Detection of epithelial apoptosis in pelvic ileal pouches for ulcerative colitis and familial adenomatous polyposis

Abstract

Background: Ileal pouch-anal anastomosis (IPAA) is the surgical procedure of choice for patients with refractory ulcerative colitis (UC) and for familial adenomatous polyposis (FAP) with many rectal polyps. Pouchitis is one of the more frequent complications after IPAA in UC patients; however, it is rare in FAP.

Objective: Evaluate pro-apoptotic activity in endoscopically and histological normal mucosa of the ileal pouch in patients with UC and FAP.

Methods: Eighteen patients (nine with UC and nine with FAP) with J pouch after total rectocolectomy were studied. Biopsies were obtained from the mucosa of the pouch and from normal ileum. The specimens were snap-frozen and the expressions of Bax and Bcl-2 were determined by immunoblot of protein extracts and by immunohistochemistry analysis. FADD, Caspase-8, APAF-1 and Caspase-9 were evaluated by immunoprecipitation and immunoblot.

Results: Patients with UC had significantly higher protein levels of Bax and APAF-1, Caspase-9 than patients with FAP, but were similar to controls. The expressions of Bcl-2 and FADD, Caspase-8 were similar in the groups. Immunohistochemistry for Bax showed less intensity of immunoreactions in FAP than in UC and Controls. Bcl-2 immunostaining was similar among the groups.

Conclusion: Patients with FAP present lower levels of pro-apoptotic proteins in all methods applied, even in the absence of clinical and endoscopic pouchitis and dysplasia in the histological analysis. These findings may explain a tendency of up-regulation of apoptosis in UC patients, resulting in higher rates of progression to pouchitis in these patients, which could correlate with mucosal atrophy that occurs in inflamed tissue. However, FAP patients had low pro-apoptotic activity in the mucosa, and it could explain the tendency to low cell turn over and presence of adenomas in this syndrome.

Figure 1

Representative Western blot analyses and determination of Bax, Bcl-2, FADD - Caspase-8, APAF-1 - Caspase-9 protein expressions in non-inflamed pouches in the Control, FAP and UC groups. For illustration purpose each line band represents one patient. For all conditions, n = 09, *p < 0.05 vs Control; §p < 0.05 vs FAP.

Figure 2

Immunohistochemistry of ileal pouch sections from UC, FAP and control group immunoreacted for Bax and Bcl-2. Immunostainig for Bax was intense in UC group. Bcl-2 positive cell among the groups was similar. (200×)