Strain-dependent induction of epithelial cell oncosis by Campylobacter jejuni is correlated with invasion ability and is independent of cytolethal distending toxin
Induction of host cell death is thought to play an important role in bacterial pathogenesis. Campylobacter jejuni is a prevalent cause of bacterial enteritis; however, its effects on enterocytes remain unclear. The present study indicates for the first time that C. jejuni induces oncotic, rather than apoptotic death of T84 enterocytes. C. jejuni-treated enterocytes exhibited extensive cytoplasmic vacuolation, rapid (3–6 h) loss of plasma membrane integrity (‘cytotoxicity’), loss of mitochondrial transmembrane potential, and ATP depletion. Enterocytes also exhibited increased oligonucleosomal DNA fragmentation, a feature characteristic of apoptosis. However, consistent with a non-apoptotic process, DNA fragmentation and cytotoxicity were not caspase dependent. During apoptosis, caspases mediate cleavage of poly(ADP-ribose) polymerase; however, cleavage was not observed in C. jejuni-treated monolayers. Cytotoxicity, ATP depletion and DNA fragmentation were not prevented by the deletion of the cytolethal distending toxin (CDT) gene, indicating that C. jejuni causes enterocyte oncosis via a mechanism that is CDT independent. The ability to cause oncosis was significantly decreased in a FlaAFlaB mutant (CDT+) that was defective in the ability to adhere and invade enterocytes. Analysis of clinical isolates revealed that oncosis was strain dependent and correlated with increased invasive ability. These observations offer new insights into the pathogenesis of C. jejuni infection.
BangD. D.,
ScheutzF.,
AhrensP.,
PedersenK.,
BlomJ.,
MadsenM.2001; Prevalence of cytolethal distending toxin ( cdt) genes and CDT production in Campylobacter spp. isolated from Danish broilers. J Med Microbiol 50:1087–1094
BielaszewskaM.,
SinhaB.,
KucziusT.,
KarchH.2005; Cytolethal distending toxin from Shiga toxin-producing Escherichia coli O157 causes irreversible G2/M arrest, inhibition of proliferation, and death of human endothelial cells. Infect Immun 73:552–562
BuretA. G.,
ChinA. C.,
ScottK. G.2003; Infection of human and bovine epithelial cells with Cryptosporidium andersoni induces apoptosis and disrupts tight junctional ZO-1: effects of epidermal growth factor. Int J Parasitol 33:1363–1371
ChinA. C.,
TeohD. A.,
ScottK. G.,
MeddingsJ. B.,
MacnaughtonW. K.,
BuretA. G.2002; Strain-dependent induction of enterocyte apoptosis by Giardia lamblia disrupts epithelial barrier function in a caspase-3-dependent manner. Infect Immun 70:3673–3680
ChinA. C.,
VergnolleN.,
MacNaughtonW. K.,
WallaceJ. L.,
HollenbergM. D.,
BuretA. G.2003; Proteinase-activated receptor 1 activation induces epithelial apoptosis and increases intestinal permeability. Proc Natl Acad Sci U S A 100:11104–11109
De MeloM. A.,
GabbianiG.,
PechereJ. C.1989; Cellular events and intracellular survival of Campylobacter jejuni during infection of HEp-2 cells. Infect Immun 57:2214–2222
DenisM.,
SoumetC.,
RivoalK.,
ErmelG.,
BlivetD.,
SalvatG.,
ColinP.1999; Development of a m-PCR assay for simultaneous identification of Campylobacter jejuni and C. coli. Lett Appl Microbiol 29:406–410
DongZ.,
SaikumarP.,
WeinbergJ. M.,
VenkatachalamM. A.1997; Internucleosomal DNA cleavage triggered by plasma membrane damage during necrotic cell death. Involvement of serine but not cysteine proteases. Am J Pathol 151:1205–1213
EarnshawW. C.,
MartinsL. M.,
KaufmannS. H.1999; Mammalian caspases: structure, activation, substrates, and functions during apoptosis. Annu Rev Biochem 68:383–424
EnariM.,
SakahiraH.,
YokoyamaH.,
OkawaK.,
IwamatsuA.,
NagataS.1998; A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature 391:43–50
EverestP. H.,
GoossensH.,
ButzlerJ. P.,
LloydD.,
KnuttonS.,
KetleyJ. M.,
WilliamsP. H.1992; Differentiated Caco-2 cells as a model for enteric invasion by Campylobacter jejuni and C. coli. J Med Microbiol 37:319–325
EyigorA.,
DawsonK. A.,
LangloisB. E.,
PickettC. L.1999; Detection of cytolethal distending toxin activity and cdt genes in Campylobacter spp. isolated from chicken carcasses. Appl Environ Microbiol 65:1501–1505
Fernandez-PradaC.,
TallB. D.,
ElliottS. E.,
HooverD. L.,
NataroJ. P.,
VenkatesanM. M.1998; Hemolysin-positive enteroaggregative and cell-detaching Escherichia coli strains cause oncosis of human monocyte-derived macrophages and apoptosis of murine J774 cells. Infect Immun 66:3918–3924
GaoJ. X.,
MaB. L.,
XieY. L.,
HuangD. S.1991; Electron microscopic appearance of the chronic Campylobacter jejuni enteritis of mice. Chin Med J (Engl 104:1005–1010
GaynorE. C.,
CawthrawS.,
ManningG.,
MacKichanJ. K.,
FalkowS.,
NewellD. G.2004; The genome-sequenced variant of Campylobacter jejuni NCTC 11168 and the original clonal clinical isolate differ markedly in colonization, gene expression, and virulence-associated phenotypes. J Bacteriol 186:503–517
HickeyT. E.,
MajamG.,
GuerryP.2005; Intracellular survival of Campylobacter jejuni in human monocytic cells and induction of apoptotic death by cytholethal distending toxin. Infect Immun 73:5194–5197
HumphreyC. D.,
MontagD. M.,
PittmanF. E.1986; Morphologic observations of experimental Campylobacter jejuni infection in the hamster intestinal tract. Am J Pathol 122:152–159
KimJ. S.,
HeL.,
LemastersJ. J.2003; Mitochondrial permeability transition: a common pathway to necrosis and apoptosis. Biochem Biophys Res Commun 304:463–470
KonkelM. E.,
KlenaJ. D.,
Rivera-AmillV.,
MontevilleM. R.,
BiswasD.,
RaphaelB.,
MickelsonJ.2004; Secretion of virulence proteins from Campylobacter jejuni is dependent on a functional flagellar export apparatus. J Bacteriol 186:3296–3303
KorlathJ. A.,
OsterholmM. T.,
JudyL. A.,
ForfangJ. C.,
RobinsonR. A.1985; A point-source outbreak of campylobacteriosis associated with consumption of raw milk. J Infect Dis 152:592–596
Lara-TejeroM.,
GalanJ. E.2001; CdtA, CdtB, and CdtC form a tripartite complex that is required for cytolethal distending toxin activity. Infect Immun 69:4358–4365
LeistM.,
SingleB.,
CastoldiA. F.,
KuhnleS.,
NicoteraP.1997; Intracellular adenosine triphosphate (ATP) concentration: a switch in the decision between apoptosis and necrosis. J Exp Med 185:1481–1486
LiP.,
NijhawanD.,
BudihardjoI.,
SrinivasulaS. M.,
AhmadM.,
AlnemriE. S.,
WangX.1997; Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell 91:479–489
LindblomG. B.,
KaijserB.1995; In vitro studies of Campylobacter jejuni/coli strains from hens and humans regarding adherence, invasiveness, and toxigenicity. Avian Dis 39:718–722
LosM.,
MozolukM.,
FerrariD.,
StepczynskaA.,
StrohC.,
RenzA.,
HercegZ.,
WangZ. Q.,
Schulze-OsthoffK.2002; Activation and caspase-mediated inhibition of PARP: a molecular switch between fibroblast necrosis and apoptosis in death receptor signaling. Mol Biol Cell 13:978–988
MaC.,
WickhamM. E.,
GuttmanJ. A.,
DengW.,
WalkerJ.,
MadsenK. L.,
JacobsonK.,
VoglW. A.,
FinlayB. B.,
VallanceB. A.2006; Citrobacter rodentium infection causes both mitochondrial dysfunction and intestinal epithelial barrier disruption in vivo: role of mitochondrial associated protein (Map. Cell Microbiol 8:1669–1686
MillerJ. F.,
DowerW. J.,
TompkinsL. S.1988; High-voltage electroporation of bacteria: genetic transformation of Campylobacter jejuni with plasmid DNA. Proc Natl Acad Sci U S A 85:856–860
MontevilleM. R.,
YoonJ. E.,
KonkelM. E.2003; Maximal adherence and invasion of INT 407 cells by Campylobacter jejuni requires the CadF outer-membrane protein and microfilament reorganization. Microbiology 149:153–165
NadeauE.,
MessierS.,
QuessyS.2003; Comparison of Campylobacter isolates from poultry and humans: association between in vitro virulence properties, biotypes, and pulsed-field gel electrophoresis clusters. Appl Environ Microbiol 69:6316–6320
NewellD. G.,
PearsonA.1984; The invasion of epithelial cell lines and the intestinal epithelium of infant mice by Campylobacter jejuni/coli. J Diarrhoeal Dis Res 2:19–26
PalombaL.,
SestiliP.,
ColumbaroM.,
FalcieriE.,
CantoniO.1999; Apoptosis and necrosis following exposure of U937 cells to increasing concentrations of hydrogen peroxide: the effect of the poly(ADP-ribose)polymerase inhibitor 3-aminobenzamide. Biochem Pharmacol 58:1743–1750
PurdyD.,
BuswellC. M.,
HodgsonA. E.,
McAlpineK.,
HendersonI.,
LeachS. A.2000; Characterisation of cytolethal distending toxin (CDT) mutants of Campylobacter jejuni. J Med Microbiol 49:473–479
SiegesmundA. M.,
KonkelM. E.,
KlenaJ. D.,
MixterP. F.2004; Campylobacter jejuni infection of differentiated THP-1 macrophages results in interleukin 1 beta release and caspase-1-independent apoptosis. Microbiology 150:561–569
SmileyS. T.,
ReersM.,
Mottola-HartshornC.,
LinM.,
ChenA.,
SmithT. W.,
SteeleG. D.Jr,
ChenL. B.1991; Intracellular heterogeneity in mitochondrial membrane potentials revealed by a J-aggregate-forming lipophilic cation JC-1. Proc Natl Acad Sci U S A 88:3671–3675
Strain-dependent induction of epithelial cell oncosis by Campylobacter jejuni is correlated with invasion ability and is independent of cytolethal distending toxin