Banca de DEFESA: MAURO VALENTINO PALOSCHI

Uma banca de DEFESA de DOUTORADO foi cadastrada pelo programa.
DISCENTE : MAURO VALENTINO PALOSCHI
DATA : 29/09/2021
HORA: 09:00
LOCAL: Sala Virtual Remota
TÍTULO:

BIOGENESIS OF HUMAN NEUTROPHIL PRO-INFLAMMATORY MEDIATORS STIMULATED BY L-AMINO ACID OXIDASE OF Calloselasma rhodostoma VENOM

PALAVRAS-CHAVES:

Snake venom. Inflammasome. Cytosolic phospholipase A2. Cyclooxygenases. Lipid droplets

PÁGINAS: 149
GRANDE ÁREA: Ciências Biológicas
ÁREA: Biologia Geral
RESUMO:

Snake venom contains compounds that have the potential to be used in bioprospecting in biology and medicine. These proteins, such as L-amino acid oxidases (LAAOs), cause edema, microbicidal activity, cytotoxicity, coagulant and anticoagulant activity, hemorrhagic and pro-inflammatory effects. LAAO isolated from the venom of Calloselasma rhodosthoma (Cr-LAAO) has been shown to be a powerful stimulator of neutrophil activation and inflammatory mediator production. However, the mechanisms underlying Cr-LAAO-induced cell activation are not well understood. Previous research has shown that this enzyme can activate the NADPH oxidase complex and the PKC-  signaling protein, resulting in the production of reactive oxygen species (ROS), as well as stimulate phagocytosis, chemotaxis with the participation of p38 MAPK and PI3K, MPO release, cytokine production (IL-8, IL-6, and TNF-), neutrophil extracellular traps (NETs), and lipid mediators (LTB4 and PGE2). The activation of the cyclooxygenase (COX) pathway can result in the formation of some lipid mediators within lipid droplets (LDs). LDs are cytoplasmic organelles that are highly associated with the inflammatory response. They are found in a variety of cell types and can increase in number and/or size during an infection. Simultaneously, ROS can activate other inflammatory mechanisms such as inflammasomes. NLRP3-type inflammasomes are cytosolic protein complexes that cleave caspase-1, resulting in the proteolytic activation of pro-inflammatory cytokines such as IL-1. Thus, the purpose of this study is to assess the effect of Cr-LAAO on the activation of the NLRP3 inflammasome, as well as the activation of the COX pathway for the production of PGE2 and its involvement in the formation of LDs. The neutrophil transcriptome was examined using the microarray technique, and it was discovered that Cr-LAAO plays a role in upregulating the expression of genes involved in the NADPH oxidase complex and inflammasomes, as well as genes involved in lipid signaling and metabolism. Immunofluorescence assays for NLRP3 revealed protein accumulation and the formation of punctas in the cytosol, indicating that this complex is activated. Furthermore, protein expression analysis in the presence and/or absence of Apocynin, an inhibitor of NADPH oxidase, revealed that when ROS are inhibited, the expression of proteins that comprise the NLRP3 inflammasome is reduced. The release of IL-1β was also detected, and the pharmacological inhibition of NLRP3, caspase 1 and ROS reduced the levels of this released cytokine. Some of the genes positively regulated in the microarray belong to the phospholipase A2 family, specifically cytosolic phospholipases A2- (cPLA2-); and enzymes involved in the synthesis of PGE2, COX-2, and prostaglandin E synthase (PTGES). In addition, the genes perilipins 2 and 3 (PLIN 2 and 3) and diacylglycerol acyltransferase 1 (DGAT1), which are involved in the formation of LDs, were up-regulated. In neutrophils, Cr-LAAO stimulated increased cPLA2-phosphorylation, LD biogenesis, and PGE2 synthesis. cPLA2- (CAY10650) and DGAT-1 (A922500) inhibitors inhibited LD formation and PGE2 secretion. Finally, this is the first study to look at the effects of Cr-LAAO on the regulation of metabolism and lipid signaling in human neutrophils, as well as its ability to stimulate the activation of the NLRP3 inflammasome complex via ROS production

MEMBROS DA BANCA:
Externo à Instituição - GUSTAVO BATISTA DE MENEZES - UFMG
Externo à Instituição - HELDER NAKAYA - USP
Externo à Instituição - JOSÉ ROBERTO MINEO - UFU
Presidente - 1726461 - JULIANA PAVAN ZULIANI TRENCH DE SOUZA
Externo à Instituição - PATRICIA TORRES BOZZA - FIOCRUZ