Effects of nicotinic receptors agonist and alosteric modulators in an in vitro pulmonary cellular model stimulated with LPS: evaluation of inflammatory markers and ACE2 expression.

• Grant number: 22/03653-4
• Support Opportunities: Scholarships in Brazil - Master
• Start date: April 01, 2023
• End date: March 31, 2025
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Carla Máximo Prado
• Grantee: Juliana Tavares Correa
• Host Institution: Instituto de Saúde e Sociedade (ISS). Universidade Federal de São Paulo (UNIFESP). Campus Baixada Santista. Santos , SP, Brazil

Abstract

Acute lung injury (ALI) is characterized by acute lung insufficiency, a sudden onset, recruitment of polymorphonuclear leukocyte recruitment and the release of inflammatory mediators, and it can induce an increase in the cytokine levels, and it is one of the consequences of severe COVID-19. There is no effective treatment, and the mortality rates are high. The stimuli of cholinergic pathway and nicotinic receptors modulate the acute lung inflammatory response in different experimental models, and more recently studies associating the nicotinic receptor with COVD-19 and the angiotensin converse enzyme - 2 (ACE-2) have been reported. However, in vitro studies that better elucidate the mechanism of action and compare the different way to stimulate nicotinic receptors are necessary. This project aims to evaluate the effect of different nicotinic receptor (±7-nAChR e ²2-nAChR) agonists and positive allosteric modulators in pulmonary cell culture models. For this purpose, human pulmonary epithelial cells BEAS-2B, A549 and a mouse macrophage lineage (RAW 264.7) will be cultivated, stimulated with LPS and treated with a nicotinic receptor agonist or an allosteric modulator. A dose-response curve was done to determine the better concentration and time. In these times, the supernatant will be collected for cytokine analysis and the cell material to analyze the gene expression of acetylcholine (AChE), vesicular acetylcholine transporter (VAChT), nicotinic acetylcholine receptors (±7 and ²2-nAChR), ACE2 receptor, as well as the JAK-2/STAT-3/SOCS-3 pathway and NrF-2, the regulator of anti-oxidative response. The oxidative stress will be evaluated. The statistical analysis will be performed through the program GraphPad Prism. Based on preliminary data from our group suggesting that the stimuli of nicotinic receptors modulate the acute inflammatory response in in vivo models, it is expected that the use of agonist and modulators reduce the inflammatory response in these cellular models and maybe some mechanism can be elucidated, as well as some possible differences between modulators and agonists.