 7 days, there was an increase in CtBP2 expression. On the other hand, exposure to 2 mg/kg of LPS for 24h induced as increase in CtBP1 expression. LPS, at different concentrations, also induced increased expression of CtBPs in organotypic hippocampal cultures. Next, we evaluated the glial response, in organotypic hippocampal slices obtained from C57BL/6J mice by Western-Blot analysis against glial fibrillary acidic protein (GFAP) and neutrophil cytosolic factor 1 (P47phox). To evaluate the involvement of CtBPs in the neuroinflammatory response, cultures were exposed to 4-methylthio-2-oxobutyric acid (MTOB), a modulator of CtBP activity that can act either as an inhibitor at high concentrations or an agonist at low concentrations. In some experiments, retinoic acid (RA) was also added as a potent anti-inflammatory agent. We observed that P47phox expression was reduced in slices exposed to LPS and/or MTOB and that RA was able to counteract this effect. The decreased expression of this phagocytic cell marker suggests implications for the inflammatory response of these cells. On the other hand, the increase in expression of this marker to levels similar to the control induced by RA may indicate an attempt to attenuate the inflammatory response. Regarding the analysis of GFAP expression, a decrease in its expression was observed in all experimental groups, suggesting that these compounds may be affecting the survival of astrocytes. In short, we can conclude that neuroinflammation induced by LPS triggers the expression of CtBPs in the hippocampus depending on the dose and time of exposure to the inflammatory agent and that RA can attenuate the effects induced by LPS+MTOB and can be considered a potential anti-inflammatory agent.