Visual diagnosis, biopsy sampling, handling, and histopathology

Despite the nonbiopsy strategy adopted in recent years for symptomatic patients, endoscopy remains essential for most children suspected for CD – for patients with no or vague symptoms, with signs of malabsorption without symptoms, with extraintestinal manifestations, in risk groups and healthy family members. However, compared to adults, where endoscopy can also be considered a case finding tool, its role in pediatric CD has been merely for mucosal sampling to get biopsy specimens for diagnosis. The same endoscopic features of CD that have been described in adult patients have been reported in children also – mucosal mosaic pattern, scalloping, visible vascular pattern or reduction of duodenal folds; these endoscopic markers however have low sensitivity in children with CD [3,4].

Over the years, endoscopic visualization of the duodenal mucosal pattern has improved, so that today one can see the villi up close, with the aid of advanced endoscopic techniques, as is the case with water immersion images as shown in Figures 23.1a and 23.1b, corresponding to histopathological views of normal mucosal morphology (Figure 23.1c) and subtotal villous atrophy. Also demonstrated is the pathognomonic feature of a manifest lesion in CD (Figure 23.1e). Endoscopists may nowadays set a real‐time diagnosis of clear villous atrophy, confirming, correcting or even replacing the pathologist’s work. However, endoscopy cannot assess for crypt hyperplasia thus missing early developing disease with villi still normal looking and tall (Figure 23.1a and 23.1d). This was true also for properly oriented biopsies evaluated under liquid surface using high‐power dissecting microscope showing the mucosal morphology continuum from normal to “flat” (Figure 23.1a–d). Crypt hyperplasia may already be occurring when the mucosa is visually evaluated as “normal” (Figure 23.1a and 23.1b). At a later stage when crypt hyperplasia is more prominent, the villi merge and convolute to low villous ridges (Figure 23.1c) and further to the gluten‐induced so‐called “flat” lesion, the endstage of the mucosal pathology continuum (Figure 23.1d). When biopsies showing low villous ridges are cut wrongly (tangentially), artificially tall villi are seen on histology slides, evaluated to represent normal mucosa of grade Marsh 0 or I when actually being Marsh 3b or 3c [5] (Figure 23.1e). Studies correlating visual endoscopic appearance to proper morphometric evaluations are lacking.

Gluten‐induced duodenal mucosal lesions can be patchy, depending on the time and amount of gluten ingestion. This is where novel endoscopic techniques such as chromoendoscopy come in, allowing for targeted biopsies in the diseased mucosa. In routine clinics the diagnosis is based on the most severe lesions of the sampled biopsies.

Visualization of villi using modern endoscopic techniques is not widely available, and also because of missing comparative studies, traditional histological assessment of biopsies is needed. CD guidelines support a multiple biopsy protocol, two specimens from the bulb and four from the distal part of the duodenum, but these recommendations are poorly followed in routine practice [6–9], mostly because of prolonged procedural time [8,10]. Therefore a multibite biopsy technique has been suggested, but a study in adults proved that the single‐biopsy technique is preferred, yielding a higher proportion of well‐oriented duodenal biopsy specimens [11]. As mentioned, the orientation and correct vertical cutting of biopsy specimens are of paramount importance as neither morphometric results nor Marsh class can be given if crypts are cut in cross‐section only.

Biopsy orientation on acetate cellulose filters in the endoscopy room has been recommended [12] but again, it is not enough to blindly press the biopsy on a filter strip resulting in messy fixed biopsies; proper orientation is needed (Figure 23.2). Recuttings are often demanded to find a representative number of intact villus‐crypt units to allow evaluation, so biopsies should be processed separately, not placed in the same paraffin block.

Duodenal bulb biopsies have been traditionally avoided because of concerns regarding their difficult interpretation due to histological confounders such as peptic injury, gastric metaplasia, Brunner’s glands or lymphoid follicles [13]. Current guidelines recommend taking biopsies from the proximal mucosa and the bulb, resulting in a diagnosis called ultra‐short CD [14]. However, a pediatric CD study has shown that bulb specimens are frequently of poor quality, tiny or having many Brunner artefacts, rendering them unreadable. Further morphological injury, even with crypt hyperplasia, is common in the duodenal bulb in nonceliacs, leading to false‐positive diagnoses [15]; for these situations, evaluation for bulb TG2‐targeted IgA subepithelial deposits is a powerful tool to confirm CD [15]. If bulb biopsies are taken, they should not be put in the same fixative vial with the distal duodenal biopsies.

Future of endoscopy in pediatric CD

The insufficient number, poor quality and bad orientation of biopsies, disease patchiness, caution regarding bulb samples, low sensitivity of endoscopic markers, and, most importantly, bad interobserver agreement between pathologists [16] have all cast a shadow on the use of endoscopy in the routine diagnosis of CD, despite efforts in overcoming each of them. The trend towards a nonendoscopic nonbiopsy diagnosis of CD is catching on, so that the future of endoscopy in CD might be limited for selected cases. One future need in children will be clinical drug trials using gluten‐induced mucosal injury as the primary outcome [17]. Modern endoscopy techniques with visual evaluation of the duodenal mucosa, including correlation with histopathology, determining the extent of the lesion, and evaluating the percentage of the lesioned mucosa, warrant further studies.