Phenotypic study of extracellular vesicles and their relationship with senescence in the context of COVID-19

• Grant number: 24/18417-0
• Support Opportunities: Scholarships in Brazil - Program to Stimulate Scientific Vocations
• Start date: May 05, 2025
• End date: June 14, 2025
• Field of knowledge: Biological Sciences - Immunology - Applied Immunology
• Principal Investigator: Ana Maria Caetano de Faria
• Grantee: Larissa Augusta de Sousa
• Host Institution: Instituto de Ciências Biológicas. Universidade Federal de Minas Gerais (UFMG). Ministério da Educação (Brasil). Belo Horizonte , SP, Brazil

Abstract

Extracellular vesicles (EVs) are nanosized particles released by almost all cell types and are recognized as new tools for local and systemic intercellular communication. It has been demonstrated that EVs carry and transfer a wide variety of molecules, such as proteins, nucleic acids, microRNAs, messenger RNAs. These molecules may be involved in physiological functions, the pathology of various diseases, or even therapeutic processes through the modulation of immunity and inflammation. Added to this, we know that during viral infections EVs incorporate nucleic acids, proteins and lipids derived from pathogens and become a vector for delivering viral materials. In turn, Severe Acute Respiratory Syndrome caused by Coronavirus (SARS-CoV-2) is characterized by the multiplication of the virus in the lower or upper respiratory tract, leading to an increase in the production of cytokines and consequent local and systemic inflammation. In this sense, it is important to understand the relationship between EVs and a possible mechanism for the metabolic dysregulation characteristic of COVID-19. During the internship period, the objectives of the work plan will be: 1) learn the methodology for isolating and studying EVs; 2) understand the relationship between microvesicles released into the blood by immune cells and the senescence phenotype presented by individuals with COVID-19; 3) interact with the group and research topics at the UFMG Immunobiology Laboratory. This work plan will be part of the Project "Role of Cellular Communication through Extracellular Vesicles in COVID-19 in Elderly Individuals" approved by the National Research Ethics Commission (CONEP - CAAE 40208320.3.1001.5149) and funded by the State Research Support Foundation of Minas Gerais (FAPEMIG, APQ 01301/24) which is currently being conducted at the Immunobiology Laboratory by doctoral student Cecília Horta Ramalho Pinto. Blood samples were collected from adult (20-60 years) and elderly (>60 years) volunteers and individuals were previously tested using qRT-PCR for SARS-CoV-2 (nasopharyngeal swab). Routine clinical and laboratory examination, nutritional and anthropometric assessments were also carried out after signing the Informed Consent Form (TCLE). The blood samples have already been processed to obtain the serum and this will be ultracentrifuged at 100,000g for 4 hours to isolate the EVs. Thus, the isolated EVs will be immunophenotyped using Flow Cytometry, using the expression of markers (coupled to fluorochromes for analysis by flow cytometry): annexin V FITC, CD66b PE, CD45APC, CD51/61 PE, CD235aPECy5, 7 CD3 PE , CD41 PerCP, CD16PE and CD14PerCP. The acquisition of microvesicles will be carried out using the CytoFLEX S equipment (Beckman Coulter®) and analysis using the CyteXPERT software and FlowJo®. The data obtained from individuals and groups composed of adults and elderly people with or without COVID-19 will be compared.Statistical analysis will be conducted using Prism GraphPad 8.0 software. The mechanisms by which cell-to-cell communication between microvesicles occurs during immunosenescence are still little explored by the scientific community. Therefore, the analysis of the immunophenotyping of these particles can offer a more specific overview of the function, communication and relationship of EVs with other molecules secreted by cells of the immune system. (AU)