Scrapie Strain 263K Research Articles

Pharmacological studies of a new derivative of amphotericin B, MS-8209, in mouse and hamster scrapie.

Transmissible subacute spongiform encephalopathies (TSSE) are neurodegenerative diseases characterized by the presence of a modified, partially proteinase-resistant host protein, PrPSc, which accumulates in the brains of infected individuals. Recently it has been reported that amphotericin B (AmB) treatment of hamsters infected with scrapie strain 263K prolongs the incubation period of the disease, and dissociates in vivo replication of the scrapie agent from PrPSc accumulation. We report here on data obtained after treatment with AmB and one of its derivatives, MS-8209, in experimental scrapie of mouse and hamster. Treatment was carried out by the intraperitoneal route 6 days per week, at three different dosages initiated at the time of infection. Two regimens were used: during the early time of infection or throughout the experimental infection. Results indicate that MS-8209 was as efficient as AmB in prolonging the incubation time and decreasing PrPSc accumulation in the hamster scrapie model. A dose-dependent response was observed in mice treated early after experimental infection. At a dose of 2.5 mg/kg, MS-8209 significantly prolonged the incubation period (by 11.9%). In long-term treatment of mice, MS-8209 and AmB markedly reduced PrPSc levels in the preclinical stage of the disease. These data demonstrate that the effect of AmB is not restricted to one model (hamster-263K). This regimen leads to an inversion of the PrPSc to proteinase-sensitive protein (PrPSens) ratio, suggesting PrPSens (presumably cellular PrPC) accumulation occurs before its conversion into PrPSc. As it has been shown that AmB does not modify the infectivity titre, we conclude that the drugs could act by inhibiting either the interaction of the scrapie agent with PrPSens during the early times of infection or the conversion of PrPSens into PrPSc.

Histopathological changes in the islets of Langerhans in scrapie 139H-affected hamsters

Previous studies showed that the 139H strain of scrapie injected into hamsters caused obesity, a marked hypertrophy of the islets of Langerhans, generalized endocrinopathy and marked hypoglycaemia-hyperinsulinaemia. In the current study, female weanling Syrian hamsters (LVG/LAK strain) were inoculated intracerebrally with scrapie strain 139H or 263K, or with normal hamster brain. Sections of the pancreas stained with haematoxylin and eosin or Gomori's one-step trichrome were examined by light microscopy. The 139H-affected hamsters showed extensive vacuolization, cellular hypertrophy, cellular atrophy, cytoplasmic vesicles and nuclear pathological changes in the islets of Langerhans. Also observed were abnormal structures, termed blood vessel cores, in the islets of 139H-affected hamsters. These structures were almost always centrally located within islets and were surrounded by B cells, some of which were abnormally elongated. None of these pathological changes were seen in the islets of Langerhans in control or 263K-affected hamsters. The level of scrapie-specific protease resistant protein (PrPSc) in pancreas was much lower than that in brain, a finding consistent with previous data showing low scrapie infectivity titres in pancreas.

Subcellular distribution and physicochemical properties of scrapie‐associated precursor protein and relationship with scrapie agent

We studied the biologic properties of hamster-adapted scrapie (strain 263K) and its relationship to the precursor protein of scrapie (PrP33-35Sc). The highest titer of infectious material and the greatest concentration of PrP33-35Sc were in the fractions containing microsomal and synaptosomal membranes. We found traces of infectivity in the absence of PrP33-35Sc associated with matrix protein. Partitioning of membranes with neutral chloroform-methanol resulted in concentration of PrP33-35Sc and infectivity within the interphase layer. Recombination of membrane glycoproteins (interphase) with lipids extracted from homologous brains decreased infectivity greater than or equal to 4 logs. Temperature-dependent phase separation of infected synaptosomal and microsomal membranes with Triton X-114 yielded a phospholipid-rich phase containing a high concentration of PrP33-35Sc and greatest infectivity titers. This material spontaneously formed liposomes, indicating that PrP33-35Sc and PrP33-35C precursor proteins are highly hydrophobic intrinsic membrane components integrated with phospholipids. Homologous membrane phospholipids appear to prevent aggregation of the scrapie isoform of PrP and maintain high levels of infectivity.

Pathogenesis of scrapie (strain 263K) in hamsters infected intracerebrally, intraperitoneally or intraocularly.

After intracerebral (i.c.) infection of hamsters, the 263K strain of scrapie replicated at a nearly constant exponential rate until clinical disease developed when titres in brain averaged 9.8 log10 LD50 i.c. units/g. After intraperitoneal infection, scrapie replication was first detected in spleen, then in thoracic spinal cord and finally in lumbar cord and brain. This pattern suggests that invasion of the central nervous system occurs by spread of infection along certain visceral autonomic nerves. Infectivity was detected in the thoracic cord only 3 to 4 weeks after infection (incubation period 16 weeks) indicating the exceptional neuroinvasiveness of this scrapie model. This observation and the failure of splenectomy to lengthen incubation period raises the possibility of direct infection of nerve tissue in the peritoneum and transport to the thoracic cord with minimal prior replication of scrapie agent extraneurally. After intraocular infection of the right eye, replication (or accumulation) of scrapie was detected in the right optic nerve and left superior colliculus, then in the right superior colliculus and finally in the left optic nerve and medulla. This pattern shows that scrapie infection can spread along nerves, possibly by intra-axonal transport. The duration of agent replication in brain (between detectable onset of replication and clinical disease) was shortest after intraperitoneal infection (51 to 58 days), longer after intracerebral infection (81 to 88 days) and longest after intraocular infection (greater than 121 days). These differences may reflect the relative efficiency of the neural pathways by which infectivity spreads from different sites of entry in the brain to the postulated 'clinical target areas'.

Virus like sensitivity of the scrapie agent to heat inactivation.

The resistance of the infectious agent of scrapie disease to sterilization at 100 degrees or 121 degrees C is reputed to be inconsistent with the structure of conventional viruses. However, in kinetic studies the majority of hamster scrapie strain 263K infectivity was (like that of previously characterized viruses) rapidly inactivated at temperatures of 100 degrees C or greater. Small resistant subpopulations remained. Similar heat-resistant subpopulations were observed at 60 degrees C for phage lambda but only in the presence of brain homogenate. Brain homogenate may also confer stability to small subfractions of scrapie infectivity. Such refractory subpopulations cannot be used to make structural inferences that are properly obtained from the behavior of the majority population as revealed in the initial inactivation.

Scrapie infectivity, fibrils and low molecular weight protein.

The development of a short incubation model of scrapie (strain 263K), in golden hamsters has added impetus to the purification of the infectious agent. Our own attempts have been based on methods pioneered by Millson and developed by Prusiner. We present here results indicating that a purification factor of up to 10(4) with respect to protein may now be possible. Fractions from brain with high infectivity had a sedimentation range of 70-300S and contained an abundance of fibrils closely similar to the scrapie-associated fibrils (SAF) discovered by Merz et al.. Material of molecular weight (Mr) 26,000, which is probably protein, appears to be a major constituent of the fibrils. The association between infectivity and fibrils raises two possibilities: the fibrils are an infectious form of the scrapie agent or they are a pathological response to scrapie infection.