Validation of a cell-based assay to differentiate between the cytotoxic effects of elapid snake venoms

Yasmean Kalam, Geoffrey K. Isbister, Peter Mirtschin, Wayne C. Hodgson, Nicki Konstantakopoulos

    Research output: Contribution to journalArticleResearchpeer-review

    Abstract

    Introduction: Acanthophis genus (i.e. death adders) and the Naja genus (i.e. cobras) belong to the family elapidae. The current study compared the in vitro cytotoxicity of venoms from four Acanthophis spp. and three Naja spp. on rat aortic smooth muscle cells, A7r5, and rat skeletal muscle cells, L6. The ability of CSL death adder antivenom and SAIMR antivenom, for Acanthophis spp. and Naja spp. venom respectively, to negate the cytotoxicity was also examined. 

    Methods: A cell proliferation assay was used to determine cell viability following treatment with venom in the presence or absence of antivenom. Sigmoidal growth curves were obtained, and IC50 values were determined. 

    Results: Acanthophis spp. and Naja spp. venoms produced concentration-dependent inhibition of cell proliferation in both cell lines. Naja spp. venoms were significantly more cytotoxic than the most potent Acanthophis venom (i.e. A. antarcticus) in both cell lines. Naja spp. venoms also displayed higher sensitivity in L6 cells. SAIMR antivenom significantly inhibited the cytotoxic actions of all Naja spp. venoms in both A7r5 and L6 cells. However, death adder antivenom (CSL Ltd) was unable to negate the cytotoxic effects of Acanthophis spp. venoms. 

    Discussion: Concentrations of the predominantly cytotoxic Naja spp. venoms used were approximately three times less than the predominantly neurotoxic Acanthophis spp. venoms. SAIMR antivenom was partially effective in neutralising the effects of Naja spp. venoms. Death adder antivenom (CSL Ltd) was not effective in negating the cytotoxic effects of venom from Acanthophis spp. These results indicate that the cell-based assay is suited to the examination of cytotoxic snake venoms and may be used in conjunction with organ bath experiments to pharmacologically characterise snake venoms. Furthermore, the results suggest that the use of a skeletal muscle cell line is likely to be more clinically relevant for the examination of cytotoxic snake venoms.

    Original languageEnglish
    Pages (from-to)137-142
    Number of pages6
    JournalJournal of Pharmacological and Toxicological Methods
    Volume63
    Issue number2
    DOIs
    Publication statusPublished - Mar 2011

    Fingerprint

    Elapid Venoms
    Snake Venoms
    Venoms
    Elapidae
    Assays
    Antivenins
    Adders
    Cells
    Muscle
    Cell proliferation
    Cytotoxicity
    Cell Line
    Muscle Cells
    Rats
    Skeletal Muscle
    Cell Proliferation

    Cite this

    Kalam, Yasmean ; Isbister, Geoffrey K. ; Mirtschin, Peter ; Hodgson, Wayne C. ; Konstantakopoulos, Nicki. / Validation of a cell-based assay to differentiate between the cytotoxic effects of elapid snake venoms. In: Journal of Pharmacological and Toxicological Methods. 2011 ; Vol. 63, No. 2. pp. 137-142.
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    abstract = "Introduction: Acanthophis genus (i.e. death adders) and the Naja genus (i.e. cobras) belong to the family elapidae. The current study compared the in vitro cytotoxicity of venoms from four Acanthophis spp. and three Naja spp. on rat aortic smooth muscle cells, A7r5, and rat skeletal muscle cells, L6. The ability of CSL death adder antivenom and SAIMR antivenom, for Acanthophis spp. and Naja spp. venom respectively, to negate the cytotoxicity was also examined. Methods: A cell proliferation assay was used to determine cell viability following treatment with venom in the presence or absence of antivenom. Sigmoidal growth curves were obtained, and IC50 values were determined. Results: Acanthophis spp. and Naja spp. venoms produced concentration-dependent inhibition of cell proliferation in both cell lines. Naja spp. venoms were significantly more cytotoxic than the most potent Acanthophis venom (i.e. A. antarcticus) in both cell lines. Naja spp. venoms also displayed higher sensitivity in L6 cells. SAIMR antivenom significantly inhibited the cytotoxic actions of all Naja spp. venoms in both A7r5 and L6 cells. However, death adder antivenom (CSL Ltd) was unable to negate the cytotoxic effects of Acanthophis spp. venoms. Discussion: Concentrations of the predominantly cytotoxic Naja spp. venoms used were approximately three times less than the predominantly neurotoxic Acanthophis spp. venoms. SAIMR antivenom was partially effective in neutralising the effects of Naja spp. venoms. Death adder antivenom (CSL Ltd) was not effective in negating the cytotoxic effects of venom from Acanthophis spp. These results indicate that the cell-based assay is suited to the examination of cytotoxic snake venoms and may be used in conjunction with organ bath experiments to pharmacologically characterise snake venoms. Furthermore, the results suggest that the use of a skeletal muscle cell line is likely to be more clinically relevant for the examination of cytotoxic snake venoms.",
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    author = "Yasmean Kalam and Isbister, {Geoffrey K.} and Peter Mirtschin and Hodgson, {Wayne C.} and Nicki Konstantakopoulos",
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    Validation of a cell-based assay to differentiate between the cytotoxic effects of elapid snake venoms. / Kalam, Yasmean; Isbister, Geoffrey K.; Mirtschin, Peter; Hodgson, Wayne C.; Konstantakopoulos, Nicki.

    In: Journal of Pharmacological and Toxicological Methods, Vol. 63, No. 2, 03.2011, p. 137-142.

    Research output: Contribution to journalArticleResearchpeer-review

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    AU - Kalam, Yasmean

    AU - Isbister, Geoffrey K.

    AU - Mirtschin, Peter

    AU - Hodgson, Wayne C.

    AU - Konstantakopoulos, Nicki

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    N2 - Introduction: Acanthophis genus (i.e. death adders) and the Naja genus (i.e. cobras) belong to the family elapidae. The current study compared the in vitro cytotoxicity of venoms from four Acanthophis spp. and three Naja spp. on rat aortic smooth muscle cells, A7r5, and rat skeletal muscle cells, L6. The ability of CSL death adder antivenom and SAIMR antivenom, for Acanthophis spp. and Naja spp. venom respectively, to negate the cytotoxicity was also examined. Methods: A cell proliferation assay was used to determine cell viability following treatment with venom in the presence or absence of antivenom. Sigmoidal growth curves were obtained, and IC50 values were determined. Results: Acanthophis spp. and Naja spp. venoms produced concentration-dependent inhibition of cell proliferation in both cell lines. Naja spp. venoms were significantly more cytotoxic than the most potent Acanthophis venom (i.e. A. antarcticus) in both cell lines. Naja spp. venoms also displayed higher sensitivity in L6 cells. SAIMR antivenom significantly inhibited the cytotoxic actions of all Naja spp. venoms in both A7r5 and L6 cells. However, death adder antivenom (CSL Ltd) was unable to negate the cytotoxic effects of Acanthophis spp. venoms. Discussion: Concentrations of the predominantly cytotoxic Naja spp. venoms used were approximately three times less than the predominantly neurotoxic Acanthophis spp. venoms. SAIMR antivenom was partially effective in neutralising the effects of Naja spp. venoms. Death adder antivenom (CSL Ltd) was not effective in negating the cytotoxic effects of venom from Acanthophis spp. These results indicate that the cell-based assay is suited to the examination of cytotoxic snake venoms and may be used in conjunction with organ bath experiments to pharmacologically characterise snake venoms. Furthermore, the results suggest that the use of a skeletal muscle cell line is likely to be more clinically relevant for the examination of cytotoxic snake venoms.

    AB - Introduction: Acanthophis genus (i.e. death adders) and the Naja genus (i.e. cobras) belong to the family elapidae. The current study compared the in vitro cytotoxicity of venoms from four Acanthophis spp. and three Naja spp. on rat aortic smooth muscle cells, A7r5, and rat skeletal muscle cells, L6. The ability of CSL death adder antivenom and SAIMR antivenom, for Acanthophis spp. and Naja spp. venom respectively, to negate the cytotoxicity was also examined. Methods: A cell proliferation assay was used to determine cell viability following treatment with venom in the presence or absence of antivenom. Sigmoidal growth curves were obtained, and IC50 values were determined. Results: Acanthophis spp. and Naja spp. venoms produced concentration-dependent inhibition of cell proliferation in both cell lines. Naja spp. venoms were significantly more cytotoxic than the most potent Acanthophis venom (i.e. A. antarcticus) in both cell lines. Naja spp. venoms also displayed higher sensitivity in L6 cells. SAIMR antivenom significantly inhibited the cytotoxic actions of all Naja spp. venoms in both A7r5 and L6 cells. However, death adder antivenom (CSL Ltd) was unable to negate the cytotoxic effects of Acanthophis spp. venoms. Discussion: Concentrations of the predominantly cytotoxic Naja spp. venoms used were approximately three times less than the predominantly neurotoxic Acanthophis spp. venoms. SAIMR antivenom was partially effective in neutralising the effects of Naja spp. venoms. Death adder antivenom (CSL Ltd) was not effective in negating the cytotoxic effects of venom from Acanthophis spp. These results indicate that the cell-based assay is suited to the examination of cytotoxic snake venoms and may be used in conjunction with organ bath experiments to pharmacologically characterise snake venoms. Furthermore, the results suggest that the use of a skeletal muscle cell line is likely to be more clinically relevant for the examination of cytotoxic snake venoms.

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