Tissue Localization and Extracellular Matrix Degradation by PI, PII and PIII Snake Venom Metalloproteinases: Clues on the Mechanisms of Venom-Induced Hemorrhage.

Snake venom hemorrhagic metalloproteinases (SVMPs) of the PI, PII and PIII classes were compared in terms of tissue localization and their ability to hydrolyze basement membrane components in vivo, as well as by a proteomics analysis of exudates collected in tissue injected with these enzymes. Immunohistochemical analyses of co-localization of these SVMPs with type IV collagen revealed that PII and PIII enzymes co-localized with type IV collagen in capillaries, arterioles and post-capillary venules to a higher extent than PI SVMP, which showed a more widespread distribution in the tissue. The patterns of hydrolysis by these three SVMPs of laminin, type VI collagen and nidogen in vivo greatly differ, whereas the three enzymes showed a similar pattern of degradation of type IV collagen, supporting the concept that hydrolysis of this component is critical for the destabilization of microvessel structure leading to hemorrhage. Proteomic analysis of wound exudate revealed similarities and differences between the action of the three SVMPs. Higher extent of proteolysis was observed for the PI enzyme regarding several extracellular matrix components and fibrinogen, whereas exudates from mice injected with PII and PIII SVMPs had higher amounts of some intracellular proteins. Our results provide novel clues for understanding the mechanisms by which SVMPs induce damage to the microvasculature and generate hemorrhage. Author Summary: Local and systemic hemorrhage are typical manifestations of envenomings by viperid snakes. Hemorrhagic activity is due to the action of snake venom metalloproteinases (SVMPs) in the microvasculature, especially in capillaries. There are large differences in the hemorrhagic potential of SVMPs, depending on their domain composition. The present study compared PI, PII and PIII hemorrhagic SVMPs for their tissue distribution and their ability to cleave proteins of the extracellular matrix (ECM), especially those of the basement membrane (BM) that provides mechanical stability to microvessels. Observations indicate that PII and PIII SVMPs, which exert a high hemorrhagic activity, are preferentially located in microvessels, whereas PI SVMP is distributed in a more widespread fashion in the tissue. In addition, when these toxins are injected at doses that induce a similar hemorrhagic effect, they cleave type IV collagen to a similar extent, showing differences in the cleavage patterns of other ECM components, such as laminin, nidogen and type VI collagen. The analysis of the exudates resultant from the action of these SVMPs in the tissue revealed many similarities and some differences in the exudate proteomes. Overall our results indicate that hydrolysis of type IV collagen is a key event in the onset of microvessel damage, and that the ability of SVMPs to bind to microvessels greatly determines their hemorrhagic potential. [ABSTRACT FROM AUTHOR]