In vitro and in vivo studies on some toxic effects of the venom from Hemiscorpious lepturus scorpion

The aim of this laboratory-based study was to investigate some of the toxic effects induced by the venom from Hemiscorpious lepturus (H. lepturus). For this aim, pharmacological, histological, biochemical methods as well as complete blood cell count were used to assess these toxic actions. In addition, in vitro haemolysis studies on human washed blood suspension and cytotoxicity on cultured fibroblasts were also undertaken. In vitro pharmacological test was made on rat isolated ileal segment. To this end, the effects of the venom on the contractile responsiveness to acetylcholine were recorded using F30 transducer and Darco chart recorder. For assessment of the haemolytic potency, varying concentrations (2, 10, 20 and 40 μg/ml) of the venom were added to 0.5 ml of 5% washed human blood and after 30 min, 2, 4, 8, 12 and 24 h of exposure, the degree of lysis (extent of redness developed in the supernatant solution after centrifugation) were measured by ELISA method. Cytotoxicity potential of the venom was assessed by trypan blue exclusion test. The venom (0.1, 1 and 10 μg/ml) was mixed with confluent fibroblast cell culture and the extent cytotoxicity was assessed microscopically. In vivo studies were conducted by a subcutaneous administration of sub-lethal dose (10 μg) of the venom and after 7 days the skin, at the site of injection, and kidney samples were stained by H & E method and examined microscopically. In addition, biochemical assessments including measurement of serum aspartate aminotransferase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP) and amylase levels and urine analysis were made. The results showed that the venom prevented the relaxation phase of the acetylcholine-induced contractions on the isolated ileal segments and finally produced sustained spasmodic contractions. This spasmodic action was abolished by 1 μM atropine. The venom produced haemolysis of red blood cells in a concentration-dependent and duration-of-exposure manner, with 100% of haemolysis produced after 24 h following exposure to 40 μg/ml of venom. While cultured fibroblasts cells were more sensitive and disintegrated after 15 min of exposure to 1 μg/ml of the venom. Histological findings showed evidences of excessive inflammatory responses accompanied with signs of necrosis in the skin at the site of injection as well as structural damage in the nephrones. There was a significant rise in the serum enzymes. In addition, the number of the RBCs were reduced. The urine showed positive readings for proteinuria, blood and intact RBCs. The overall results suggest that the venom from H. lepturus primarily is a cytotoxic agent and has haemolytic, nephrotoxic and to some extent hepatotoxic activity.