Immune mechanisms in hepatic and pulmonary fibrosis associated to telomeropathies in murine model and human disease

• Grant number: 19/25002-2
• Support type: Scholarships in Brazil - Doctorate
• Effective date (Start): March 01, 2020
• Effective date (End): February 29, 2024
• Field of knowledge: Health Sciences - Medicine - Medical Clinics
• Principal researcher: Rodrigo do Tocantins Calado de Saloma Rodrigues
• Grantee: Willian Robert Gomes
• Home Institution: Hemocentro de Ribeirão Preto. Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto da USP (HCMRP). Secretaria da Saúde (São Paulo - Estado). Ribeirão Preto , SP, Brazil

Abstract

Telomeres are complex structures made up of repetitive nucleotide sequences that associate with proteins to form the ends of linear chromosomes. Telomeric extension decreases with each mitotic cycle and once the telomere length becomes too short, it induces cells to enter senescence or apoptosis. Some cell types, such as embryonic or adult stem cells, must maintain their proliferative capacity and, for that, they have the telomerase enzyme complex, which is able to lengthen telomeres and attenuate their erosion. Several mutations may lead to excessive telomere shortening, leading to the development of telomeropathies. Hematopoietic stem cell failure (aplastic anemia) is one of the main clinical manifestations of telomeropathies, but patients may also develop fibrotic processes in some organs, such as in the lungs (pulmonary fibrosis) and in the liver (cirrhosis). Although there is evidence showing the correlation between telomeric shortening and liver cirrhosis, the mechanisms involved in this process are not well understood. The present work aims to evaluate the immune mechanisms involved in the development of hepatic and pulmonary fibrotic process caused by telomeric shortening. For this, Terc-/- and Tert-/- knockout mice, along with wild type animals with the same genetic background, will be studied. Cirrhosis will be induced by controlled percutaneous infection with Schistosoma mansoni. On day 120 post-infection, fibrosis will be evaluated in liver samples dyed with Picro-Sirius Red, while gene expression will be evaluated by next generation RNA sequencing (RNASeq) and phenotypic characterization of infiltrated macrophages will be performed by immunohistochemistry. Bone marrow derived macrophages will be harvested and studied for their ability to be polarized, in vitro. Subsets of peripheral blood-derived T-cells will be identified and quantified through flow-cytometric immunophenotyping. In serum samples, the cytokine profiles will be evaluated through Luminex assay, as markers of the inflammatory pathways involved with the disease. Additionally, patients with telomeropathies associated to pulmonary fibrosis and/or cirrhosis will also be studied. Patients with telomeropathies and isolated bone marrow failure, without fibrotic manifestation, will be studied as a control group. Phenotypic characterization and stratification of T-cells subsets, in peripheral blood, will be performed by Cytometry by Time-Of-Flight (CyTOF) and serum cytokine pattern will be determined through Luminex to possibly identify the cellular and molecular immune signature in patients who develop fibrotic disease. It is expected that the results contribute to a better understanding of how telomeric dysfunctions play a role on the development of fibrosis and, also, new therapeutic targets against these diseases could be identified to be used for future treatments. (AU)