Effect of ammodytin L from Vipera ammodytes on L-6 cells from rat skeletal muscle.
Ammodytin L (AMDL) is a myotoxic phospholipase-like protein from the venom of Vipera ammodytes with a serine in position 49 instead of an aspartate, therefore this toxin is devoid of phospholipase activity, and the membrane-damaging effect does not involve any step of phospholipase activity. The aim of the present study was to analyze the effect of AMDL on L-6 cells from rat skeletal muscle to investigate its mechanism of action and the role of calcium ions in its muscle-damaging activity. Our data indicate that the effect of ammodytin L is strongly dependent on the degree of cell differentiation. Low doses of myotoxin gave rise to a marked release of creatine kinase in myotubes differentiated from L-6 myoblasts and the presence of calcium ions plays a role in the cytotoxic effect. The presence of EGTA in the incubation buffer reduced by 50% the release of creatine kinase. No membrane damage was observed in myoblasts, but there was a significant increase of intracellular calcium concentration measured with Fura-2. A non-specific membrane effect of AMDL was ruled out using platelets as reference cells: no platelet aggregation pattern and no increase in intracellular calcium were observed.
Incerpi
S
S
Department of Biology, University of Rome Tor Vergata, Rome, Italy.
de Vito
P
P
Luly
P
P
Rufini
S
S
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line / Q000187:drug effects; D003402:Creatine Kinase / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid; D009132:Muscles / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D013997:Time Factors; D014757:Viper Venoms / Q000494:pharmacology*
D014757:Viper Venoms; C065419:ammodytin L; D004533:Egtazic Acid; D003402:Creatine Kinase; D002118:Calcium