Vacuolization of HeLa cells by a partially purified Clostridium histolyticum cytotoxin.
Clostridium histolyticum vacuolating cytotoxin was partially purified from culture broth using ammonium sulfate precipitation, gel filtration and hydrophobic interaction chromatography. The toxin caused vacuolization of HeLa cells visible under a light microscope after 2 h and distinct after 8 h. Transmission electron microscopy revealed the presence of numerous vacuoles, condensation of the mitochondrial matrix, increased cytoplasm density and increased amounts of heterochromatin. Apoptosis was not detected either by electron microscopy or by an apoptosis/necrosis discrimination assay with fluorescein-labeled annexin V and propidium iodide, or DNA fragmentation assay. Calcium ion influx was detected by flow cytometry after labeling cells with Fluo-4 AM. Vacuolation of HeLa cells by C. histolyticum cytotoxin was inhibited by bafilomycin A1, suggesting involvement of H+ -ATPase in the formation of vacuoles.
Department of Histology and Embryology, Warsaw Medical University, Warsaw, Poland.
Ewa
E
Jankowska-Steifer
Jarosław
J
Jóźwiak
Tomasz
T
Grzela
Aldona
A
Komar
Justyna
J
Niderla
Maciej
M
Lazarczyk
Grazyna
G
Korczak-Kowalska
Stanisław
S
Moskalewski
Gayane
G
Martirosian
D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000378:metabolism; D003015:Clostridium Infections / Q000382:microbiology; D047009:Hathewaya histolytica / Q000737:chemistry* / Q000502:physiology; D003603:Cytotoxins / Q000302:isolation & purification* / Q000494:pharmacology*; D004247:DNA / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D020287:In Situ Nick-End Labeling; D018942:Macrolides / Q000494:pharmacology; D046529:Microscopy, Electron, Transmission; D009499:Neutral Red / Q000493:pharmacokinetics; D012964:Sodium / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D014617:Vacuoles / Q000187:drug effects* / Q000382:microbiology / Q000502:physiology
D003603:Cytotoxins; D004791:Enzyme Inhibitors; D018942:Macrolides; C057620:bafilomycin A; D009499:Neutral Red; D004247:DNA; D012964:Sodium; D025262:Vacuolar Proton-Translocating ATPases; D002118:Calcium