macrophages.
Amyloidosis. Accumulation of amyloid occurs most commonly in mesenteric lymph nodes. Amyloid appears as pale eosinophilic amorphous material in H&E‐stained sections (Figure 7.4). Amyloid deposits first in the subcapsular sinus and may expand into the paracortex.
Sinus dilatation: Dilatation of the medullary sinuses is a common finding in the mesenteric and mediastinal lymph nodes of old mice. The sinuses contain proteinaceous fluid, lymphocytes, and macrophages.
Sinus erythrocytosis: The presence of erythrocytes in the subcapsular and medullary sinuses may be the result of antemortem hemorrhage in the tributary area or be caused by tissue manipulation and euthanasia especially in the bronchial and mediastinal lymph nodes when the mice are euthanized by CO2 asphyxiation. In chronic hemorrhage, erythrocyte and hemosiderin‐laden macrophages are typically present.
Angiomatous hyperplasia: This is a common finding in the mesenteric lymph nodes of old mice characterized by an increased number of endothelium‐lined spaces in the cortex and medulla. The vessels may contain proteinaceous fluid and erythrocytes. There is no evidence that this lesion will progress to a neoplasm.
Pigmentation: Accumulation of hemosiderin and ceroid‐lipofuscin is common in sinusoidal macrophages that drain areas of hemorrhage and inflammation. Melanin may be observed in skin‐draining lymph nodes of black mice.
Macrophage hyperplasia: Diffuse or focal increases of the different macrophage populations may occur in lymph nodes as a result of influx from the blood or draining area or as a result of proliferation. A diffuse accumulation of subcapsular and medullary sinus macrophages is commonly referred to as sinus histiocytosis. The macrophages may contain pigment, red blood cells, or material injected in the tributary area such as mineral or oil from vaccine adjuvants (Figure 7.4).
Mucosa‐Associated Lymphoid Tissues
The mucosa‐associated lymphoid tissues are composed of the inductive sites for adaptive immune responses in the intestinal and upper respiratory tract. In the mouse, they include the Peyer's patches, solitary lymphoid follicles, and colonic lymphoid patches in the intestine, and the nasopharynx‐associated lymphoid tissue (NALT) and lacrimal duct‐associated lymphoid tissue (LDALT) in the nose (Figure 7.5). Mice do not have tonsils. Mucosa‐associated lymphoid organs do not have afferent lymphatics, and antigens gain access to the lymphoid tissue from the mucosal surface via specialized epithelial cells (microfold or M cells). In addition, dendritic cells may extend dendritic processes between epithelial cells to sample the lumen of the intestinal and respiratory tract. M cells are derived from epithelial stem cells. Mature M cells are characterized electron microscopically by short, irregular microvilli and a thin brush border, and express glycoprotein 2 as a distinct surface marker [52]. The basolateral membrane of M cells is invaginated forming a pocket that often contains lymphocytes, macrophages, and dendritic cells. Antigens taken up by transcytosis through M cells are processed by dendritic cells and activate B and T cells in the underlying lymphoid tissue.
Examination of mucosa‐associated lymphoid tissues: Peyer's patches can be observed and counted macroscopically in adult mice. However, for a detailed investigation of lymphoid tissues in the small intestine, the intestine should be cut in two pieces, and each piece rolled up in a cassette as a “Swiss roll”. Serial longitudinal sections allow for a thorough examination of the intestine. The large intestine is first opened longitudinally to empty the contents and then rolled up in a cassette. To examine the development of Peyer's patches in late‐stage embryos and newborn pups, whole mount immunohistochemistry can be performed [53]. Microscopic examination of the NALT and LDALT requires cross sections of the nose after fixation and decalcification of the head. A cross section just rostral of the first molar will include the NALT.Figure 7.3 Lymph nodes. C57BL/6J (a–c), B6.Cg‐Foxn1nu (d–f), B6.129S7‐Rag1tm1Mom (g–i), and NOD.Cg‐Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice (j–l). a, d, g, j – cervical lymph node; b, e, h, k – jejunal lymph node with higher magnification of subcapsular sinus and cortex (c, f, i, l). Arrows point to lymph nodes in NSG mice.Figure 7.4 Amyloid. (a) Amyloid in mesenteric lymph node of 615‐day old C57BL/6J mouse. (b) Macrophages with intracellular accumulation of aluminum adjuvant in medullary cords. Iliac lymph node of male CD‐1 mouse after injection of an aluminum adjuvant.Figure 7.5 Mucosal lymphoid tissues. (a) Peyer's patch in C57BL/6 J mouse. (b) Colonic lymphoid patch in C57BL/6J mouse. (c) Cross section of the nose of a CD‐1 mouse. Arrows point to the nasopharynx‐associated lymphoid tissue (NALT) and * identifies the lacrimal duct with the lacrimal duct‐associated lymphoid tissue (LDALT). Higher magnification of the squamous epithelium overlying the LDALT (d) and ciliated epithelium overlying the NALT (e).
Peyer's patches: Peyer's patches are located in the anti‐mesenteric wall of the small intestine. The number of Peyer's patches varies between 6 and 10 depending on the mouse strain. Each Peyer's patch is composed of several lymphoid follicles [8–12] which extend from the submucosa into the lamina propria. The follicles have a prominent germinal center as a result of immune stimulation by antigens from the intestinal lumen, and are separated by small interfollicular areas which are analogous to the paracortex of lymph nodes and contain high endothelial venules. The formation of Peyer's patches begins before birth and continues in the first weeks after the pups are born. Several mutations are associated with a lack or reduced number and size of Peyer's patches (Table 7.2). In SHARPIN‐deficient mice (Sharpincpdm and Sharpincpdm‐Dem), Peyer's patches are initially formed and populated by B and T cells. However, lymphoid follicles are not formed, and the lymphoid tissues undergo regression at about two to three weeks of age resulting in the absence of Peyer's patches in adult mice [54]. The majority of lymphocytes in Peyer's patches are B cells. Foxn1 mutant mice that lack T cells have well‐developed Peyer's patches with sparsely populated interfollicular areas. Peyer's patches are absent from mice with null mutations in Rag1, Rag2, Prkdc and Il2rg genes, although anlagen of VCAM1‐positive stromal cells could be detected in neonatal C.B17/Icr‐SCID Jc mice [53]
Solitary intestinal lymphoid tissues (SILTs): These are individual follicles located in the lamina propria of the small and large intestine. They are formed after birth, but are considered secondary lymphoid tissues with a similar function to Peyer's patches. There are about 100–200 SILTs in the mouse intestine, the number depending on the mouse strain. Cryptopatches are small collections of leukocytes, about 80 μm in diameter, in the intestinal lamina propria that do not extend to the epithelium. Their cellular composition includes dendritic cells and innate lymphoid cells. Cryptopatches may represent immature forms of SILTs.
Large intestinal lymphoid tissue: There are two to five colonic patches in the colon of C57BL/6J mice. Each is composed of two large lymphoid follicles in the submucosa extending into the lamina propria of the colon separated by an interfollicular T cell area. M cells are present in the overlying mucosal epithelium. In addition, there are SILTs scattered throughout the lamina propria of the colon with increasing density toward the distal colon. Lymphoid patches similar to colonic patches are also present in the cecum and rectum.
Nasopharynx‐associated lymphoid tissue: The NALT consists of two bilateral rows of five to six lymphoid follicles in the ventral meatus of the nasal cavity. The overlying epithelium consists of ciliated cells interspersed with M cells. The NALT is absent or hypoplastic in several mutant mouse strains (Table
