J Vet Sci.  2014 Dec;15(4):465-474. 10.4142/jvs.2014.15.4.465.

Developmental changes in cell proliferation and apoptosis in the normal duck bursa of Fabricius

• Fang J
• Peng X

• KMID: 2070232
• DOI: http://doi.org/10.4142/jvs.2014.15.4.465

Abstract

The aim of this work was to investigate developmental changes in cell proliferation and apoptosis in normal duck bursa of Fabricius using flow cytometry and immunohistochemistry. Studies were carried out on Tianfu ducks on days 24 and 27 of embryogenesis (E24 and E27) along with days 20, 70, and 200 of postnatal development (P20, P70, and P200). Results showed that the percentage of G0/G1 bursa cells significantly increased between E24 and P200 while the percentage of cells in the S phase or G2 + M phase as well as the proliferating index obviously decreased during the same period. Proliferation cell nuclear antigen was detected in lymphocyte and interfollicular epithelium. The proliferative lymphocyte density tended to decrease from E24 to P200. Apoptotic bodies in macrophages, free apoptotic bodies, or nuclei with condensed chromatin in lymphocytes in follicles were identified by transferase-mediated dUTP nick-end labeling. Both flow cytometry and microscopic analysis reveal that the proportion of apoptotic cells and apoptotic lymphocyte density increased from E24 to P20, fell on P70, then rose again on P200. Our foundings demonstrate that cell proliferation decreases and apoptosis increases with age. These changes may account for duck bursa development and involution.

Keyword

apoptosis; bursa of Fabricius; development; duck; proliferation

MeSH Terms

Animals
*Apoptosis
Bursa of Fabricius/*cytology/embryology/growth & development/*physiology
Cell Proliferation
Ducks/embryology/*physiology
Embryo, Nonmammalian/cytology/embryology
Embryonic Development
Epithelium/physiology
Female
Flow Cytometry/veterinary
Immunohistochemistry/veterinary
Lymphocytes/physiology
Male

Figure

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  Fig. 1 Haematoxylin and eosin staining of (A) the bursa of Fabricius on E24. The star (⋆) indicates a lymphoid follicle with indistinct medulla and cortex. (B) The bursa of Fabricius on E27. (C) The bursa of Fabricius on P20. (D) The bursa of Fabricius on P70. (E and F) The bursa of Fabricius on P200. Asterisks (*) indicate large mucoid cysts in the lymphoid follicles. FAE: follicle-associated epithelium, IFE: interfollicular epithelium. C, lymphoid follicle cortex; M, lymphoid follicle medulla. Scale bars = 1,000 µm (A~F).

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  Fig. 2 DNA histogram of the bursa cell cycle obtained by flow cytometry. Panels A-E correspond to E24, E27, P20, P70, and P200, respectively.

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  Fig. 3 Scattergram of apoptotic bursa cells obtained by flow cytometry. Panels A-E correspond to E24, E27, P20, P70, and P200, respectively.

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  Fig. 4 Proliferation cell nuclear antigen (PCNA) assay results with hematoxylin counterstaining. (A) PCNA-positive reaction in the follicular cortex and medulla as well as the IFE on E24 (arrow); the FAE was PCNA-negative. (B) PCNA-positive reaction in the follicular cortex and medulla along with IFE on P20 (arrow); FAE was PCNA-negative. (C) PCNA-positive reaction in the follicular cortex and medulla on E24 (arrow). (D) PCNA-positive reaction in the follicular cortex and medulla on E27 (arrow). (E) PCNA-positive reaction in the follicular medulla on P20 (arrow). (F) PCNA-positive reaction in the follicular cortex and medulla on P70 (arrow). (G) PCNA-positive reaction in the follicular cortex and medulla on P200 (arrow). (H) Negative control for the follicle lymphocytes on E27. Scale bars = 20 µm (A and B), 10 µm (C~H).

• 

  Fig. 5 Transferase-mediated dUTP nick-end labeling (TUNEL) assay findings with haematoxylin counterstaining. (A) TUNEL-positive follicular lymphocytes containing free apoptotic bodies or nuclei with condensed chromatin (arrow with a longer tail), and apoptotic bodies in the cytoplasm of macrophages (arrow with a shorter tail) in the lymphoid follicle on P200. (B) TUNEL-positive follicular cortex and medulla as well as FAE on E24 (arrow). (C) TUNEL-positive follicular cortex and medulla on E27 (arrow); the IFE and FAE were negative. (D) TUNEL-positive follicular cortex and medulla, and FAE on P20 (arrow). (E) TUNEL-positive follicular cortex and medulla on P70 (arrow). (F) TUNEL-positive follicular cortex and medulla on P200 (arrow). (G) TUNEL-positive mucosa epithelium on P200 (arrow). (H) Negative control follicle lymphocytes and mucosa epithelium on P20. Scale bars = 10 µm (A), 20 µm (B~G), 40 µm (H).

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