Venom Gland Mass Spectrometry Imaging of Saw-Scaled Viper, Echis carinatus sochureki, at High Lateral Resolution

Abstract

The snake venom gland is the place for the synthesis, storage, and secretion of a complex mixture of proteins and peptides, i.e., the venom. The morphology of the gland has been revealed by classical histology and microscopic studies. However, knowledge about the gland’s cellular secretory and functional processes is still incomplete and has so far been neglected by the omics disciplines. We used autofocusing atmospheric-pressure matrix-assisted laser desorption/ionization (AP-SMALDI) mass spectrometry imaging (MSI) to investigate endogenous biomolecular distributions in the venom glands of the saw-scaled viper, Echis carinatus sochureki, employing different sample preparation methods. Fresh-freezing and formalin-fixation were tested for the gland to obtain intact tissue sections. Subsequently, MSI was conducted with 12 μm pixel resolution for both types of preparations, and the lateral distributions of the metabolites were identified. Experiments revealed that lipids belonging to the classes of PC, SM, PE, PS, PA, and TG are present in the venom gland. PC (32:0) and SM (36:1) were found to be specifically located in the areas where cells are present. The snake venom metalloprotease inhibitor pEKW (m/z 444.2233) was identified in the venom by top-down LC–MS/MS and localized by MALDI-MSI in the gland across secretory epithelial cells. The peptide can inhibit the venom’s enzymatic activity during long-term storage within the venom gland. With a high degree of spectral similarities, we concluded that formalin-fixed tissue, in addition to its high ability to preserve tissue morphology, can be considered as an alternative method to fresh-frozen tissue in the case of lipid and peptide MS imaging in venom gland tissues.

Comparative Venom Proteomics of Iranian, Macrovipera lebetina cernovi, and Cypriot, Macrovipera lebetina lebetina, Giant Vipers

“…The venoms of M. l. cernovi , M. l. obtusa and M. l. lebetina were found to contain significant quantities of endogenous tripeptide metalloprotease inhibitors (SVMPi). The tripeptides were also detected previously from venoms of different species, e.g., Trimeresurus mucrosquamatus [ 57 ], Bothrops asper [ 58 ], Echis carinatus sochureki [ 59 ], Echis ocellatus , Cerastes cerastes cerastes [ 60 ], Deinagkistrodon acutus [ 61 ], Vipera ammodytes transcaucasiana and Vipera ammodytes montandoni [ 62 ], and various species of rattlesnakes [ 63 ], confirming their role in reducing the proteolytic activity of SVMPs during storage in the venom gland to prevent self-intoxication [ 59 ]. They appear to be among the essential components of snake venoms, which contain significant amounts of SVMP toxins.…”

Nanoparticle-based applications by atmospheric pressure matrix assisted desorption/ionization mass spectrometry

“…4d ). 101 In 2021, venom gland mass spectrometry imaging of saw-scaled viper, Echis carinatus sochureki , was realized by Parviz Ghezellous's group with a high lateral resolution of 12 μm via autofocusing AP MALDI MS. 102 Novelly, C. Jacques visualized sun filter skin spatial distribution and penetration and detected endogenous metabolites induced by a sun filter in human skin in 2022. 103 Metabolic study of aristolochic acid I (AAI)-exposed murine liver section by AP MALDI MSI was conducted by Wenjing Guo's group in 2022, demonstrating the distinct metabolic alternations.…”

Spatial Venomics─Cobra Venom System Reveals Spatial Differentiation of Snake Toxins by Mass Spectrometry Imaging

“…It allows unsupervised and simultaneous analyses of molecules (e.g., metabolites, proteins, peptides, lipids, and glycans) in situ on a single tissue section, preserving their spatial coordinates and generating a molecular intensity map reflecting the relative molecule abundance. − Since Caprioli et al introduced MALDI-MSI for biological samples in the late 90s, continual technical improvements have raised its suitability and applicability to become a valuable tool for use in the field of venomics. , Hence, it facilitates mapping of individual toxins on-tissue and provides deeper insights into the spatial venom distribution as well as heterogeneity and its effects on venom regulation or modulation . The localization of different venom toxins within the context of morphological structures has previously been described by MALDI-MSI in a multidimensional manner. − However, previous MSI experiments are restricted in their venom identification, which allows intact profiling of selected toxin classes and excludes the identification of higher molecular weight toxins, like snake venom serine proteases (svSP), snake venom metalloproteases (svMP), or l -amino acid oxidases (LAAO).…”