Old and New Diagnostic Paths in Inflammatory Bowel Diseases

Abstract

In the setting of the inflammatory bowel diseases, the role played by pathologists in the histological assessment of mucosal biopsy can be ideally divided into four steps. As a sequential proceeding, the histological evaluation has to deal with (1) the identification of the intestinal inflammation as consistent with ulcerative colitis or Crohn’s disease, distinguishing them from their own mimickers; (2) the assessment of the inflammatory activity, thus highlighting their main descriptors, also including grade and distribution; (3) the evaluation of the histological features suggesting treatment response; (4) the histological detection of dysplasia and other complications in long-standing diseases. Requirements that should support the biopsy interpretation include (1) the thorough endoscopic data from the colonoscopy; (2) a comprehensive mucosal mapping; (3) a proper methodological approach to the mucosal specimens during the pre-analytical phase; (4) the methodological approach to the histological evaluation. In the last few years, efforts have been undertaken to identify novel histological markers of mucosal dysfunction in inflammatory bowel disease, which could help to identify novel markers of mucosal healing.

1. Introduction

The term inflammatory bowel disease (IBD) refers to ulcerative colitis (UC) and Crohn’s disease (CD), two chronic relapsing diseases involving the gastrointestinal (GI) tract and deeply affecting the patient’s quality of life. Genetic, immunological and environmental factors have been advocated as pathogenetic contributors [1]. Recently, the weakness in the mucosal barrier has been proposed as an early causative co-agent in IBD onset [2]. The age of onset of UC and CD is variable and albeit adults are more commonly affected, both early and late onset can be seen in children and elderly respectively [3]. The main treatment goal in IBD is represented by the stable clinical remission, characterized by both symptomatic relief and mucosal healing. The medical treatment in IBD is mostly based on immunosuppression, through the administration of both anti-inflammatory and immunosuppressive drugs [4]. The suboptimal treatment and the long-lasting duration of the disease increase the risk of complications, including IBD-associated dysplasia and colorectal cancer (CRC). Both UC and CD are multifaceted diseases that need to be managed by multi-disciplinary teams [5].

2. Histological landmarks

2.1 Methodological approach

Before highlighting the main diagnostic clues in IBD, the methodological approach, encompassing both endoscopic and technical recommendations, should be considered. Notably, mucosal sampling should be performed on six gut segments, including the terminal ileum, cecum-ascending, transverse, descending, sigmoid colon and the rectum, collecting at least two pinch biopsies per each segment [6]. Then, mucosal specimens should be placed onto an acetate cellulose filter, in order to maximize the specimen’s orientation [7, 8]. Since CD is known to not be limited to the gut, the clinical features can prompt to extend the endoscopic evaluation to the whole GI tract. In such cases, collecting two biopsies in both the esophagus and the duodenum, and five biopsies in the stomach, according to the Sydney scheme, is advisable [9]. The role played by processing and cutting should not be down-placed. Samples must be processed with the filter. A 90 degrees rotation of the filter-mucosal specimen must be performed before the embedding, in order to provide the proper trans-sectional orientation and to minimize manipulation artifacts. Relevant clinical, radiological and laboratory data should also be collected [8, 10]. Overall, the optimal recognition of the diagnostic clues of IBD, mostly the architectural abnormalities and basal plasmacytosis, also relies on methodology.

2.2 Ulcerative colitis

The UC can involve the colorectum with different extensions at the onset, ranging from proctitis and procto-sigmoiditis to left-sided disease and pancolitis. Disease progression across the colorectum can also occur. Unusual presentations include rectal sparing and the so-called cecal patch. Terminal ileum is spared by definition and, if involved, the suspicion of CD should be raised. However, mild inflammatory infiltration of terminal ileum can be observed mostly in pancolitis, and it is composed by lymphocytes, few plasma cells and neutrophils. Such finding is referred to as backwash ileitis and represents a reactive phenomenon that should not be misinterpreted as CD [11, 12, 13]. The histological features in UC can be grouped into architectural and inflammatory abnormalities. The architectural abnormalities refer to spatial alterations of glands, including glandular distortion, characterized by gland branching, fused or angulated glands and pseudo-villous mucosal surface. Mucosal inflammation is usually diffused in treatment-naïve UC, and it can be subdivided into non-active inflammation and neutrophil-mediated (active) glandular injury [8]. Non-active mucosal inflammation is composed by scattered lymphocytes, plasma cells and eosinophils into the lamina propria. Mucosal samples from active UC commonly exhibit abundant plasma cells infiltrate throughout the mucosal lining, and the involvement of the lower third (known as basal plasmacytosis) constitutes, along with the finding of architectural abnormalities, a diagnostic clue to distinguish UC from acute self-limiting colitis. Basal plasmacytosis is the strongest predictor of IBD, although differences in its segmental distribution were highlighted in UC and CD [14]. Few eosinophils are present in the healthy gut, with a slight predominance in the right colon and a gradient fading out towards the left colon. Increased eosinophils are detected in treatment-naïve UC, intermingled with plasma cells. The number, degranulation, clustering and intra-epithelial location of eosinophils have been related to a less responsiveness to corticosteroid drugs in UC patients [15].

Active inflammation is qualified by the presence of neutrophils into the lamina propria, also producing injuries on glands (cryptitis and crypt abscesses) and surface epithelium (erosions and ulcerations) [8]. Inflammatory pseudo-polyps result from severe mucosal damage in more aggressive diseases, representing the end-stage of the superficial epithelium injury. On the other hand, crypt lysis and atrophy, sometimes paired to cryptolytic granulomas, mark the end-stage of glandular injury [10, 16]. The active inflammation should be histologically graded and the three-tiered scheme, encompassing the mild, moderated and severe grades, is advisable (Figure 1) [17, 18].

Figure 1.

A-B-C: Crohn’s disease with the typical granulomas. H&E x 20. D-E-F: Ulcerative colitis D-E active H&E Dx10, E x 20, Fx10. G-H-I: Autoimmune colitis H&E Gx10, Hx20, Ix40 in the blu circle apoptotic body.

2.3 Crohn’s disease

The CD is characterized by trans-mural inflammation of the terminal ileum, but every segment can be involved in the GI tract. Architectural changes are commonly patchy, consisting in villous blunting in the small intestine, whilst glandular distortion is less prominent than that in UC and may involve few glands [8]. As in UC, non-active inflammation is composed by lymphocytes, plasma cells and eosinophils. The presence of neutrophils in the lamina propria qualifies the active inflammation and leads to epithelial injuries consisting in both glandular and surface lesions, even though gland cryptitis and abscesses are less extensive as compared to UC.

The discontinuous mucosal involvement represents another critical feature distinguishing CD from UC, resulting in patchy or segmental inflammation. Moreover, basal plasmacytosis is a less characteristic finding than in UC [19, 20]. The early lesion commonly consists in neutrophil infiltration of the superficial epithelium in the terminal ileum, leading to mucosal erosion and ulceration. Such erosive lesions can involve the mucosa overlying Peyer’s patches, producing a peculiar histological feature called aphtoid ulcer [21]. The glandular injury leading to the mucus leakage in the lamina propria can evoke a local granulomatous reaction, promoting granuloma formation as in UC. However, such cryptolytic granulomas must be distinguished by typical epithelioid and non-caseating granulomas in CD.

Granulomas represent a characterizing feature in CD and are composed by discrete clusters of epithelioid histiocytes with intermingled T-lymphocytes. Granulomas in CD should be distinguished by cryptolytic granulomas, which are found close to injured glands. Conversely, CD granulomas are commonly detected in the lamina propria or at the edges of the inflammatory infiltration, and not in contact with glands. The sequential sectioning represents a basic requirement for the differential diagnosis [22]. In contrast with granulomatous enterocolitis caused by infective agents (e.g. Micobacterium sp., Yersinia sp), in which necrotizing granulomas can be detected, the CD is characterized by non-necrotizing granulomas (Figure 1) [23].

2.4 Typical findings in early and late onsets

The onset of IBD commonly involves the adult patients, but early and late onset in children and elderly respectively can be observed in a subset of patients [24, 25, 26]. IBD onset has been estimated to occur in about 25% of cases during childhood or adolescence [27].

In paediatric UC patients, the disease onset is characterized by the extensive colonic involvement in the majority of cases. Nonetheless, unusual presentations with patchy disease including caecal patch, and rectal sparing are not infrequent in children [28]. The early mucosal lesions are characterized by increased inflammatory infiltration into the lamina propria, including eosinophils and basal plasma cells [29]. However, architectural distortion and crypt atrophy pointing to the chronic nature of disease in the adult counterpart, are usually lacking in patients younger than 11 years [30]. Reactive changes in the terminal ileum, the so-called backwash ileitis, occur in approximately 20% of children presenting with pancolitis [31]. It consists of blunting of intestinal villi and mild neutrophil infiltration, often with patchy distribution, occasionally associated with crypt injury [32]. Detecting granulomas or pyloric metaplasia can support in distinguishing backwash ileitis from CD [33].

The CD onset usually involves the terminal ileum. The colonic involvement is seen in a lesser number of cases, often with rectal sparing. Isolated gastro-duodenal disease is observed in less than 5% of patients [34]. The early mucosal lesion consists in the patchy inflammatory infiltration of the lamina propria, with basal plasmacytosis, increased eosinophils and neutrophils. Non-caseating non-cryptolytic granulomas are more frequently observed in children than in adults [35]. The differential diagnosis mainly includes infectious colitis and food allergy. Moreover, the autoimmune enteropathy should be considered as a mimicker of IBD, mostly in patients suffering from IPEX syndrome (Immune dysregulation, polyendocrinopathy, enteropathy X-linked) and primary immunodeficiencies [36]. Gland atrophy, crypt hyperplasia and apoptotic bodies allow to discriminate autoimmune enteropathy from IBD (Figure 1). On the other hand, common variable immunodeficiency or severe combined immunodeficiency are characterized by reduced plasma cells into the inflammatory infiltrate [37].

Elderly patients are characterized by frequent co-morbidities, and a late onset of IBD commonly worsens their vulnerability, giving rise to daily challenges in the clinical management. There are not significant differences at histology in late onset UC, as compared to adult onset. However, elderly patients have a raised prevalence of non-IBD colitis that typically affects that subset of patients (e.g. the segmental colitis associated to diverticula - SCAD), which should be cautiously excluded. The CD can affect elderly patients, both resulting from a long-standing disease and as a late onset. Worth noting, terminal ileum has been reported to be less commonly involved in the late onset CD, the colonic involvement more frequently observed [38]. Moreover, a drug-induced injury should be considered in the differential diagnosis of Chron’s ileitis in the elderly [39].

2.5 The mucosal healing: concept, practice and future perspectives

Medical treatment represents the first approach to IBD patients, aiming at earning symptomatic relief, as well as endoscopic and histological disappearing of mucosal inflammation. The general concept of mucosal healing (MH) emerged as the major treatment goal in IBD, predicting long-term remission. Moreover, MH correlates with a lower risk of IBD-associated dysplasia and CRC, being also related to lower rates of hospitalization and surgical procedures [40, 41]. The endoscopic MH is focused on the disappearing of inflammatory mucosal lesions at ileo-colonoscopy. However, there are no valid studies encompassing clinical-histological correlations on the topic [42]. Particularly, the histological features cannot be inferred by endoscopy, and the histological assessment of MH should be improved by pathologists, promoting simplified and highly reproducible scoring systems, as well as novel biomarkers.

At histology, neutrophil infiltration into the lamina propria is the most reliable histological marker of activity in IBD, also including neutrophil-mediated epithelial injuries. Hence, the absence of neutrophils and neutrophil-related mucosal injuries can be considered the most reproducible marker of histological MH. Furthermore, plasma cell infiltration is commonly seen in the lamina propria in IBD specimens and it is referred to as a marker of chronic inflammation [8]. However, the persistence of subtle plasma cell infiltration does not stand for active disease in treated patients, but conversely, it highlights the persistence of chronic inflammation [43]. Overall, the presence of plasma cells into mucosal biopsies from treated patients does not exclude the histological definition of MH, as long as neutrophils are absent.

During the last few years, efforts have been undertaken in order to identify markers that could support the histological definition of MH in IBD. The glucocorticoid-induced leucine zipper (GILZ) has been established to mediate glucocorticoid effects in immune cells, including T lymphocytes, B lymphocytes, macrophages and neutrophils [44, 45, 46, 47]. Several in vivo models have been used to study the role that GILZ plays in inflammation, including IBD. Genetic ablation studies demonstrated the protective function of GILZ in IBD, due to the reduction of immune cell recruitment and activity in inflammation [48, 49]. Moreover, the administration of the recombinant GILZ protein has been proven to ameliorate symptoms of colitis in animal model studies, representing a novel pharmacological tool in IBD [50, 51]. Interestingly, GILZ has been recently established to play a secretory role in the gut, mostly involving goblet cells and enteroendocrine cells. Particularly, goblet cells showed a basal GILZ expression in normal colonic mucosa, whilst it was affected by active inflammation in IBD patients, with the expression reduced in active disease and restored in clinical remission [4]. Such GILZ dynamics correlated with neutrophil infiltration into the lamina propria, which represents the only reliable histological marker of activity in IBD. Importantly, the reduction or lack of GILZ in neutrophils exacerbates experimental colitis [46, 52]. Overall, GILZ expression can be considered a protective factor in the gut, its down-regulation marks the active phase of disease and its restoration strongly correlates with remission in IBD.

Furthermore, based on the recognition of the mucosal weakness as one of the major promoters of UC onset [2], colon-specific mucins (MUC) have been investigated in order to identify molecular traits of mucus barrier dysfunction. A novel signature involving MUC1, MUC4 and MUC5B derangements was found to characterize treatment-naïve UC patients. Moreover, the mucin expression profile was also found not to be restored in treated patients, who expressed the same types of mucin derangement. Notably, both relapsing and remitting patients showed commonalities in mucin expression profile with onset patients. Hence, evaluating the MUC1, MUC4 and MUC5B signatures should be integrated into the histological assessment of mucosal biopsies in treated UC patients. The assessment of gut mucin restoration should be integrated in the routine clinical practice as a functional descriptor of the histological MH. Worth-nothing, the proposed signature was also highlighted to represent an exploitable actionable target for specific MUC enhancers, opening novel treatment perspectives towards a synergistic approach, including immunosuppressive agents and mucin agonists [53].

3. The scoring systems in IBD

Several scoring systems have been proposed over time, aiming at grading disease activity and identifying histological MH. Such scoring systems considered and scored several histological parameters, including variations in inflammatory infiltrate, architectural abnormalities and epithelial injuries. However, validation studies are lacking for the most part of them, and their application into the routine clinical practice resulted to be complex, time-consuming and burdened by subjectivity [54, 55]. A Simplified Histological MH Scheme (SHMHS) has been recently proposed, being suitable for both CD and UC. It consisted in a two-steps approach, aiming at considering neutrophil infiltration as the major histological descriptor of activity, and gut segments. The SHMHS exhibited a high correlation rate with the endoscopic findings and resulted in simple, time-sparing and suitable for the clinical practice (Table 1) [43, 56, 57].

Features	Value	Score
Neutrophil infiltration into the lamina propria	Present	1
	Absent	0
Cryptitis and crypt abscesses	Present	1
	Absent	0
Erosion and ulcerations	Present	1
	Absent	0
Gut segments	Value	Score
Terminal Ileum (for CD only)	Quiescent	0
	Not involved	0
	Active	1
Right colon	Quiescent	0
	Not involved	0
	Active	1
Transverse colon	Quiescent	0
	Not involved	0
	Active	1
Descending colon	Quiescent	0
	Not involved	0
	Active	1
Sigmoid colon	Quiescent	0
	Not involved	0
	Active	1
Rectum	Quiescent	0
	Not involved	0
	Active	1

Table 1.

The simplified histological mucosal healing scheme (SHMHS).

4. IBD-associated dysplasia and colorectal carcinoma

IBD is a well-recognized risk factor for CRC, with the risk of malignant transformation referred to be higher than that in a non-IBD population [58]. Increased risk of high-grade dysplasia and CRC in IBD patients with recurrent low-grade dysplasia [58]. Long-standing disease and ineffective treatments expose enterocytes to progressive oxidative stress, leading to mutagenic activity and carcinogenesis [59]. Particularly, p53 mutations mark the early phase of IBD-related carcinogenesis, in contrast with conventional CRC in which such mutations are found in the invasive phase [60]. Endoscopic surveillance plays a critical role in detecting early dysplastic lesions in IBD, monitoring clinical course and planning clinical management. The endoscopic features are listed into the scenic classification and encompassed both visible and not visible lesions. Visible lesions can present as polypoid, flat and depressed, as summarized in Table 2. Detecting dysplasia at histology represents the most reliable marker of malignant transformation in IBD patients, supporting clinical management and treatment planning. Worth noting, dysplasia that is found within the disease field, and referred to as dysplasia associated lesion or mass (DALM), should be distinguished from the so-called adenoma-like mass (ALM), that arises outside, in bowel segments spared by inflammation, and that can be managed as sporadic adenomas. Unfortunately, detecting IBD-related dysplasia is burdened by high inter-observer variability, mostly in low grade and serrated lesions, and a second look is advisable to ensure the proper management of these patients [5].

Lesion type	Classification		Borders	Ulceration
Non-visible
Visible	Polypoid	Pedunculated	Distinct	Present
		Sessile	Indistinct	Absent
	Non-polypoid	Superficial elevated
		Flat
		Depressed

Table 2.

Scenic classification of endoscopic findings in IBD patients.

5. The differential diagnosis in IBD

5.1 Drug-induced colitis

Drug-induced colitis (DIC) is an inflammatory condition in which the correlation with drugs administration is proven. DIC represents a prevalent cause of colitis in a specific population subset, mostly composed by elderly patients, often characterized by several vulnerabilities and co-morbidities [25]. Thus, the history of drug use should be clinically investigated, particularly in that subset of patients. Drugs and chemical compounds, including antibiotics, non-steroidal anti-inflammatory drugs (NSAIDs), laxatives, anticancer drugs and immunosuppressive agents, can induce cellular toxicity through allergic reactions and vascular insufficiency [61]. Watery diarrhea is the most complained symptom, whereas bloody discharge is seldom found and, if any, it is usually mild. Endoscopic findings can vary, ranging from unremarkable features to mucosal oedema and erythema with or without mucosal erosions. Worth noting, a normal-looking mucosa at endoscopy does not stand for healthy mucosa, thus a thorough mucosal sampling during ileo-colonoscopy is advisable [6].

The diagnosis of DIC needs thorough clinical-pathological correlations and the identification of one or more inflammatory patterns at histology (Table 3).

			Histological features
DIC Patterns	Lamina propria	Cell composition*	Glandular features	Histological hallmarks
Pseudo-melanotic	Hypocellular	Lymphocytes, plasma cells (few), histiocytes.; Granulocytes uncommon.	Architectural sparing.#; Epithelial injury uncommon.	Lipofuscin-laden histiocytes.; Basal plasma cells absent.
Hyper-eosinophilic	Mild hyper-cellular	Eosinophils, lymphocytes, plasma cells (few), histiocytes. Neutrophils uncommon§	Architectural sparing.#; Epithelial injury uncommon.	Eosinophil count (>60/10HPF).; Basal plasma cells absent.
Acute inflammation	Hyper-cellular	Granulocytes (neutrophils and eosinophils), Lymphocytes, plasma cells (few), eosinophils (few).	Architectural sparing.; #Epithelial injury common.	Acute inflammation with neutrophil-mediated epithelial injury. Basal plasma cells absent.
Ischemic	Hypocellular, Fibrotic	Lymphocytes, histiocytes, eosinophils (few).; Neutrophils uncommon§	Architectural sparing.; Atrophic changes	Atrophic glandular changes in fibrotic stroma. Basal plasma cells absent.
GVHD-like	Hypocellular	Lymphocytes, plasm cells eosinophils (few), histiocytes. Neutrophils uncommon§	Architectural sparing.; Glandular hyperplasia	Raised apoptotic activity in glandular epithelium. Basal plasma cells absent.
Lymphocytic	Hypercellular	Lymphocytes, plasm cells eosinophils (few), histiocytes. Neutrophils uncommon§	Architectural sparing.; Intra-epithelial T-lymphocytes	Diffuse intra-epithelial T- lymphocytes (>20/100 epithelial cells). Basal plasma cells absent.
Collagenous	Hypocellular; Mild; Hyper-cellular	Lymphocytes, plasm cells eosinophils (few), histiocytes.	Architectural sparing.; Intra-epithelial T-lymphocytes (few)	Subepithelial collagenous band thickening (>10 μm).; Basal plasma cells absent.

Table 3.

Synopsis of the histological features in drug-induced colitis (DIC), according to the histological pattern.

*

Cell composition can vary according to the overlap with other DIC patterns;

^§

unless an erosive process is associated, mostly in terminal ileum; and.

^#

referring to cryptitis and crypt abscesses, surface erosions and ulcerations.

5.1.1 Pseudo-melanotic pattern

Lymphoid cells are commonly found in the gut mucosa, and their composition varies throughout the GI tract, including lymphocytes, granulocytes and histiocytes. Histiocytes are commonly found as scattered in the lamina propria and their foamy-like appearance is unusual in healthy mucosa. In the setting of DIC, the pseudo-melanotic pattern is characterized by the accumulation of histiocytes in the colonic mucosa, mostly in the upper third, but diffuse engulfment of the lamina propria can be seen in cases of long-term exposure to drugs. Pigment-laden histiocytes represent the most common finding, and can be found in loco-regional lymph nodes as well. Such lipofuscin-like pigment results from epithelial cell apoptosis with subsequent histiocytic phagocytosis occurring during contact laxative use (e.g. anthraquinone-containing drugs), even though it has also been described in NSAIDs abusers [61, 62]. Inflammatory infiltration is usually slightly increased into the lamina propria, but basal plasma cells are usually absent. The pseudo-melanotic pattern of injury can be also found to be associated with other patterns of DIC, particularly the focal active, hyper-eosinophilic and lymphocytic patterns. In those cases, the cellular composition varies in the lamina propria due to the enrichment of neutrophils, eosinophils and lymphocytes respectively. Glandular architecture is spared and there is not significant mucin depletion. Worth noting, the so-called “melanosis coli” has been linked to an increased risk of colonic dysplasia and CRC [63, 64].

5.1.2 Hyper-eosinophilic pattern

Colonic mucosa harbors eosinophils with a gradient decrease from the right to the left colon, and scattered eosinophils can be found in the small bowel as well. The main diagnostic clue in the hyper-eosinophilic pattern is detecting more than 60 eosinophils/10 high power fields [HPF] without peripheral hyper-eosinophilia [65]. The inflammatory infiltrate is slightly increased in the lamina propria, including lymphocytes and plasma cells, but basal plasmacytosis is usually absent. The hyper-eosinophilic pattern can occur alone or pair with other DIC patterns, including the pseudo-melanotic, lymphocytic and collagenous patterns [25]. In contrast with eosinophilic gastro-enteritis, eosinophilic colitis is a non-IgE-associated disease, with watery diarrhea as the most common clinical presentation [66]. Once parasitic infestations have been excluded, clinical-pathological correlations should be undertaken to distinguish drug-related colitis from gut diseases with an allergic etiology [65].

5.1.3 Acute inflammation pattern

This inflammatory pattern characterizes a broad and heterogeneous group of colitis, with different histological features, clinical presentation and causative agents. They range from focal acute colitis to pseudo-membranous and IBD-like colitis. Several drugs have been implicated, such as NSAIDs, antibiotics and laxatives [61].

In focal active colitis, the main histological clue is represented by neutrophil infiltration into the lamina propria, also promoting epithelial injuries (e.g. cryptitis, mucosal erosion). As focal active ileitis, it could give rise to the suspicion for CD onset. Worth noting, clues of chronic gut disease, including basal plasma cells and architectural distortion, are usually absent. Moreover, the presence of surface mucosal erosion along with a history of drugs use favor a drug-related etiology.

Pseudo-membranous colitis is a peculiar form of active colitis linked to Clostridium difficile infection, with both antibiotic therapy and older age representing the major risk factors. Pathologists should be confident with mucosal lesions in different evolutive phases. Particularly, the earliest type I lesion is rare and characterized by focal fibrin accumulation at the mucosal surface, oedematous lamina propria with scattered neutrophils and eosinophils. A small amount of weakly eosinophilic sub-epithelial exudate, surface erosion and focal cryptitis can be seen during this phase. In the type II lesion crypts are filled by eosinophils. The type III mucosal lesion represents the most severe mucosal injury, with diffuse coagulative necrosis, mucus accumulation, fibrin strands, cellular debris, neutrophils and eosinophils, constituting the typical pseudo-membranes. Type II and type III lesions should be distinguished from ischemic injury. Detecting collections of eosinophils in the crypt lumen aids the differential diagnosis, especially when a proper clinical-pathological correlation has been provided [67, 68, 69]. Some chemical compounds (e.g. gold salt) and drugs (e.g. NSAIDs, Micofenolate Mofetil) can produce an IBD-like pattern of injury consisting in some histological features frequently found in IBD, both in UC and CD. Active inflammation with surface mucosal erosions and ulcers paired to focal architectural abnormalities can be detected in the colonic mucosa, whilst the terminal ileum can present a cobblestone appearance resembling Crohn’s ileitis [70, 71].

The acute inflammation pattern can be seen in combination with other DIC patterns of injury, most of all the hyper-eosinophilic and pseudo-melanotic patterns.

5.1.4 Ischemic pattern

Ischemic colitis can affect both young and elderly patients due to vascular and iatrogenic causes. Iatrogenic colitis with ischemic pattern is characterized by necrosis and inflammation, resulting from mucosal hypoxia. It results from non-occlusive etiology based on vasoconstrictive effect of drugs on small vessels [61, 72, 73]. Cocaine, amphetamines, diuretics and oral contraceptives have been implicated in this pathogenic mechanism [74, 75]. Glandular withering and coagulative necrosis characterize the mucosal injury at histology. Vascular congestion, haemorrhagic foci and capillary microthrombi may be also detected into the submucosal layer. Inflammatory infiltrate is usually mild, consisting in lymphocytes, plasma cells, histiocytes, and eosinophils, with a variable number of neutrophils. Basal plasma cells are usually absent. Architectural distortion is not a prominent finding, whereas small atrophic glands, lined by irregular cells with dark-stained nuclei, and variable degrees of fibrosis in the lamina propria can be found [76].

5.1.5 Apoptotic (GVHD-like) pattern

In transplant patients, the use of immunosuppressive agents (e.g. Micofenolate Mofetil) can give rise to adverse events including graft versus host disease (GVHD) and GVHD-like mucosal injuries. The same pattern of injury has been reported in patients who have not undergone transplants [77]. It refers to a peculiar mucosal damage caused by anti-inflammatory (e.g. anti- tumor necrosis factor (TNF) α, anti-interleukins/interleukin receptors) and anti-tumour drugs (immune checkpoint inhibitors, anti-CD20 and anti-endothelial growth factor). Diarrhea is the most frequently mentioned symptom and it is due to GI toxicity [78]. It should be noted that drug toxicity can cause other types of mucosal injury, including autoimmune enteropathy, acute and fulminant colitis, ischemic colitis, UC-like and CD-like inflammation. The diagnostic clue of the apoptotic pattern is represented by apoptotic epithelial cells at the base of the crypts [79, 80, 81].

5.2 Microscopic colitis

Microscopic colitis is a clinical term used to identify a clinical setting characterized by watery diarrhea and unremarkable endoscopic findings. Lymphocytic and collagenous colitis are the two histological findings in such cases, even though mixed forms can be seen. Microscopic colitis has several causative factors, including drugs (e.g. NSAIDs, PPI, ticlopidine) [82]. In lymphocytic colitis, the main diagnostic clue at histology is represented by diffuse infiltration of T (CD3+; CD8+)-lymphocytes (more than 20/100 epithelial cells) into the surface epithelium, the lamina propria is usually hypercellular, with neither architectural effacement nor basal plasma cells. On the other hand, collagenous colitis is qualified by thickening sub-epithelial collagen bundle (more than 10 μm), and inflammatory cells may present as either mildly or heavily increased into the lamina propria, but architectural abnormalities and basal plasma cells are usually unseen [83, 84]. Both lymphocytic and collagenous colitis can be found in combination with other DIC patterns of injury.

5.3 Infectious colitis

Infectious colitis is a self-limiting inflammatory disease involving the colonic mucosa, which may spontaneously resolve after time, most commonly a few weeks. Depending on the etiology, the onset of the disease is characterized by the increased number of inflammatory cells into the lamina propria, composed by lymphocytes, plasma cells, eosinophils and neutrophils. Epithelial injuries can be seen and could represent a prominent feature in the early phase of disease. Neither architectural distortion nor basal plasma cells are detected at histology, both representing a diagnostic clue for the differential diagnosis with active IBD. Moreover, some infectious species can promote histological features resembling IBD, making misdiagnoses possible. Particularly, colitis due to Campylobacter infection can be difficult to distinguish from active UC, whilst other microorganisms (e.g. Shigella species) can be associated with mucosal granulomatous reaction, suggesting to rule out CD [85]. Biopsy interpretation of infectious colitis relies on the absence of histological features pointing towards a chronic gut inflammation, particularly the absence of crypt distortion. Depending on the time of biopsy, we could have to deal with the resolving (late) phase of disease, presenting with different features at histology. The resolving phase of infectious colitis has a less cellular lamina propria, neutrophils are usually absent and, if any, they are scattered in the lamina propria and glandular damage is seldom found. The main diagnostic clue for the resolving phase is the recognition of increased apoptotic activity in the surface epithelium, which exhibits nuclear dust in a variable amount. Scattered intraepithelial lymphocytes (less than 20/100 epithelial cells) and regenerative hyperplastic changes in the surface and glandular epithelium can also be seen at histology [68, 86].

5.4 Segmental colitis associated to diverticulosis (SCAD)

The development of intestinal diverticula usually occurs asymptomatic. In symptomatic patients, medical treatment should be attempted, as significant morbidity may be associated with resection, especially in the elderly [87]. The SCAD refers to an inflammatory condition in which the sigmoid harboring diverticula shows different degrees of architectural distortion and inflammation, in a segmental fashion [88]. The inflammatory infiltrate is composed of lymphocytes, plasma cells, eosinophils and histiocytes. Basal plasma cells can be seen and neutrophilic infiltration promotes epithelial injuries [89]. Distinguishing SCAD from UC can be challenging on biopsy specimens, but the segmental distribution of inflammation represents the main diagnostic clue, since the rectum is spared by definition in SCAD, whereas it is involved in UC. Thus, the thorough mucosal sampling including biopsies from at least the rectum to the descending colon should not be omitted, in order to highlight the segmental nature of the inflammation in SCAD.

5.5 Ischemic colitis

Ischemic injuries can affect the large bowel following any of the following patterns: gangrenous colitis, haemorrhagic entero-colitis and ischemic colitis, the latter being due to subacute ischemic damage. It usually occurs in elderly patients mostly due to vascular causes. Hypoxia can produce two types of acute mucosal damage (gangrenous colitis and haemorrhagic entero-colitis) and a subacute injury (ischemic colitis) [90]. Young patients may also be affected due to drug abuse or systemic diseases (e.g. Behçet disease, systemic lupus erythematous) [91, 92]. It is characterized by mucosal injury, epithelial withering and coagulative epithelial necrosis at histology. In the submucosal layer vascular congestion, haemorrhagic foci and capillary microthrombi may be detected. Inflammation is usually mild and mainly composed by lymphocytes, histiocytes and granulocytes. Basal plasma cells are absent. Atrophic features of the glands, including clusters of small-sized glands with dark nuclei and condensed cytoplasm should not be misinterpreted as architectural distortion. Moreover, interstitial fibrosis can be found in the setting of chronic injury [76].

5.6 Radiation colitis

Radiation colitis is a treatment-related condition due to therapeutic irradiation of the abdomen, causing mucosal damage. The acute injury is represented by acute mucositis, whereas the chronic phase is characterized by progressive vascular damage and fibrosis [93]. Short-term lesions consist in mucosal inflammation and crypt abnormalities, epithelial reactive changes and apoptotic bodies are easily detectable in both the surface and glandular epithelium. Delayed injury can be subdivided into epithelial and stromal lesions, the former consisting in glandular alterations in size and shape, also exhibiting reactive epithelial changes that should not be misdiagnosed as dysplasia. Stromal alterations include fibrosis with atypical fibroblasts, and mild inflammation. Capillaries are usually dilated, with swollen endothelial cells and micro-thrombi. Small-sized arteries may show various degrees of lipid-laden macrophage into the intimal layer, wall hyalinization and fibrosis [94].

6. Conclusions

Managing patients suffering from IBD represents a daily challenge that must be addressed in the main clinical scenarios, consisting in 1) histological diagnosis; 2) evaluation of treatment response; 3) management of long-term complications, also including IBD-associated dysplasia and CRC. A multidisciplinary approach is advisable in every IBD clinical settings.

Acknowledgments

The present chapter is dedicated to Doctor Vincenzo Villanacci for the 10 years-long mentorship on gastrointestinal pathology, with gratitude.

Giuseppe Leoncini, MD.