Intravascular hemolysis has been described in envenomings by the Eastern coral snake, Micrurus fulvius, in dogs. An experimental model of
intravascular hemolysis was developed in mice after intravenous (i.v.) injection of M. fulvius
venom. Within one hr, there was prominent
hemolysis, associated with a drastic drop in hematocrit, morphological alterations of erythrocytes, hemoglobinemia, and
hemoglobinuria.
Hemoglobin was identified in urine by mass spectrometry. Histological sections of kidney revealed abundant hyaline casts, probably corresponding to
hemoglobin. This effect was abrogated by
p-bromophenacyl bromide, indicating that it is caused by
phospholipases A2 (PLA2). A monospecific anti-Micrurus nigrocinctus
antivenom neutralized hemolytic activity in vivo. When tested in vitro with erythrocytes of various species, a clear difference in susceptibility was observed. Mouse and dog erythrocytes showed the highest susceptibility, whereas human and rabbit erythrocytes were not affected at the experimental conditions tested. The higher susceptibility of dog and mouse erythrocytes correlates with a high ratio of
phosphatidylcholine/
sphingomyelin in erythrocyte plasma membrane. When mouse erythrocytes were subjected to mechanical stress, after incubation with
venom,
hemolysis increased significantly, suggesting that both
phospholipid hydrolysis by PLA2s and mechanical stress associated with rheological factors are likely to contribute to cell lysis in vivo. Several PLA2s isolated from this
venom reproduced the hemolytic effect, and the complete amino acid sequence of one of them (fraction 17), which also induces
myotoxicity, is reported. Since very few PLA2s inducing
intravascular hemolysis have been described from
snake venoms, this
enzyme is a valuable tool to identify the structural determinants of hemolytic activity. The mouse model described in this study may be useful to explore the pathophysiology of
intravascular hemolysis.