Classification of Murine Gastrointestinal Tumors
The following standards for histological assessment
came out of the MMHCC-sponsored symposium, "Mouse Models of Intestinal Neoplasia",
convened at the Jackson Laboratories with the purpose of bringing specialists together to
examine GEM with intestinal neoplasia systematically. The meeting was designed to allow a
panel including 7 pathologists and 4 basic scientists to examine and describe the morphology
of intestinal neoplasia in GEM, to develop standardized nomenclature for these lesions, to
develop recommendations for histologic handling of intestinal tissues from GEM, and to
compare the morphology of colorectal lesions from GEM to human colorectal neoplasia.
Nomenclature for Histologic Assessment of Intestinal Tumors in the Rodent
(Reprinted with permission from reference 58)
Hyperplasia: gross thickening of the mucosa; some growths may be pedunculated; mitosis
are always located in the lower two thirds of the mucosa; nuclei lack significant atypia, are
basally located, ovoid to round, usually uniformly dark, with occasional visible nucleoli
; crypts take on a star-shaped appearance in tangential sections; herniation (pseudoinvasion)
of epithelium through the muscularis mucosae may occur. Hyperplasia can be further categorized
as diffuse, focal, multifocal, associated with inflammation (type and severity noted), and also
can be graded as to severity
Aberrant crypt foci (ACF): microscopically in whole-mount colonic tissue that usually
is stained with methylene blue; one or more crypts larger than most crypts in the field, have
a thickened layer of epithelial cells that stain more intensely with methylene blue, often have
a slit-shaped luminal opening, have an increased pericryptal space, and are elevated from the
focal plane of the microscope (ACF are not a histologic diagnosis).
Gastrointestinal Intraepithelial neoplasia (GIN): histologically apparent areas of
dysplasia that are not visible grossly (<0.5-1.0 mm); in human and veterinary pathology, these
lesions may be referred to as microadenoma, microcarcinoma, carcinoma in situ, and focal areas
of dysplasia; lesions may involve single or multiple glands; is compatible with dysplastic
aberrant crypt foci (ACF) that previously have been identified in the same unembedded tissue
Adenoma: benign, circumscribed neoplasm composed of tubular and/or villous structures
lined by dysplastic epithelium; herniation (pseudoinvasion) through the muscularis mucosae may
occur; categorized by the following criteria:
1. macroscopic growth pattern: sessile, broad-based or pedunculated (polypoid)-attached by a stalk
2. histologic type: tubular - at least 75% of adenoma composed of branching tubules in lamina propria (usually pedunculated); villous - at least 75% of adenoma composed of leaf- or finger-like processes of lamina propria covered by epithelium (usually sessile; also known as papillary); tubulovillous - adenoma composed of 25-75% of both tubular and villous structures (usually pedunculated; also known as papillotubular)
3. grade of dysplasia: based on the most severely dysplastic area of each tumor
low grade: branching or elongation of crypts with some reduction of interglandular stroma; low N/C ratio; cell nuclei elongate, crowded, appear stratified, with regular nuclear membranes, fine chromatin, and inconspicuous nucleoli; nuclei maintain polarity with respect to the basement membrane; mucus secretion usually present
high grade: exhibits both architectural and cytologic changes; marked reduction of interglandular stroma with complex irregularity of glands with cribriform (sieve-like) structures and back-to-back glands; high N/C ratio; cell nuclei large, ovoid to round, with loss of polarity with respect to the basement membrane; cytologic atypia is more pronounced with marked, irregular nuclear membranes, with aberrant chromatin pattern not basally located (cleared, vesicular, clumped, or densely hyperchromatic chromatin); large, conspicuous nucleoli; mucus secretion usually absent; numerous mitoses with abnormal mitotic figures
Herniation: glands have penetrated through the muscularis mucosae (see Table 3).
Adenocarcinoma: malignant neoplasia of glandular epithelium composed of tubular and/or villous structures penetrating through the muscularis mucosa; categorized by the following criteria:
1. grade of differentiation: well differentiated, moderately differentiated, or poorly differentiated;
2. histologic type: tubular/tubulovillous/villous adenocarcinoma; mucinous adenocarcinoma (>50% of tumor composed of extracellular mucin, signet-ring cells can be present); signet-ring cell adenocarcinoma (>50% of tumor composed of signet-ring cells); undifferentiated carcinoma (no glandular structure to differentiate; can be uniform or pleomorphic)
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Summary Different GEM models of intestinal cancer display characteristic disease
manifestations. For example, lesions appear primarily in the cecum and colon in
some mouse lines and in the small intestine in others. Gross morphology is also
influenced by genotype; pedunculated polyps carry Apc mutations (regardless of
the initial mutation) while sessile lesions do not. Disease severity also differs
among the GEM models. For example, the Apc models and some of the MMR GEM have a
high incidence of adenoma but only rarely develop carcinomas. Others, such as the
Smad3 and Tgfβ1xRag2 mutants exhibit adenoma and carcinomas with equal frequency.
In these models carcinomas are observed more frequently in the large intestine than
in the small intestine.
Mouse models of intestinal cancer exhibit some important differences with respect to human
disease. In general, adenomas are the most common lesion in murine intestinal cancer models.
However, unlike humans, adenomas occur throughout the intestine in mice, with no predilection
for the large bowel. Another important difference is that metastasis is rare in mice.
Only one mouse line, Smad3-/-(83), exhibits metastasis of colonic adenocarcinomas to
lymph nodes and liver, which is a common occurrence in human colorectal carcinomas. There
is a critical need for a consistent model of metastasis.
Gut flora is an important modifier of intestinal tumor development. For example, when derived
in a germ-free environment, Il-10-/-, IL-2-/-, and Tgfβ1-/- X Rag2-/- mice do not develop
colonic lesions (77,91,94). Despite evidence of its importance, the role of the gut flora
is often overlooked in studies involving GEM. Understanding the role of bacterial flora in
the intestine is necessary to understand disease initiation and progression in murine models
of intestinal tumor formation. The precise role of bacteria in the development of lesions is
not understood, but is an important issue that should be addressed in each model with an
unexpected or invasive phenotype.
An important issue in interpreting the phenotypes of GEM is accurate discrimination between
invasive cancer and herniation. In the mouse, the muscularis mucosa is very thin, thus,
the mucosa is able to penetrate the muscle easily. Herniation can occur through weak points
where blood vessels and lymphatics traverse the muscle. Herniation can also result from
pressure exerted by the polyp that can push basal crypt cells through the muscularis mucosa.
Both of these mechanisms are enhanced by inflammatory conditions.
The Consensus report on the intestinal pathology of GEM (58) includes a classification system
(link to box: Features that distinguish…) to facilitate consistent and accurate distinction
between carcinoma and mucosal herniation. At least one of the following features should be
present to classify a lesion as a carcinoma: desmoplasia, loss of mucosal lining in invading
glands, and the presence of irregular, sharp or angulated glands. Particular caution is
necessary when evaluating inflamed areas because of the severe fibrosis and frequent disruption
and herniation of glandular tissue that occurs in many chronic inflammatory bowel disease
models.
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| Herniation of the intestinal mucosa of an Msh3-/- mouse. |
The invading cells include multiple cellular types, and there is no desmoplastic response in the herniated intestinal lesion. |
There is severe desmoplasia and loss of mucosa in the carcinoma. |
Features that Distinguish Invasive Carcinoma versus Herniation
(Reprinted with permission from reference 58)
1. Invasive cells are different from overlying mucosal component with atypia exceeding low-grade dysplasia.
2. Presence of desmoplasia that is not associated with a prominent inflammatory cell infiltrate.
3. Presence of irregular, sharp or angulated glands in invasive component.
4. Invading crypts spread laterally deep to the surface mucosal component.
5. Cell loss from invading mucosa.
6. More than two invading glands.
7. Absence of basement membrane around invading glands.
8. Evidence of progression to invasive cancer in other mice of the same genotype.
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For information on histology and cell culture techniques and other biological protocols see:
http://emice.nci.nih.gov/emice/resources/protocols/organ_site
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