Interaction between P2X7 and TRPV1 receptors in the activation of NLRP3 inflammasome and extracellular vesicle release in the murine BV2 microglial lineage

Author(s)

Sávio Lima Bastos

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2024-05-13

Examining board members

Sabrina Francesca de Souza Lisboa; Carolina Demarchi Munhoz; Robson Coutinho Silva

• Advisor: Sabrina Francesca de Souza Lisboa

Abstract

Microglia cells are the resident macrophages of the central nervous system, acting as the primary defense line and highly responsive to stress stimuli. There are several receptors and intracellular pathways involved in the microglial activation process, with a focus on the P2X7 and TRPV1 receptors, two sensors activated in stressful situations. P2X7 receptors are activated by high concentrations of ATP in the extracellular medium, while TRPV1 receptors are activated by capsaicin and other stimuli associated with damage, such as acidity and high temperatures, in addition to endogenous cannabinoids. The activation of both receptors is related to the activation of the NLRP3 inflammasome and the release of extracellular vesicles (EVs) in microglia, but it is not clear how these receptors interact in these processes. In this study, we investigated how the pharmacological modulation of TRPV1, through activation with capsaicin or blockade with the antagonist SB366791, interacts with the activation of the NLRP3 inflammasome and the release of EVs induced by the activation of P2X7 receptors by ATP. For this purpose, murine BV2 microglia cell line cells were challenged with lipopolysaccharide, treated with SB366791 (TRPV1 antagonist), and finally stimulated with ATP and/or capsaicin. It was observed that treatment with ATP and capsaicin induced changes in the expression of TRPV1 dimers, which may be indicative of receptor sensitization and desensitization mechanisms. Treatment with ATP induced activation of the NLRP3 inflammasome in the cells, although we did not detect mature forms of caspase-1 and IL-1β in the cell lysate. Treatment with capsaicin also induced an increase in the expression of NLRP3 and pro-IL-1β and interacted with the effects promoted by ATP treatment on the expression of ASC, NLRP3, and pro-IL-1β. In the evaluation of EVs secreted by microglia, treatment with ATP induced an increase in the release of EVs, an effect that was reversed by treatment with capsaicin. Thus, we observed an intricate relationship between TRPV1 and P2X7 receptors in the activation of the inflammasome and the release of microglial EVs, with both synergistic and antagonistic effects observed upon the activation of both receptors. However, further experiments are needed to confirm the mechanisms behind these phenomena. (AU)

• FAPESP's process: 21/05406-1 - Study of possible changes in the endocannabinoid, endovaniloid, and neuroimmune systems in brain after stimulation of microglia cells: involvement of epigenetic mechanisms
• Grantee: Sávio Lima Bastos
• Support Opportunities: Scholarships in Brazil - Master