Participation of pre-transcriptional and post-translational mechanisms in hypothalamic reduction of a7nAChR receptor expression by consumption of hyperlipidic diet

• Grant number: 19/09420-9
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): November 01, 2019
• Effective date (End): August 31, 2020
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Marcio Alberto Torsoni
• Grantee: Ísis de Cássia Alves Martins
• Host Institution: Faculdade de Ciências Aplicadas (FCA). Universidade Estadual de Campinas (UNICAMP). Limeira , SP, Brazil

Abstract

The parasympathetic nervous system is a fundamental regulator of the intensity of the inflammatory response, with acetylcholine as the main neurotransmitter. This mechanism is called the cholinergic anti-inflammatory pathway, which promotes the release of acetylcholine by the nerve terminal that activates nicotinic cholinergic receptors (a7nAChR) in target cells. Activation of these receptors reduces the production of proinflammatory cytokines in different cell types. Data from our group showed that animals that consumed a diet rich in saturated fats for a short period of time presented an impairment of the cholinergic anti-inflammatory response with reduction of a7nAChR receptor expression in the hypothalamus. The ubiquitin-proteasome system (UPS) constitutes an intracellular pathway of protein degradation, being the main post-translational mechanism. Evidence indicates that ubiquitin ligases may indirectly regulate insulin signaling molecules, targeting proinflammatory markers and that their deletion can prevent obesity induced by the hyperlipidic diet. Methylation, as a pre-transcriptional mechanism, consists of the addition of a methyl group to the DNA that leads to the recruitment of proteins that cause chromatin compaction, preventing gene transcription. Studies have shown that short exposure to the hyperlipic diet has introduced widespread changes in DNA methylation, mainly affecting genes involved in inflammation. Thus, we hypothesized that the short exposure to the hyperlipidic diet may activate early pre-transcriptional and post-translational mechanisms, leading to the reduction of the a7nAChR receptor. This hypothesis will be tested in vitro, using lipopolysaccharide (LPS)-stimulated murine microglial cell line BV-2, and in vivo, by exposing C57 mice to the hyperlipid diet for 3 days. The expression of the a7nAChR receptor, the activity of the ubiquitin-proteasome system, the pharmacological blocking effect of the ubiquitin-proteasome pathway, the general methylation profile, and DNA-methylase levels will be analyzed. Expression analyzes will be performed by real-time PCR, Western blotting, immunoprecipitation, and ELISA.