Overall, these studies suggest that prion neuroinvasion following i.t. inoculation of LT�� and muMT null mice does not depend on scrapie agent replication kinase inhibitor Regorafenib in the LRS. To investigate the route(s) of neuroinvasion following i.t. inoculation of mice with the RML scrapie agent, the distribution of PrPSc was measured in the tongue by Western blot and immunohistochemistry. PrPSc was enriched from tongues (100 mg, which represents 70% to 100% of the tongue) of mock- and RML scrapie-infected mice after i.t. inoculation and analyzed by Western blot. A weak PrPSc signal of three polypeptides between 18 kDa and 28 kDa was sometimes observed in tongues of C57BL/6J, LT�� null, and muMT null mice, but there was also a weak signal observed in tongues of mock-infected LT�� null mice (Fig. (Fig.3A).3A).

The molecular mass of the polypeptide fragments in this mock-infected animal was between 25 and 32 kDa. Based on analysis of additional tongues from these mice, we were unable to conclude that the immunoreactivity in the scrapie-infected mice was PrPSc specific despite the apparent correct molecular mass of these polypeptides. Using immunohistochemistry, tongues from symptomatic mice following i.t. inoculation did not reveal PrPSc deposition in nerve fibers, ganglion, skeletal muscle, muscle spindles, epithelium, or mucus and serous glands of C57BL/6 or LT�� null mice (data not shown). Strong PrP immunostaining was found in the taste buds of the foliate and circumvallate papillae, but this pattern was found in both mock- and scrapie-infected mice, suggesting either nonspecific immunoreactivity or high levels of PrPC that are immunoreactive (data not shown).

These findings are consistent with the Western blot data, and they indicate that PrPSc was not found in the tongues of scrapie-infected mice despite the finding that scrapie neuroinvasion from the tongue was independent of LRS infection (Table (Table44). Pathogenicity of the RML scrapie agent in wild-type, LT�� null, and muMT null mice following Cilengitide i.n. inoculation. To investigate scrapie agent neuroinvasion from the nasal cavity, C57BL/6J, LT�� null, and muMT mice were inoculated in the nasal cavity with the RML scrapie agent at two different doses and mice were monitored for scrapie agent infection. For the i.n. route of inoculation, only one C57BL/6J mouse developed scrapie while none of the LT�� null mice developed disease at the lower dose of scrapie agent (Table (Table1).1). In the one scrapie agent-positive C57BL/6J mouse, PrPSc was found in the brain and spleen at clinical disease (Fig. 1A and B, lane 13), but it was not found in the brains of asymptomatic C57BL/6J (Fig. (Fig.1A,1A, lane 14) or LT�� null (Fig. (Fig.1A,1A, lane 16) mice at 365 days postinoculation.