An investigation of scrapie pathogenesis in neonatal mice with special reference to germinal centre maturation

Studies carried out during the 1970s-1980s, demonstrated that neonatal mice are less susceptible than adult mice to intraperitoneal (i.p) but not intracerebral (i.c) challenge with the ME7 strain of scrapie. Using doses that are 100% lethal to adult mice, some peripherally challenged neonatal mice resist scrapie infection and survive or have significantly longer incubation periods than mice challenged as adults. Susceptibility to scrapie increases between 7-10 days after birth and no survivors are observed.These experiments were repeated in order to determine whether neonatal mice could sustain scrapie replication within the spleen. The presence of scrapie associated protease resistant prion protein (PrPsc) was used as a marker for infectivity. PrPsc could not be detected by Western blotting, 70 days after infection in the spleens of neonatally challenged mice, a time when infectivity has plateaued within spleens of mice inoculated as adults. This suggests that neonatal mice have an impaired ability to replicate scrapie and therefore resist scrapie infection.In adult mice, cellular prion protein (PrPc) expressing follicular dendritic cells (FDCs) in the spleen are required for replication of scrapie after i.p challenge. It is therefore hypothesised that neonatal mice resist scrapie infection due to lack of mature PrPc expressing FDCs within spleen. The presence of PrPc and the maturation of FDCs were studied in the spleens of developing mice using immunocytochemistry and confocal microscopy. It was found that FDC-M1+ cells can be detected from birth within the immature spleen white pulp. However detectable levels of the FDC functional marker, complement receptor 1 were not observed until 14 days after birth, inferring that the FDCs are functionally immature prior to this timepoint. PrPc was detected from 10 days old immunocytochemically, and was found to co-localise with FDC-M1+ cells with an immature dendritic morphology compared to 30 day old mice. It can be concluded that PrPc is expressed on immature FDCs in the 10 day old mouse. The detection of PrPc on FDC-M1+ cells between 10 and 14 days coincides with a dramatic increase in susceptibility to scrapie. This is consistent with XV the hypothesis that neonatal mice have an impaired ability to replicate scrapie due to insufficient PrPc expressing FDCs in the spleen.Paradoxically, previous experiments have shown a proportion of peripherally inoculated neonatal mice to develop scrapie after very short incubation periods ranging between 150-230 days. During the first two weeks of life, the number of cases with short incubation periods decreases. It is hypothesised that due to the immaturity of peripheral nerves at birth, infectivity can be taken up more efficiently by neonatal peripheral nerve endings and reach the central nervous system (CNS) without prior replication in the spleen. Adult severe combined immunodeficient mice (SCID) are resistant to scrapie due to a lack of mature FDCs. Neonatal SCID mice offer a model with which to test this hypothesis without the complicating factor of a developing lymphoreticular system. After subcutaneous challenge, a large proportion of neonatally inoculated SCID mice developed scrapie after a very short incubation period compared to SCIDs challenged as adults. This suggests that infectivity can access the CNS with greater ease in neonates compared to adults and since SCID mice do not possess functional FDCs, demonstrates that the mechanism is not dependent on prior FDC replication.In conclusion, the results presented in this thesis demonstrate that FDC dependent and non-FDC dependent mechanisms exist in the neonatal mouse for peripheral scrapie pathogenesis