Novel histological repertoire of crypt-associated anomalies in inflamed colon mucosa

Carlos A Rubio; Michael Vieth; Corinna Lang-Schwarz

doi:10.1136/jclinpath-2022-208152

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Original research

Novel histological repertoire of crypt-associated anomalies in inflamed colon mucosa

http://orcid.org/0000-0003-3879-5729Carlos A Rubio1,
Michael Vieth2,
http://orcid.org/0000-0001-8631-2600Corinna Lang-Schwarz2

¹ Department of Pathology, Karolinska Institute and University Hospital, Stockholm, Sweden
² Institute of Pathology, Friedrich-Alexander University Erlangen-Nuremberg, Klinikum Bayreuth, Bayreuth, Germany

Correspondence to Professor Carlos A Rubio, Department of Pathology, Karolinska Institute and University Hospital, Stockholm, 171 76, Sweden; carlos.rubio{at}ki.se

Abstract

Aims Studying crypt branching in ulcerative colitis (UC) and in infectious colitis (IC), we detected previously unreported crypt-associated anomalies (CAAs). The objective was to describe, illustrate and assess the frequency of CAAs in inflamed colon mucosa in patients with UC and IC.

Methods Sections from 100 consecutive biopsies with UC, in 50 with IC and in 27 with UC in remission (UCR) were reviewed. The following CAAs were identified: crypt eosinophilia, intracryptal epithelial hyperplasia, intracryptal epithelial budding, intracryptal supernumerary crypts, intracryptal epithelial bridges, crypt rings in rows and off-centre epithelial budding.

Results The frequency of crypts with extensive crypt eosinophilia and with intracryptal epithelial budding was significantly higher in UC than in IC and UCR (p<0.05); the frequency in the remaining histological parameters was similar in UC, IC and UCR.

Conclusions CAAs were found interspersed with branching crypts. CAAs persisted in long-lasting UC mucosal inflammation, but declined when the inflammation waned. Since similar anomalies are not present in normal colon mucosa, the results suggest that CAAs had been boosted by the ongoing mucosal inflammation. The development of these previously unreported CAAs in the colon mucosa with inflammation might embody pathological aberrations of cryptogenesis.

colitis
inflammation
inflammatory bowel diseases
intestine, large

Data availability statement

Data are available upon reasonable request.

https://creativecommons.org/licenses/by/4.0/

This is an open access article distributed in accordance with the Creative Commons Attribution 4.0 Unported (CC BY 4.0) license, which permits others to copy, redistribute, remix, transform and build upon this work for any purpose, provided the original work is properly cited, a link to the licence is given, and indication of whether changes were made. See: https://creativecommons.org/licenses/by/4.0/.

https://doi.org/10.1136/jclinpath-2022-208152

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Introduction

The sagittal profile of the normal colon mucosa at low magnification discloses a wavy silhouette due to small mucosal protrusions called anthemia, a Greek term used to define alternating decorative motifs.1 An anthemion is built of parallel close mucosal invaginations called crypts. In well-oriented sections, the crypts are arranged perpendicular to the surface epithelium and to the muscularis mucosae, as ‘test tubes’ in a rack.2 In the middle of the anthemion, the crypts are the tallest, but decrease in height, sideways. This explains the slight variations in the height of the crypts found in well-oriented tissue sections.1 The anthemia are limited by furrows called innominate grooves.3

In early infancy, the building of new crypts is achieved by symmetric branching or fission.4 Building of new crypts begins at their base with a crevice; the cleft progresses upwards until two identical crypts are eventually formed.5 During late infancy and adulthood, crypt replication wanes; crypts divide only once or twice, with average crypt cycle length of 36 years.4 Thus, crypt branching in the colon of adults is a rare finding.6–8 In contrast, the colonic mucosa in patients with inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn colitis, displays increased number of branching crypts.9 Several authors claim that branching can easily be detected in crypts cut in the vertical plane6 10 11 (ie, in well-oriented, upright crypts), but not in those cut in the horizontal plane (ie, in cross-cut crypts) as they ‘often produced acinar profiles’.11 12 ⇓ However, daily praxis shows that sections from endoscopic biopsies are fortuitously cut at the laboratory in a horizontal plane, a procedure often yielding cross-cut ring-shaped crypts. More recently, Rubio et al 13–16 reported the occurrence of increased numbers of branching crypts in endoscopic biopsies from patients with IBD. The presence of epithelial crests in ring-shaped cross-cut crypts identified the beginning of crypt branching.16 Their results showed that the number of crypts in asymmetric branching surpassed that of crypts in symmetric branching.14

Recently, while reviewing sections from patients with UC, a series of previously unreported crypt-associated histological anomalies were detected; they were regarded as histological artefacts. Notwithstanding, the review of endoscopic biopsies from patients with infectious colitis (IC) revealed similar artefacts. Against this background, the following questions arose:

Could those anomalies merely mirror crypt alterations conveyed by the mucosal inflammation per se?
Could one or more of those anomalies be pathognomonic of UC?

The aim of the present investigation was to explore these questions by analysing a cohort of patients with UC, IC and UC in remission (UCR).

Materials and methods

Biopsies from 100 consecutive patients with UC under surveillance and from 79 controls (50 with IC and 27 with UCR) were analysed. Following the local endoscopic scheme, a minimum of two endoscopic biopsies were taken from various colon segment and rectum. Biopsies were cut in 4 µm thick sections, stained with H&E, scanned with a Hamamatsu NanoZoomer S360 digital scanner and made available online to authors.

Novel histological parameters

The following histological parameters were found in the crypts in patients with colon inflammation: (1) crypt eosinophilia, (2) intraepithelial cell hyperplasia, (3) intracryptal epithelial budding, (4) intracryptal supernumerary crypts, (5) intraluminal epithelial bridges, (6) crypt rings in rows and (7) off-centre epithelial budding.

Definitions

Crypt eosinophilia

Under normal conditions, colon crypts are mainly lined with non-eosinophilic, mucus-producing goblet cells, which compress sideways columnar absorptive cells. Goblet cells decrease towards the upper portion of the crypts, and near the luminal portion of the crypts cells with eosinophilic cytoplasm are usually seen. Scrutiny of crypt cells revealed that crypt cells with intense eosinophilic cytoplasm were the columnar absorptive cells (figure 1A–D). Crypt eosinophilia was classified by semiquantitative estimates into (1) focal crypt eosinophilia (ie, haphazardly distributed eosinophilic crypts) and (2) extensive crypt eosinophilia (ie, crypt eosinophilia found in most crypts).

Figure 1

Crypt eosinophilia, intraepithelial cell hyperplasia and intracryptal budding found in inflamed colon mucosa. (A) Colon crypts with eosinophilia. Note the occasional goblet cells (H&E, original ×4). (B) Detail of crypt eosinophilia showing eosinophilia in columnar cells (H&E, original ×20). (C) Another crypt with eosinophilic cells with one goblet cell (arrow) (H&E, original ×20). (D) Colon crypt with intense eosinophilia (H&E, original ×20). (E) Crypt with intraepithelial cell hyperplasia. Note the focal nuclear stratification (H&E, original ×20). (F) Crypt with intracryptal budding protruding into the crypt lumen (H&E, original ×20). (G) Another view from a crypt with intracryptal budding protruding into the crypt lumen (H&E, original ×10). (H) Crypt with two ‘face-to-face’ intracryptal buddings protruding into the crypt lumen. Note the small incomplete gap between the two arrows (H&E, original ×4).

Intraepithelial cell hyperplasia

These are intraepithelial hubs showing hyperplastic epithelium, easily recognised by focal nuclear stratification (figure 1E).

Intracryptal epithelial budding

These are epithelial growths bulging into the crypt lumen developing from a ‘mother’ crypt (figure 1F–H). Some intracryptal epithelial budding revealed a minor central lumen (figure 2A); other intracryptal epithelial budding develops into more distinct crypts attached to the ‘mother’ crypt (figure 2B,C).

Figure 2

Putative pathway from intracryptal budding with a minor central lumen to ‘crypt-in-crypt’ found in inflamed colon mucosa. (A) Intracryptal budding with a minor central lumen (arrow) (H&E, original ×20). (B, C) Intracryptal budding connected with the ‘mother’ crypt; note the large central lumen (H&E, original ×20). (D) ‘Crypt-in-crypt’ without apparent connection with the ‘mother’ crypt (H&E, original ×20). (E) Another view of ‘crypt-in-crypt’ without apparent connection with the ‘mother’ crypt (H&E, original ×10). (F) Vertical crypt with a transversal line; a histological section cut at that level unveils the ‘crypt-in-crypt’ mystery (H&E, original ×10).

Intracryptal supernumerary crypts

In cross-cut sections, intracryptal supernumerary crypts were epitomised by a luminal crypt ring without apparent connection with the ‘mother’ crypt (figure 2D,E). Figure 2F shows a crypt with a transversal line; a histological section cut at that level unveils the ‘crypt-in-crypts’ mystery, such as in figure 2D,E. Growing intracryptal supernumerary crypts adopted bizarre pretzel or 8-like profiles (figure 3A). Figure 3B shows a bent crypt with U-turn configurations and a transversal line; a histological section cut at that level would reveal one cross-cut crypt with a pseudo-epithelial bud (figure 3C). Figure 3D shows vertical crypts with bizarre profiles and a transversal line; a histological section cut at that level would reveal cross-cut ‘two-crypts-in-crypts’ (figure 3E).

Figure 3

Intracryptal supernumerary crypts found in inflamed colon mucosa. (A) Low-power view colon biopsy showing vertical crypts with bizarre crypts growing inside the ‘mother’ crypt. Note that the internal crypts assume U-turn or pretzel/8-like configurations (H&E, original ×4). (B) A closer view of vertical crypts with intracryptal supernumerary crypts unveiling U-bent configuration and a transversal line; a histological section cut at that level would reveal one cross-cut crypt with a pseudo-epithelial bud (C). (D) Vertical crypts with bizarre pretzel/8-like profiles and a transversal line; a histological section cut at that level would reveal cross-cut ‘two-crypts-in-crypts’ (E) (B, D: H&E, original ×10; C, D: H&E, original ×20).

Intraluminal epithelial bridges

These are one or more bands of epithelium found across the crypt lumen, dividing the lumen into ≥2 compartments (figure 4A,B).

Figure 4

Intraluminal epithelial bridges, crypt rings in rows and off-centre epithelial budding found in inflamed colon mucosa. (A) Two bands of epithelium (arrows) across the crypt lumen, dividing the lumen into ≥2 compartments (H&E, original ×10). (B) Closer view of crypts with intraluminal epithelial bridges (H&E, original ×20). (C) Crypts in rows in the upper portion of the colon mucosa (H&E, original ×10). (D) Another view of crypts in rows or files in the upper portion of the colon mucosa (H&E, original ×10). (E, F) Close view of two crypts with off-centre epithelial budding (H&E, original ×20).

Crypt rings in rows

These are crypts with several intraluminal epithelial bands, building a chain of crypt rings in file (figure 4C,D).

Off-centre epithelial budding

These are buds merging from the lateral aspect of a crypt, bulging into the lamina propria. Off-centre epithelial budding was mainly found in vertically oriented sections (figure 4E,F).

Degree of mucosal inflammation

Mucosal inflammation was classified by semiquantitative estimates into (1) slight-moderate mucosal inflammation and (2) severe mucosal inflammation.

Statistical analysis

Non-parametric Mann-Whitney U two-tailed test was applied to compare differences between two independent groups. Statistical significance was defined as p<0.05.

Results

The results are condensed in table 1.

View this table:

Table 1

Frequency of crypt-associated anomalies found in H&E-stained sections of 100 cases with ulcerative colitis, 50 cases with infectious colitis and 29 cases with ulcerative colitis in remission