Nasal associated lymphoid tissue of the Syrian golden hamster expresses high levels of PrPC

Abstract

The key event in the pathogenesis of the transmissible spongiform encephalopathies is a template-dependent misfolding event where an infectious isoform of the prion protein (PrPSc) comes into contact with native prion protein (PrPC) and changes its conformation to PrPSc. In many extraneurally inoculated models of prion disease this PrPC misfolding event occurs in lymphoid tissues prior to neuroinvasion. The primary objective of this study was to compare levels of total PrPC in hamster lymphoid tissues involved in the early pathogenesis of prion disease. Lymphoid tissues were collected from golden Syrian hamsters and Western blot analysis was performed to quantify PrPC levels. PrPC immunohistochemistry (IHC) of paraffin embedded tissue sections was performed to identify PrPC distribution in tissues of the lymphoreticular system. Nasal associated lymphoid tissue contained the highest amount of total PrPC followed by Peyer's patches, mesenteric and submandibular lymph nodes, and spleen. The relative levels of PrPC expression in IHC processed tissue correlated strongly with the Western blot data, with high levels of PrPC corresponding with a higher percentage of PrPC positive B cell follicles. High levels of PrPC in lymphoid tissues closely associated with the nasal cavity could contribute to the relative increased efficiency of the nasal route of entry of prions, compared to other routes of infection.

Fig 1. NALT contains significantly more PrP^C than other lymphoid tissues.

A) Western blot analysis and B) normalized quantification of lymphatic tissue PrP^C abundance. SP—spleen, SLN—submandibular lymph node, MLN—mesenteric lymph node, PP—Peyer’s patch, NALT—nasal associated lymphoid tissue.

Fig 2. PrP^C migration patterns were distinct for different lymphoid tissues.

A) Migration banding patterns are consistent when comparing individual animal homogenates. SP, SLN and PP homogenates from individual animals migrate similarly. B) Line graph of lane intensity analysis demonstrating peak differences in intensity of PrP^C migration. C) Western blot of lymphoid tissue (NALT, SP, and PP) and BR control. D) Line graph of lane analysis of PrP^C intensity of lymphoid tissue indicates intensity difference and migration profile of NALT. E) Comparison of PNGase treated and untreated PrP^C from lymphoid tissue (SLN, PP and NALT and BR) demonstrates variable levels of full length and truncated PrP^C in samples. BR—brain, SP—spleen, PP—Peyer’s patch, SLN—submandibular lymph node, NALT—nasal associated lymphoid tissue.

Fig 3. Immunohistochemistry (IHC) of lymphoid tissues demonstrating presence and localization of PrP^C within B cell follicles.

IHC was performed on A/B) NALT, C) MLN, D) SLN, E) PP and F) SP with the anti-PrP antibody 3F4 (B-F) or an isotype control (A). The tissue sections were processed identically using the same reagents at the same time to illustrate relative differences in PrP^C expression between tissues. The scale bar represents 100 μm. NALT—nasal associated lymphoid tissue, MLN—mesenteric lymph node, SLN—submandibular lymph node, PP—Peyer’s patch, SP—spleen.