Bovine PrP expression levels in transgenic mice influence transmission characteristics of atypical bovine spongiform encephalopathy

Abstract

Until recently, transmissible spongiform encephalopathy (TSE) disease in cattle was thought to be caused by a single agent strain, bovine spongiform encephalopathy (BSE) (classical BSE or BSE-C). However, due to the initiation of a large-scale surveillance programme throughout Europe, two atypical BSE strains, bovine amyloidotic spongiform encephalopathy (BASE, also named BSE-L) and BSE-H have since been discovered. These atypical BSE isolates have been previously transmitted to a range of transgenic mouse models overexpressing PrP from different species at different levels, on a variety of genetic backgrounds. To control for genetic background and expression level in the analysis of these isolates, we performed here a comprehensive comparison of the neuropathological and molecular properties of all three BSE agents (BASE, BSE-C and BSE-H) upon transmission into the same gene-targeted transgenic mouse line expressing the bovine prion protein (Bov6) and a wild-type control of the same genetic background. Significantly, upon challenge with these BSE agents, we found that BASE did not produce shorter survival times in these mice compared with BSE-C, contrary to previous studies using overexpressing bovine transgenic mice. Amyloid plaques were only present in mice challenged with atypical BSE and neuropathological features, including intensity of PrP deposition in the brain and severity of vacuolar degeneration were less pronounced in BASE compared with BSE-C-challenged mice.

Fig. 1.

Kaplan–Meier survival curve showing percentage survival of Bov6 mice following challenge with BSE-C, BASE and BSE-H. All cases used were both positive for PrP deposition in the brain and showed vacuolar pathology (a). Pattern of vacuolation observed in brains of Bov6 mice inoculated with BASE, BSE-C and BSE-H. A profile was produced from nine grey matter areas (1, medulla; 2, cerebellum; 3, superior colliculus; 4, hypothalamus; 5, thalamus; 6, hippocampus; 7, septum; 8, retrospinal cortex; and 9, cingulate and motor cortex) and three white matter areas (1*, cerebellar white matter; 2*, midbrain white matter; and 3*, cerebral peduncle). Mean scores were taken from a minimum of 15 mice per group and plotted against brain area±sem (b).

Fig. 2.

Comparative analysis of variation in intensity of PrP^TSE and presence of amyloid plaques in the hippocampus and thalamus regions of brains from Bov6 mice challenged with BSE-C, BASE and BSE-H. BSE-C: 378 days p.i. (a), 604 days p.i. (d); BASE: 406 days p.i. (b), 645 days p.i. (e); BSE-H: 476 days p.i. (c), 637 days p.i. (f). Images obtained after staining with anti-PrP antibody 6H4 and counterstained with haematoxylin. Magnification, ×4. Thioflavin-S fluorescent amyloid plaques (arrows) are clearly visible in the hippocampal region of Bov6 mice challenged with BASE (h) and BSE-H (i), but were not observed with BSE-C (g). Magnification, ×4.

Fig. 3.

Comparative analysis of the CA1 region of the hippocampus of Bov6 mice challenged with BSE-C (604 days p.i.), BASE (645 days p.i.) and BSE-H (637 days p.i.). PrP deposition is visible by anti-6H4 antibody (a–c). Astro- and microgliosis is present in all mice, detected by anti-GFAP (e–g) and anti-Iba1 (i–k), respectively. Uninfected aged matched Bov6 mice showing mild gliosis were used as controls (d), (h) and (l). Magnification, ×10.

Fig. 4.

PK-resistant fragment (PrP^TSE) of the prion protein in Bov6 mice challenged with BSE-H, 581 days p.i. (lanes 1 and 2), BASE, 610 days p.i. (lanes 3 and 4) and BSE-C, 604 days p.i. (lanes 5 and 6). Lanes 1, 3 and 5 represent untreated brain homogenate and lanes 2, 4 and 6 show PK-treated brain homogenate. Anti-PrP mAb 6H4 was used to detect bands.