Immunohistochemical characterization of lymphocytes in microscopic colitis

Background

Microscopic colitis, encompassing the subgroups collagenous colitis (CC) and lymphocytic colitis (LC), is clinically characterized by chronic non-bloody watery diarrhoea. The diseases present with a macroscopically normal or near-normal colonic mucosa, but show increased numbers of intraepithelial lymphocytes (IELs) and mononuclear cell infiltration in the underlying lamina propria (LP). In CC a thickened (≥10 µm) subepithelial collagen band is also present. The IELs have previously been characterized as predominantly CD8+ T lymphocytes. In contrast,lymphocytes in the LP have mostly been described as CD4+.

Aims & Methods

The study aimed to further characterize the mucosal inflammation using immunohistochemistry to stain for lymphocytic surface markers.
Paraffin-embedded biopsies from proximal colon from 13 CC, 10 LC and 17 controls (Ctrl) were sectioned, mounted and stained using antibodies for CD3 (general marker of T lymphocytes), CD4 (helper/regulatory T lymphocytes), CD8 (cytotoxic T lymphocytes), CD20 (B lymphocytes) and CD30 (marker for activated T and B lymphocytes). The biopsies from the patients were all taken at the first diagnostic colonoscopy and no patient was on anti-inflammatory medication. Computerized image analysis, Qwin, was used to calculate areas of stained lymphocytes in the epithelium, LP, and crypts. The results in LP were confirmed with point counting of 10 fields from each patient using a 10x10 grid. Mann-Whitney-U test was used to analyze differences between patients and controls.

Results

The results are presented as median % stained lymphocytes as seen in the table and graphs.

Conclusion

This is one of the largest studies so far characterizing subgroups of lymphocytes in microscopic colitis. In CC and LC an increase of predominantly CD8+ lymphocytes was seen in both epithelium and lamina propria whereas, in contrast to other studies, a decreased amount of CD4+ lymphocytes was found in lamina propria.

The cause of this is unknown, but it can be speculated that a paucity of regulatory CD4+ T cells can play a role in the pathogenesis of microscopic colitis.

 

Fig 1A. Immunostained CD8+ lymphocytes in the colonic mucosa of a patient with lymphocytic colitis.

Fig 1B Shows same image with the quantified areas colored.

Fig 2. Increased areas with CD8+ lymphocytes in epithelium in MC patients

Fig 3. Decreased areas with CD4+ lymphocytes in lamina propria in MC patients.

Stained areas representing CD8+ lymphocytes were significantly increased in patients with microscopic colitis compared with controls (Fig 2). In contrast, the CD4+ areas in MC patients were significantly decreased in lamina propria (fig 3). Horizontal lines represent median values.