J Vet Sci.  2013 Mar;14(1):1-6. 10.4142/jvs.2013.14.1.1.

C6, a new monoclonal antibody, reacts with the follicle-associated epithelium of calf ileal Peyer's patches

• Tozaki K
• Kimura J
• Yasuda M
• Ryu N
• Nasu T
• Pernthaner A
• Hein WR

• KMID: 1482806
• DOI: http://doi.org/10.4142/jvs.2013.14.1.1

Abstract

The follicle-associated epithelium (FAE) of Peyer's patches (PPs) contains M cells that are important for reducing mucosal immune responses by transporting antigens into the underlying lymphoid tissue. We generated a monoclonal antibody (C6) that reacted with the FAE of calf ileal PPs, and analyzed the characteristics of C6 using immunohistochemistry and Western blotting. FAE of the ileal PP was stained with C6 during both late fetal developmental and postnatal stages. Neither the villous epithelial cell nor intestinal crypt basal cells were stained at any developmental stage. During the prenatal stages, FAE of the jejunal PP was C6-negative. However, a few C6-positive cells were distributed diffusely in some FAE of the jejunal PPs during the postnatal stages. The protein molecular weight of the antigen recognized by C6 was approximately 45 kDa. These data show that C6 is useful for identifying the FAE in ileal PPs and further suggest that differentiation of the FAE in these areas is independent of external antigens.

Keyword

bovine; follicle-associated epithelium; monoclonal antibody; Peyer's patch

MeSH Terms

Animals
Antibodies, Monoclonal/*immunology
*Cattle
Fetus
Hybridomas
Ileum/*ultrastructure
Intestinal Mucosa/*immunology
Peyer's Patches/*immunology/ultrastructure
Antibodies, Monoclonal

Figure

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  Fig. 1 C6+ cell distribution in the FAE of ileal and jejunal Peyer's patches (PPs) during the prenatal stages. (A) No ileal PP lymphoid tissue formation was observed in a 5-month old fetus. All cells were C6-negative. (B) In a 7-month old fetus, ileal PP formation was observed. A subset of cells in the FAE but not the villus epithelium was C6 positive. (C) In a 9-month old fetus, C6+ cells were found in FAE of the ileal PP but the villous epithelial cells were C6-negative. (D) No C6-positive cells were found in FAE of the jejunal PP or villus epithelium of the 7~9-month old fetuses. Do: dome region and Vi: villus. Scale bars = 50 µm.

• 

  Fig. 2 C6+ cell distribution in the FAE of ileal and jejunal PPs during the postnatal stages. (A and C) FAE in the ileal PP was stained by C6 during the postnatal stages. In the ileal PP, the epithelial cells converted from negative to C6-positive along the crypt-to-dome axis but all epithelial cells were negative along the crypt-to-villus axis. The asterisk indicates C6+ FAE in the ileal PP dome region. The broken lines represent the outline of villous epithelial cells which were C6-negative. (B and D) Very few C6+ cells were found in some FAE of the jejunal PP (arrowhead). The broken lines represent the outline of villous epithelial cells. Arrows indicate the smooth muscular fibers in the lamina propria. Do: dome region, Vi: villus, and Cr: crypt. Scale bars = 50 µm (A and B) and 10 µm (C and D).

• 

  Fig. 3 The distribution of C6+ cells in other lymphoid tissues. (A) Hassall's corpuscle (arrow) in the thymus medulla was C6-positive. Epithelial layers of the palatine tonsil (B; stratified squamous epithelium) and pharyngeal tonsil (C; pseudostratified epithelium) were also C6-positive. Reticular cells in lymphatic follicles of the mesenteric lymph node (D), ileal PP lymphatic follicles (E), and the spleen (F) were C6-positive. Me: thymus medulla, Co: thymus cortex, Ep: epithelium, and Fo: lymphatic follicle. Scale bars = 50 µm.

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  Fig. 4 Scanning electron micrograph (SEM) of the FAE in ileal and jejunal PPs. (A) The dome region (arrowhead) of the jejunal PP was surrounded by villi. (B) M cells with irregular microvilli were sparsely scattered in FAE of the jejunal PP. (C) Many dome regions of the ileal PP were observed. (D) The majority of cells in the FAE were M cells with irregular and sparse microvilli. Scale bars = 50 µm (A), 5 µm (B), 70 µm (C), and 3 µm (D).

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  Fig. 5 Western blotting. The C6 monoclonal antibody reacted with a bovine ileal PP protein as well as an ovine protein approximately 45 kDa in size present in migrating afferent intestinal lymph cells. Lane 1: bovine, lane 2: molecular weight maker, and lane 3: ovine.

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