TNBS-induced colitis and inflammation scoring
Acute colitis was triggered on day 5 by the intra-rectal admin-
istration of a 50 ml solution of TNBS 100 mg kgÀ1 (Sigma-Aldrich
Chemical, France) in 0.9% NaCl/ethanol (50/50 v/v) (Drouault-
Holowacz et al., 2006). The animals were subsequently monitored
daily for any loss of body weight. Three days after the induction of
colitis, the mice were sacrificed; blood samples were collected
immediately and stored in heparinized tubes. After dissection of
the mice, two independent observers blindly scored the macro-
scopic inflammation of the colon on the Wallace scale (Wallace
et al., 1989). Two-cm-long fragments of the distal colon were
collected and frozen at À80 C while samples for histological
analysis were processed for paraffin-embedded 5-mm sections to be
stained with May Grünwald-Giemsa as described above. Following
examination under the microscope, tissue lesions were scored ac-
cording to the Ameho criteria (Ameho et al., 1997). In addition, the
degree of polymorphonuclear neutrophil infiltration into the distal
colon was assessed by quantifying myeloperoxidase (MPO) activity,
as reported previously (Bradley et al., 1982).