Defective mucosal T cell death is sustainably reverted by infliximab in a caspase dependent pathway in Crohn's disease.
OBJECTIVE
To verify whether targeting defective mucosal T cell death underlies the sustained therapeutic benefit of infliximab in Crohn's disease, we explored its in vivo proapoptotic effect after 10 weeks of treatment, and its in vitro killing activity on lamina propria T cells (LPT) and peripheral blood T cells (PBT), both isolated from Crohn's disease patients.


METHODS
Endoscopic intestinal biopsies were collected from 10 Crohn's disease patients (six steroid refractory and four fistulising) before and after three consecutive infusions of infliximab, administered at week 0, 2, and 6 in a single intravenous dose (5 mg/kg), and from 10 subjects who proved to have functional diarrhoea. Apoptosis was determined in vivo by TUNEL assay, and in vitro by fluorescein isothiocyanate-annexin V/propidium iodide staining on LPT and PBT from Crohn's disease patients cultured with infliximab. The effect of the broad caspase inhibitor Z-VAD-FMK and the neutralising anti-Fas antibody ZB4 was tested in vitro on LPT and PBT treated with infliximab. Caspase-3 activity was determined by immunoblotting.


RESULTS
In Crohn's disease patients, infliximab treatment induced a sustained LPT apoptosis, still evident four weeks after the last infusion. In vitro infliximab induced death of LPT from Crohn's disease patients occurred via apoptosis rather than necrosis. LPT showed a higher susceptibility to infliximab induced apoptosis than PBT in Crohn's disease patients. The signalling pathway underlying the restoration of infliximab induced LPT apoptosis occurred via the caspase pathway but not Fas-Fas ligand interaction in Crohn's disease.


CONCLUSIONS
These findings demonstrate that apoptosis is the major mechanism by which infliximab exerts its killing activity on LPT in Crohn's disease. The sustained LPT proapoptotic action of infliximab, which extends far beyond its circulating half life, may be responsible for the sustained remission induced in Crohn's disease patients by infliximab retreatment.
Di Sabatino
A
A
Gastroenterology Unit, IRCCS Policlinico S Matteo, University of Pavia, Pavia, Italy.
Ciccocioppo
R
R
Cinque
B
B
Millimaggi
D
D
Morera
R
R
Ricevuti
L
L
Cifone
M G
MG
Corazza
G R
GR
D000328:Adult; D000911:Antibodies, Monoclonal / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism / Q000502:physiology; D002478:Cells, Cultured; D003424:Crohn Disease / Q000188:drug therapy / Q000276:immunology* / Q000473:pathology; D004306:Dose-Response Relationship, Immunologic; D005260:Female; D005765:Gastrointestinal Agents / Q000494:pharmacology* / Q000627:therapeutic use; D006801:Humans; D018928:Immunity, Mucosal / Q000187:drug effects; D000069285:Infliximab; D007413:Intestinal Mucosa / Q000276:immunology* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D015398:Signal Transduction / Q000187:drug effects; D016176:T-Lymphocyte Subsets / Q000187:drug effects* / Q000276:immunology; D016896:Treatment Outcome; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors
D000911:Antibodies, Monoclonal; D005765:Gastrointestinal Agents; D014409:Tumor Necrosis Factor-alpha; D000069285:Infliximab; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases