Ultrastructural findings in pigs experimentally infected with bovine
spongiform encephalopathy agent
Pawel P. Liberski1, Beata Sikorska1, Gerald A.H. Wells2, Steve A.C.
Hawkins3, Michael Dawson3, Marion M. Simmons2 1Department of Molecular Pathology
and Neuropathology, Medical University of Lodz, Poland, 2TSE Department, and
3Specialist Scientific Services Department, Animal Health and Veterinary
Laboratories Agency, Weybridge, Addlestone, Surrey, United Kingdom Folia
Neuropathol 2012; 50 (1): 89-98
A b s t r a c t
We report here an electron microscopic study of selected nervous system
tissues from pigs infected experimentally with the agent of bovine spongiform
encephalopathy (BSE). Generally, the ultrastructural neuropathology of
BSE-affected pig brain resembled that of BSE-affected cattle brain. Spongiform
change, in the form of membrane-bound vacuoles separated by septae into
secondary chambers, dominated the pathology. Numerous astrocytic processes were
visible in close conjunction with elongated microglial cells. Neuronal
degeneration presented as either dystrophic neurites or by the formation of
autophagic vacuoles. Altered subcellular organelles: mitochondria,
electron-dense bodies, autophagic vacuoles, neurofilaments and
“branching-cisterns” accumulated in abnormal neurites. Autophagic vacuoles
appeared as neuronal cytoplasm of increased electron-density sequestrated by
intracytoplasmic membranes. Tubulovesicular structures were numerous,
particularly in the cerebellum. Unusual crystalloids were observed in the white
matter. In conclusion, experimental BSE in pigs demonstrated ultrastructural
pathology in keeping with that observed in other spongiform encephalopathies.
Key words: prions, BSE, pigs, ultrastructure.
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Material and methods
Experiment design and inoculation procedure
The pigs were infected at 1-2 weeks of age by multiple- route parenteral
inoculation with a homogenate of bovine brain from natural BSE cases, as
described in full previously [76,86,87]. All challenges were carried out in
accordance with the Animals (Scientific Procedures) Act, 1986, under licence
from the UK Home Office. Animals were sedated with azoperone (Stresnil; Janssen
Animal Health) and killed by the intravenous injection of pentobarbitone sodium
followed by exsanguination. When clinical disease developed, animals were killed
and samples collected immediately postmortem.
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Electron microscopy
Multiple samples, comprised 2-3 mm3, of cerebral cortex, brain stem at the
level of vestibular nuclei, ventral horns of the spinal cord, cerebellum and
dorsal root ganglia, selected on the basis of the previously determined
prevalence of light-microscopy changes [76,87], were fixed immediately after
dissection in 2.5% glutaraldehyde, freshly prepared in phosphate buffer (pH
7.4), then postfixed in 1% osmium tetroxide and processed for routine electron
microscopy. Comparable areas of brain from uninoculated pigs served as controls.
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Results
In general, the ultrastructural features of BSE-affected pig brain were
similar to those of BSE-affected cattle [46,66,67] and humans with TSEs [60,61].
Spongiform change in the form of membrane-bound vacuoles (Fig. 1) separated by
membranes curled into secondary chambers dominated the pathology. A dense
astrocytic reaction was accompanied by abundant elongated microglial cells. Of
particular note was the finding of numerous astrocytic processes in close
conjunction with microglial cells. Neuronal degeneration presented as either
neuroaxonal dystrophy, as evidenced by dystro - phic neurites, or autophagic
vacuoles. Dystrophic neurites accumulated altered subcellular organelles:
mitochondria (Fig. 1B), electron-dense bodies, neurofilaments and
“branching-cisterns” (Fig. 2). Autopha - gic vacuoles appeared as a part or
parts of the neuronal cytoplasm sequestrated by intracytoplasmic membranes (Fig.
3). Sequestrated cytoplasm was of higher electron density than the remaining
cytosol. Discontinuity of plasma membranes was occasionally seen (Fig. 3B,
arrow). Tubulovesicular structures (TVS) were numerous with the highest number
of affected processes in the cerebellum (Fig. 4). Many large multivesicular
bodies were seen (Fig. 5).
PORCINE SPONGIFORM ENCEPHALOPATHY PSE
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