Skip to main content
SpringerLink
Log in

Effect of Brown Spider Venom on Basement Membrane Structures

  • Published:
The Histochemical Journal Aims and scope Submit manuscript

Abstract

Loxoscelism or necrotic arachnidism are terms used to describe lesions and reactions induced by bites (envenomation) from spiders of the genus Loxosceles. Envenomation has been reported to provoke dermonecrosis and haemorrhage at the bite site and haemolysis, disseminated intravascular coagulation and renal failure. The purpose of this work was to study the effect of the venom of the brown spider Loxosceles intermedia on basement membrane structures and on its major constituent molecules. Light microscopy observations showed that L. intermedia venom obtained through electric shock, which reproduces two major signals of Loxoscelism in the laboratory, exhibits activity toward basement membrane structures in mouse Engelbreth-Holm-Swarm (EHS) sarcoma. Basement degradation was seen by a reduced periodic acid-Schiff (PAS) and alcian blue staining as well as by a reduced immunostaining for laminin when compared to control experiments. Electron microscopy studies confirmed the above results, showing the action of the venom on EHS-basement membranes and demonstrating that these tissue structures are susceptible to the venom. Using purified components of the basement membrane, we determined through SDS-PAGE and agarose gel that the venom is not active toward laminin or type IV collagen, but is capable of cleaving entactin and endothelial heparan sulphate proteoglycan. In addition, when EHS tissue was incubated with venom we detected a release of laminin into the supernatant, corroborating the occurrence of some basement membrane disruption. The venom-degrading effect on entactin was blocked by 1,10-phenanthroline, but not by other protease inhibitors such as PMSF, NEM or pepstatin-A. By using light microscopy associated with PAS staining we were able to identify that 1,10-phenanthroline also inhibits EHS-basement membrane disruption evoked by venom, corroborating that a metalloprotease of venom is involved in these effects. Degradation of these extracellular matrix molecules and the observed susceptibility of the basement membrane could lead to loss of vessel and glomerular integrity, resulting in haemorrhage and renal problems after envenomation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References cited

  • Aumailley M, Wiedemann H, Mann K, Timpl R (1989) Binding of nidogen and the laminin-nidogen complex to basement membrane collagen type IV. Eur J Biochem 184: 241-248.

    Google Scholar 

  • Baramova EN, Shannon JD, Bjarnason JB, Fox JW (1989) Degradation of extracellular matrix proteins by hemorrhagic metalloproteinases. Arch Biochem Biophys 275: 63-71.

    Google Scholar 

  • Barbaro KC, Cardoso JLC, Eickstedt VRD, Mota I (1992) Dermonecrotic and lethal components of Loxosceles gaucho spider venom. Toxicon 30: 331-338.

    Google Scholar 

  • Bascur L, Yevenes I, Barja P (1982) Effects of Loxosceles laeta spider venom on blood coagulation. Toxicon 20: 795-796.

    Google Scholar 

  • Beçak W, Paulete J (1976) Técnicas de Citologia e Histologia. Rio de Janeiro, Livros Técnicos e Científicos Editora S.A.

  • Buonassisi V, Venter JC (1976) Hormone and neurotransmitter receptors in an established vascular endothelial cell line. Proc Natl Acad Sci USA 73: 1612-1616.

    Google Scholar 

  • Civello DJ, Moran JB, Geren CR (1983) Substrate specificity of a hemorragic proteinase from timber rattlesnake venom. Biochemistry 22: 755-762.

    Google Scholar 

  • Denny WF, Dillaha, Morgan PN (1964) Hemotoxic effect of Loxosceles reclusa venom: in vivo and in vitro studies. J Lab Clin Med 64: 291-298.

    Google Scholar 

  • Dietrich CP, Dietrich SMC (1976) Electrophoretic behaviour of acidic mucopolysaccharides in dyamine buffers. Anal Biochem 70: 645-647. 408 S.S. Veiga et al.

    Google Scholar 

  • Eskafi FM, Norment BR (1976) Physiological action of Loxosceles reclusa venom on insect larvae. Toxicon 14: 7-12.

    Google Scholar 

  • Farquhar MG (1991) The glomerular basement membrane: a selective macromolecular filter. In: Cell Biology of Extracellular Matrix, New York: Plenum Press.

    Google Scholar 

  • Feitosa L, Gremski W, Veiga SS, Elias MCQB, Graner E, Mangili OC, Brentani RR (1998) Detection and characterization of metalloproteinases with gelatinolytic, fibronectinolytic and fibrinogenolytic activities in brown spider (Loxosceles intermedia) venom. Toxicon 36: 1039-1051.

    Google Scholar 

  • Forrester LJ, Barret JT, Campbell BJ (1978) Red blood cell lysis induced by the venom of brown reclusa spider. The role of sphingomyelinase D. Arch Biochem Biophys 187: 335-365.

    Google Scholar 

  • Fox JW, Bjarnason JB (1995) Atrolysins: metalloproteinases from Crotalus atrox venom. Methods Enzymol 248: 368-387.

    Google Scholar 

  • Fox JW, Mayer U, Nischt R, Aumailley M, Reinhardt D, Wiedmann H, Mann K, Timpl R, Kreig T, Engel J, Chu M-L (1991) Recombinant nidogen consists of three globular domains and mediates binding of laminin to collagen type IV. EMBO J 10: 3137-3146.

    Google Scholar 

  • Futrell J (1992) Loxocelism. Am J Med Sci 304: 261-267.

    Google Scholar 

  • Gremski W, Cutler LS (1991) Immunocytochemical localization of heparan sulphate proteoglycan in the rat submandibular gland. Histochem J 23: 339-344.

    Google Scholar 

  • Harlow E, Lane D (1988) Antibodies: A Laboratory Manual. New York: Coldspring Harbor.

    Google Scholar 

  • Jong Y-S, Norment BR, Heitz JR (1979) Separation and characterization of venom components in Loxosceles reclusa-II. Protease enzyme activity. Toxicon 16: 529-537.

    Google Scholar 

  • Karnovsky MJ (1965) A formaldehyde-glutaraldehyde fixative of high osmolality for use in electron microscopy. J Cell Biol 27: 137A.

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685.

    Google Scholar 

  • Mayer U, Zimmermann K, Mann K, Reinhardt D, Timpl R, Nischt R (1995) Binding properties and protease stability of recombinant human nidogen. Eur J Biochem 227: 681-686.

    Google Scholar 

  • Mackay AR, Hartzler JL, Pelina MD, Thorgeirsson UP (1990) Studies on the ability of 65-and 92-kDa tumour cell gelatinases to degrade type IV collagen. J Biol Chem 265: 21929-21934.

    Google Scholar 

  • McManus JFA(1948) Histological demonstration of mucin after periodic acid. Nature 158: 202.

    Google Scholar 

  • Nader HB, Dietrich CP, Buonassisi V, Colburn P (1987) Heparin sequences in the heparan sulphate chains of an endothelial cell proteoglycan. Proc Natl Acad Sci USA 84: 3565-3569.

    Google Scholar 

  • Nader HB, Buonassisi V, Colburn P, Dietrich CP (1989) Heparan stimulates the synthesis and modifies the sulfation pattern of heparan sulfate proteoglycan from endothelial cells. J Cell Physiol 140: 305-310.

    Google Scholar 

  • Ohsaka A, Just M, Habermann E (1973) Action of snake venom hemorrhagic principles on isolated glomerular basement membrane. Biochim Biophys Acta 323: 415-428.

    Google Scholar 

  • Paulsson M, Aumailley M, Deutzmann R, Timpl R, Beck K (1987) Laminin-nidogen complex: extraction with chelating agents and structural characterization. Eur J Biochem 166: 11-19.

    Google Scholar 

  • Plow EF, Pierschbacher MD, Ruoslahti E, Marguerie GA (1985) The effect of arg-gly-asp-containing peptides on fibrinogen and von Willebrand factor binding to platelets. Proc Natl Acad Sci USA 82: 8057-8061.

    Google Scholar 

  • Rees RS, Nanney LB, Yates RA, King LE (1984) Interaction of brown recluse spider venom on cell membranes: the inciting mechanism? J Invest Derm 83: 270-275.

    Google Scholar 

  • Reinhardt D, Mann K, Nischt R, Fox JW, Chu M, Krieg T, Timpl R (1993) Mapping of nidogen binding sites for collagen type IV, heparan sulfate proteoglycan, and zinc. J Biol Chem 268: 10881-10887.

    Google Scholar 

  • Rohrbach DH, Timpl R (1993) Molecular and Cellular Aspects of Basement Membranes. San Diego, CA: Academic Press.

    Google Scholar 

  • Timpl R, Rohde H, Robey PG, Rennard SI, Foidartc JM, Martin GR (1979) Laminin-a glycoprotein from basement membranes. J Biol Chem 254: 9933-9937.

    Google Scholar 

  • Timpl R, Paulsson M, Dziadek M, Fujiwara S (1987) Basement membranes. Meth Enzymol 145: 363-391.

    Google Scholar 

  • Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedures and some applications. Proc Natl Acad Sci USA 76: 4350-4354.

    Google Scholar 

  • Veiga SS, Chammas R, Cella N, Brentani RR (1995) Glycosylation of beta-1integrins in B16-F10 mouse melanoma cells as determinant of differential binding and acquisition of biological activity. Int J Cancer 61: 420-424.

    Google Scholar 

  • Veiga SS, Gremski W, Santos VLP, Feitosa L, Mangili OC, Nader HB, Dietrich CP, Brentani RR (1999) Oligosaccharide residues of Loxosceles intermedia (brown spider) venom proteins: dependence on glycosylation for dermonecrotic activity. Toxicon 37: 587-607.

    Google Scholar 

  • Yurchenco P, Ruben GC (1987) Basement membrane structure in situ: evidence for lateral associations in the type IV collagen network. J Cell Biol 105: 2559-2568.

    Google Scholar 

  • Yurchenco P, Schittny J (1990) Molecular and cellular aspects of basement membranes. FASEB J 4: 1577-1590.

    Google Scholar 

Download references

Author information

Author notes

  1. Vera Lúcia P. dos Santos

    Present address: Department of Biochemistry, Federal University of São Paulo, São Paulo, Brazil

Authors and Affiliations

  1. Department of Cell Biology, Federal University of Parana, Jardim das Americas, 81531-990, Curitiba, Parana, Brazil

    Silvio S. Veiga, Vera Lúcia P. dos Santos, Gustavo A. de Souza, Andrea S. Ribeiro & Waldemiro Gremski

  2. Department of Biochemistry, Federal University of São Paulo, São Paulo, Brazil

    Loara Feitosa, Marimélia A. Porcionatto, Helena B. Nader & Carl P. Dietrich

  3. Department of Physiology, Federal University of Parana, Curitiba, Parana, Brazil

    Oldemir C. Mangili

  4. Ludwig Institute for Cancer Research, São Paulo, Brazil

    Ricardo R. Brentani

Authors
  1. Silvio S. Veiga
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Loara Feitosa
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Vera Lúcia P. dos Santos
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Gustavo A. de Souza
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Andrea S. Ribeiro
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Oldemir C. Mangili
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Marimélia A. Porcionatto
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Helena B. Nader
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Carl P. Dietrich
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Ricardo R. Brentani
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Waldemiro Gremski
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Veiga, S.S., Feitosa, L., dos Santos, V.L.P. et al. Effect of Brown Spider Venom on Basement Membrane Structures. Histochem J 32, 397–408 (2000). https://doi.org/10.1023/A:1004031019827

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1004031019827

Keywords

Search

Navigation