Effect of epsilon toxin-GFP on MDCK cells and renal tubules in vivo.
Epsilon toxin (epsilon-toxin), produced by Clostridium perfringens types B and D, causes fatal enterotoxemia, also known as pulpy kidney disease, in livestock. Recombinant epsilon-toxin-green fluorescence protein (epsilon-toxin-GFP) and epsilon-prototoxin-GFP were successfully expressed in Escherichia coli. MTT assays on MDCK cells confirmed that recombinant epsilon-toxin-GFP retained the cytotoxicity of the native toxin. Direct fluorescence analysis of MDCK cells revealed a homogeneous peripheral pattern that was temperature sensitive and susceptible to detergent. epsilon-Toxin-GFP and epsilon-prototoxin-GFP bound to endothelia in various organs of injected mice, especially the brain. However, fluorescence mainly accumulated in kidneys. Mice injected with epsilon-toxin-GFP showed severe kidney alterations, including hemorrhagic medullae and selective degeneration of distal tubules. Moreover, experiments on kidney cryoslices demonstrated specific binding to distal tubule cells of a range of species. We demonstrate with new recombinant fluorescence tools that epsilon-toxin binds in vivo to endothelial cells and renal tubules, where it has a strong cytotoxic effect. Our binding experiments indicate that an epsilon-toxin receptor is expressed on renal distal tubules of mammalian species, including human.
Soler-Jover
Alex
A
Department de Biologia Cellular i Anatomia Patològica, Universitat de Barcelona, L'Hospitalet de Llobregat, Barcelona, Spain.
Blasi
Juan
J
Gómez de Aranda
Inma
I
Navarro
Piedad
P
Gibert
Maryse
M
Popoff
Michel R
MR
Martín-Satué
Mireia
M
D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics / Q000493:pharmacokinetics / Q000633:toxicity*; D002417:Cattle; D016923:Cell Death / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D004285:Dogs; D005404:Fixatives; D005557:Formaldehyde; D049452:Green Fluorescent Proteins; D006801:Humans; D066298:In Vitro Techniques; D007684:Kidney Tubules / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008024:Ligands; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D008856:Microscopy, Fluorescence; D011108:Polymers; D011485:Protein Binding; D011498:Protein Precursors / Q000235:genetics / Q000493:pharmacokinetics / Q000633:toxicity; D011869:Radioligand Assay; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000493:pharmacokinetics / Q000633:toxicity; D012756:Sheep; D013045:Species Specificity; D014018:Tissue Distribution
D001427:Bacterial Toxins; C039811:Clostridium perfringens epsilon-toxin; D005404:Fixatives; D008024:Ligands; D008164:Luminescent Proteins; D011108:Polymers; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D005557:Formaldehyde; C003043:paraform