Low molecular weight hemorrhagins were purified from crude Bothrops atrox
snake venom by gel filtration followed by ionic strength chromatography. The
protein fractions obtained, designated HI-1 to HI-8, contained
proteins with molecular masses lower than 30 kDa. HI-5, the most representative among of these fractions, exhibited, in vitro, proteolytic and C inactivating properties, as analyzed by proteolysis of a
protein substrate, and C system consumptive activities as assayed by reduction of the hemolytic C activity in normal human serum and by cleavage of partially purified component C3. HI-5
hemorrhagin injected i.m. into C-sufficient BALB/c mice induced a local
inflammation characterized by
edema, accumulation of polymorphonuclear leucocytes (PMN) and
hemorrhage. In contrast, when injected into BALB/c mice previously C-depleted, the number of PMN per tissue section, but not
hemorrhage, was significantly reduced (129.668 +/- 31.341 cells per microscopic field) as compared with the control C-sufficient mice (812.168 +/- 111.194 cells per microscopic field). The observations were confirmed by using C5-deficient mice instead of C-depleted mice. The average number of PMN per tissue section in C5-defficient A/J mice was 72.666 +/- 19.416 cells per microscopic field. These data indicate that the C system is involved in PMN accumulation, but not in the
hemorrhage, at the local induced lesions by low molecular mass B. atrox hemorrhagins. HI-5 apparently is not contaminated with other direct or indirect
inflammation mediators, PMN accumulation and
hemorrhage, however, an independent phenomenon, could be mediated by the same
hemorrhagin proteinase domain.