Mapping of spatial distribution of tissue protease activity in mice injected with Bothrops jararaca snake venom by mass spectrometry and multispectral microscopy.

• Grant number: 24/22646-4
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate (Direct)
• Start date: April 01, 2025
• End date: March 31, 2026
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Solange Maria de Toledo Serrano
• Grantee: Jorge da Cruz Moschem
• Supervisor: Oliver Schilling
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Institution abroad: Universitätsklinikum Freiburg, Germany
• Associated to the scholarship: 23/17619-5 - Bothrops jararaca venom and renal failure: molecular bases of toxicity on mesangial cells and murine renal tissue, the synergistic effect of plasma protein degradation products, and the role of cathepsins, BP.DD

Abstract

Snake envenomation is classified by the WHO as a neglected tropical disease, which claims millions of victims annually around the world. In Brazil, Bothrops snakes are responsible for more than 90% of snakebites, causing local and systemic pathological effects in the envenomation process. Damage at the site of the bite is primarily characterized by edema, hemorrhage, and myonecrosis, in addition to severe pain. Systemically, the clinical picture involves consumptive coagulopathy, hemorrhages, rhabdomyolysis, and renal injury, which may progress to renal failure. These complex symptoms are caused by the synergistic action of venom toxins, including (i) abundant enzymes, such as Snake Venom Metalloproteases (SVMP) and Serine Proteases (SVSPs), and Phospholipases A2 (PLA2), and (ii) non-enzymatic proteins and biologically active peptides. Tissue damage at the site of the bite often leaves victims with lifelong severe injuries and is generally only partially treatable by antivenoms based on hyperimmune sera containing polyclonal antibodies. Furthermore, systemic pathological effects are not completely mitigated by antivenom treatment in some cases. In a recent collaboration study between the groups of Solange Serrano and Oliver Schilling, the local and systemic effects of Bothrops jararaca venom was extensively investigated using a murine model and applying multiomics approaches. The findings of this study, as well as of other investigations of venom proteolytic activity, clearly indicated the activation of endogenous proteases and the disruption of proteolytic homeostasis at the site of venom injection and systemically, contributing to the pro-inflammatory and deleterious events characteristic of Bothrops envenomation. Thus, the general objective of this proposal is to further explore the participation of mammalian host proteases in the envenomation process by assessing the spatial distribution of proteolytic activity in the muscle and in the kidney tissue of mice injected with venom in the gastrocnemius, mimicking a snakebite. To this end, the following specific objective is proposed: to evaluate the in situ proteolytic activity in the gastrocnemius muscle and renal tissue of mice injected with B. jararaca venom alone and in the presence of cathepsin-type cysteine protease (JPM-OEt) and meprin (actinonin) inhibitors using mass spectrometry (MALDI-Mass Spectrometry Imaging, MALDI-MSI, and Parallel Reaction Monitoring, PRM), and multispectral microscopy. The findings of MALDI-MSI in combination with PRM analysis, and multispectral microscopy for immune profiling of specific markers, will provide: (i) valuable insights into the participation of endogenous proteases in local (gastrocnemius muscle) and systemic (kidney) tissue damage and (ii) clues for new approaches to treating snakebite envenomation.