P2X7 receptor antagonist recovers ileum myenteric neurons after experimental ulcerative colitis

Table 1 Characteristics of primary and secondary antibodies

Antigen	Host	Dilution	Source
P2X7 receptor	Rabbit	1:200	Millipore (AB5246)
nNOS	Sheep	1:2000	Millipore (AB1529)
ChAT	Goat	1:50	Chemicon (AB144P)
Anti-HuC/D	Mouse	1:100	Molecular probes (A-21271)
GFAP	Rabbit	1:400	DAKO (Z0334)
GFAP	Mouse	1:200	Sigma (G3893)
Secondary antibodies
Donkey anti-rabbit IgG 488		1:500	Molecular probes (A21206)
Donkey anti-sheep IgG 594		1:100	Molecular probes (A11016)
Donkey anti-mouse IgG 594		1:200	Molecular probes (A21203)

nNOS: Neuronal nitric oxide synthase; ChAT: Choline acetyltransferase; GFAP: Glial fibrillary acidic protein.

Table 2 Macroscopic and microscopic scores of ileums from the sham, 2,4,6-trinitrobenzene sulfonic acid and brilliant blue G groups

Variables	Sham	TNBS	BBG
Macroscopic score	0	0	0
Microscopic score
Ulcerations	0	0	0
Edema (submucosa)	0	0	0
Inflammatory cell infiltration	0	0	0

For macroscopic and microscopic analyses colitis was assessed according to colonic injury[30]. The scores were stratified as follows: 0 = normal, 1 = presence of hyperemia without ulcers, 2 = ulcerations without hyperemia, 3 = ulcerations at one site, 4 = two or more sites of ulcerations, 5 = sites of damage extending > 1 cm, and 6 to 10 = sites of damage extending > 2 cm, with the score increasing by 1 for each additional cm[30]. The microscopic colitis scores were assessed using a scoring system adapted from Erdogan et al[31] and Fabia et al[32]. The scores were categorized as follows according to the corresponding parameters. Ulcerations: 0 = no ulcer, 1 = single ulceration not exceeding the lamina muscularis mucosa, 2 = ulcerations not exceeding the mucosa, and 3 = ulcerations exceeding the submucosa. Edema (submucosa): 0 = no edema, 1 = mild edema, 2 = moderate edema, and 3 = severe edema. Inflammatory cell infiltration: 0 = no infiltration, 1 = mild infiltration, 2 = moderate infiltration, and 3 = dense infiltration[31,32]. TNBS: 2,4,6-trinitrobenzene sulfonic acid; BBG: Brilliant blue G.