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Doctoral Thesis
DOI
https://doi.org/10.11606/T.17.2020.tde-11022020-143954
Document
Author
Full name
Sarah Capelupe Simões
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2019
Supervisor
Committee
Costa Neto, Claudio Miguel da (President)
Godinho, Rosely Oliveira
Passaglia, Rita de Cassia Aleixo Tostes
Teixeira, Felipe Roberti
Title in Portuguese
Análise proteômica celular após estímulo do receptor AT1 por ligantes com diferentes perfis farmacológicos
Keywords in Portuguese
AT1R
Fracionamento celular
GPCR
Proteômica
Abstract in Portuguese
Os receptores acoplados a proteína G (GPCRs) são importantes alvos de estudos biomédicos. O receptor de angiotensina II do tipo 1 (AT1R) medeia importantes efeitos fisiopatológicos em diferentes órgãos e tecidos e tem como principal efetor a AngII. Neste trabalho investigamos por análise proteômica como o estímulo por 24hrs de células HEK293T expressando AT1R com o ligantes AngII, TRV027, Ang1-7 e L1623,313, que apresentam diferentes perfis farmacológicos, afetam a abundância total de proteínas e como essas alterações podem modificar eventos celulares, tais como apoptose e autofagia. Após a análise global de proteínas, demos início ao estudo de subfrações celulares através da padronização do fracionamento celular e análise por espectrometria de massas. Dessa forma este amplo estudo usando diversas abordagens desde a escolha dos ligantes com diferentes características farmacológicas até o monitoramento de proteínas enriquecidas em diferentes subfrações, permitiu a descrição de eventos celulares e vias de sinalização intimamente ligadas a ativação do AT1R. Este estudo enriquece significativamente o conhecimento a respeito do AT1R permitindo no futuro correlacionar eventos celulares microscópicos, como o tráfego proteico a eventos fisiológicos mais complexos e direcionar o desenvolvimento de novos fármacos.
Title in English
Cell Proteomics analysis after AT1 receptor stimulation by ligands with diferente pharmacological profiles
Keywords in English
AT1 receptor
Cell fractionation
GPCR
Proteomics
Abstract in English
G-protein coupled receptors (GPCRs) are important targets in biomedical studies. Angiotensin II type 1 receptor (AT1R) mediates important pathophysiological effects in different organs and tissues, where the peptide AngII is its main effector. In this work, we investigated by proteomic analysis how stimulation of HEK293T cells expressing AT1R 24hrs by the ligands AngII, TRV027, Ang1-7 or L1623,313, which have different pharmacological profiles, modulate the total abundance of proteins, and how these alterations can modify events such as apoptosis and autophagy. After the global analysis of proteins, we performed a of cellular subfractions study by standardization of cell fractionation and mass spectrometry analysis. This study, using different approaches such as choosing ligands with different pharmacological profiles and monitoring proteins enriched in different cellular subfractions, allowed the description of cellular events and signaling pathways closely related to AT1R activation. Our study significantly enriches the knowledge about AT1R, which shall allow in the future to correlate microscopic cellular events, such as protein traffic, to more complex physiological events, and hence to contribute and drive the development of new drugs.
 
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Publishing Date
2020-05-05
 
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