title: Membrane independent activation of fibroblast proMMP-2 by snake venom: novel roles for venom proteinases.
abstract: ProMMP-2 activation by Bothrops asper venom was investigated in mouse gastrocnemius muscle, mammalian cell culture and a cell-free system. Zymography revealed an increment of latent and activated forms of MMP-2 in muscle homogenates 1-3 days after venom injection. To clarify if venom can induce expression and activation of MMP-2, independently of the inflammatory response, venom was added to cultured human fibroblasts, endothelial and skeletal muscle cells, which expressed proMMP-2 constitutively. Venom activated proMMP-2 without promoting its expression. Venom also activated and degraded proMMP-2 in supernatants collected from fibroblast cultures, indicating that cells are not required for this activation. Pretreatment with EDTA increased MMP-2 activation and reduced degradation. Venom serine proteinases activated proMMP-2, whereas BaP1, a P-I metalloproteinase, predominantly degraded the latent and active forms of MMP-2. Moreover, pretreatment of conditioned medium with serine proteinase inhibitors greatly reduced the venom-induced activation, suggesting that venom proteinases activate MMP-2 via a serine proteinase secreted by fibroblasts. Venom also directly activated and degraded purified proMMP-2, albeit requiring a high concentration. Thus, B. asper venom proteinases activate and degrade proMMP-2 without inducing its synthesis. Serine proteinases play a dominant role in the activation, whereas metalloproteinases predominantly degrade MMP-2. Activation of proMMP-2 by snake venom proteinases, independently of the MT1-MMP/TIMP-2 pathway, extracellular matrix degradation or apoptosis, represents a novel mechanism in human fibroblasts.
authors.affiliation: Microbiology and Tumorbiology Center (MTC), Karolinska Institute, Box 280, 171 77 Stockholm, Sweden. Patricia.Saravia@mtc.ki.se
authors.forename: Patricia
authors.initials: P
authors.lastname: Saravia-Otten
authors.forename: Teresa
authors.initials: T
authors.lastname: Frisan
authors.forename: Monica
authors.initials: M
authors.lastname: Thelestam
authors.forename: José María
authors.initials: JM
authors.lastname: Gutiérrez
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D017837:Bothrops; D002478:Cells, Cultured; D003435:Crotalid Venoms / Q000633:toxicity*; D017931:DNA Primers; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects*; D018093:Gelatinases / Q000378:metabolism*; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008666:Metalloendopeptidases / Q000378:metabolism*; D051379:Mice; D018482:Muscle, Skeletal / Q000187:drug effects
chemical_list: D003435:Crotalid Venoms; D017931:DNA Primers; D004792:Enzyme Precursors; D007770:L-Lactate Dehydrogenase; D018093:Gelatinases; D008666:Metalloendopeptidases; C069860:progelatinase