Mouse-adapted scrapie strains 139A and ME7 overcome species barrier to induce experimental scrapie in hamsters and changed their pathogenic features

Abstract

Background: Transmissible spongiform encephalopathy (TSE) diseases are known to be zoonotic diseases that can infect different kinds of animals. The transmissibility of TSE, like that of other infectious diseases, shows marked species barrier, either being unable to infect heterologous species or difficult to form transmission experimentally. The similarity of the amino acid sequences of PrP among species is believed to be one of the elements in controlling the transmission TSE interspecies. Other factors, such as prion strains and host's microenvironment, may also participate in the process.

Methods: Two mouse-adapted strains 139A and ME7 were cerebrally inoculated to Golden hamsters. Presences of scrapie associate fibril (SAF) and PrPSc in brains of the infected animals were tested by TEM assays and Western blots dynamically during the incubation periods. The pathogenic features of the novel prions in hamsters, including electrophoretic patterns, glycosylating profiles, immunoreactivities, proteinase K-resistances and conformational stabilities were comparatively evaluated. TSE-related neuropathological changes were assayed by histological examinations.

Results: After long incubation times, mouse-adapted agents 139A and ME7 induced experimental scrapie in hamsters, respectively, showing obvious spongiform degeneration and PrPSc deposits in brains, especially in cortex regions. SAF and PrPSc in brains were observed much earlier than the onset of clinical symptoms. The molecular characteristics of the newly-formed PrPSc in hamsters, 139A-ha and ME7-ha, were obviously distinct from the original mouse agents, however, greatly similar as that of a hamster-adapted scrapie strain 263 K. Although the incubation times and main disease signs of the hamsters of 139A-ha and ME7-ha were different, the pathogenic characteristics and neuropathological changes were highly similar.

Conclusions: This finding concludes that mouse-adapted agents 139A and ME7 change their pathogenic characteristics during the transmission to hamsters. The novel prions in hamsters' brains obtain new molecular properties with hamster-specificity.

Figure 1

Comparative incubation periods of the experimental scrapie in hamsters cerebrally infected with the hamster-adapted strain 263 K and mouse-adapted strains 139A and ME7. The abscissa shows the survival days and the y-axis shows the percentages of the survival.

Figure 2

Morphological assays of SAFs in the hamsters' brains of three different scrapie strains in TEM. (A) agent 263 K. (B) agent 139A-ha. (C) agent ME7-ha. (D) SAF in the brain homogenates infected with agent 263 K stained by immune colloidal gold technique with PrP specific mAb 3 F4.

Figure 3

Neuropathological assays of the regions of cortex and cerebellum from the hamsters infected with agent 263 K, 139A-ha and ME7-ha. (A) Top and middle panels are HE stains of cortex and cerebellum. Bottom panels are GFAP IHC stains of cortex. The arrows show typical vacuoles. (B) Scores the severity and distribution of vacuolation. The average value for each brain region is calculated from three independent counts and indicated as mean ± SD.

Figure 4

Analyses for PrP^Sc in brain homogenates with Western blots. (A) PrP^Sc in hamsters' brains of agent 263 K, 139A and ME7 detected with mAb 3 F4. (B) PrP^Sc in mice brains of agent 139A and ME7 detected with mAb 1E4. (C) Deglycosylated PrP^Sc in hamsters' brains of agent 263 K, 139A and ME7 detected with mAb 3 F4. (D) Immunoreactivities of PrP^Sc in hamsters' brains of agent 263 K, 139A and ME7 with a panel of PrP specific mAbs, including 1E4, 6D11, 3 F4, 6H4 and 8H4. Molecular weight standards are shown on the left and different mAbs are shown on the right. "+": with PK or PNGase F; "-": without PK The products of PK-digested PrP^Sc from the hamsters' brains infected by mouse-adapted agents 139A and ME7 were further exposed to glycosidase. One single PrP-specific bands were observed, which were at the exactly same position in SDS-PAGE as that in the preparation of 263 K PrP^Sc (Figure 4C). It implies that the newly-formed PrP^Sc in the hamsters' brains by infections of mouse-adapted scrapie agents have the same glycosylated profiles as the hamster-adapted scrapie strain 263 K.

Figure 5

PK resistance of PrP^Sc in hamsters' brains of agent 263 K, 139A-ha and ME7-ha. (A) Western blots with mAb 3 F4. The concentrations of PK in individual preparation are indicated on the top. Molecular weight standards are shown on the left. (B) Quantitative analyses of each gray numerical value of PrP blots. Relative gray values of the PrP signals in each experiment are normalized by division with that of the respective reaction without PK. The average values are calculated from three independent blots and presented with as mean ± SD.

Figure 6

Conformational stability of PrP^Sc in hamsters' brains of agent 263 K, 139A-ha and ME7-ha. (A) Western blots of total PrP with mAb 3 F4. Hamsters' brain homogenates are incubated with 1 to 6 M GdnHCl. The concentrations of GdnHCl in individual preparation are indicated on the top. Molecular weight standards are shown on the left. (B) Quantitative analyses of each gray numerical value of total PrP blots. The average values are calculated from three independent blots and presented with as mean ± SD. (C) Western blots of PrP^Sc with mAb 3F4. Hamsters' brain homogenates are incubated with 1 to 6 M GdnHCl and subsequently exposed to 50 μg/ml PK. The concentrations of GdnHCl in individual preparation are indicated on the top. Molecular weight standards are shown on the left. (D) Quantitative analyses of each gray numerical value of PrP^Sc blots. The average values are calculated from three independent blots and presented with as mean ± SD.

Figure 7

Morphological assays of SAFs in the hamsters' brains of three different scrapie strains after incubated with 3 M GdnHCl. (A) agent 263 K; (B) agent 139A-ha; (C) agent ME7-ha.