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6.2 Molecular pathology
BSE resembles other members of the scrapie family in not having any gross pathological lesions consistently associated with disease, norany biochemical or haematological abnormalities (Aldridge et al., 1988; Johnson and Whitaker, 1988; Scott et al., 1988; 1 990a). Characteristic histopathological and molecular changes are found in the central nervous system.
In common with the other diseases in the scrapie family, BSE has a distinctive non-inflammatory pathology with three main features (Wells, Wilesmith and McGill, 1991):
The most important diagnostic lesion is the presence of bilaterally symmetrical neuronal vacuolation, in processes and in some. The former consists of a microcystic vacuolation (spongiform change) of the grey matter neuropil (Fig. 4a). This is the major vacuolar lesion of BSE. The other type of vacuolation consists of large, empty spaces distending neuronal perikarya (Fig. 4b). This type of vacuolation is a conspicuous feature of natural scrapie but it is less prominent in BSE. Neurons with somal vacuolation frequently have an otherwise normal appearance. However, scattered necrotic some are seen and, as with natural scrapie, neuronal loss is an occasional but rarely conspicuous feature.
Hypertrophy of astrocytes often accompanies vacuolation. This has been demonstrated in routinely stained sections and also in sections immunostained for glial fibrillary acidic protein (Fig. 4c).
Cerebral amyloidosis is an inconstant histological feature of the scrapie family of diseases. It is present in BSE but mostly as sparse, focal deposits in a small proportion of cases. Congophilic plaques showing characteristic dichroism in polarized light were found in the thalamus of one out of 20 cases examined systematically for amyloid. The plaques immunostained for PrP (see Molecular pathology).
A number of studies have examined the quantitative distribution of the vacuolar changes in BSE. A study of 22 clinically affected brains (Scott fit al., 1990b) showed that the mean vacuolar densities were greatest in: the medulla oblongata (in the solitary tract nucleus, the spinal tract nucleus of the trigeminal nerve, vestibular nuclei and the reticular formation); the central grey matter in the midbrain; and the paraventricular area in the hypothalamus, thalamus and the septal area. In contrast, the vacuolar change was often minimal in the cerebellum, hippocampus, cerebral cortex and basal nuclei.
Another quantitative study examined a series of 100 cases, sampled before July 1989 (Wells, Wilesmith and McGill 1991). The vacuolar patterns in the brain were remarkably uniform in contrast to the variability described in sheep scrapie.
These findings indicate a uniformity in the pathogenesis of BSE in terms of the route of infection (through the alimentary tract) and the major strain(s) of the infectious agent in cattle.
Electron microscope (EM) observations on thin sections of BSE-affected brain revealed the expected findings of a scrapie-like disease (Liberski, 199()). These included numerous membrane-bound intracellular vacuoles, predominantly in dendrites. Many dendrites and axons contained accumulations of neurofilaments, mitochondria and electron-dense bodies. In addition, tubulovesicular structures were seen. These are similar to the structures found in scrapie except that those seen in BSE-affected brains were membrane-bound. This might be a distinctive lesion of BSE but it requires further investigation.
6.2 Molecular pathology
In addition to the histological lesions that characterize the scrapie family of diseases, extracts of clinically affected brains contain an abundance of characteristic abnormal fibrils (SAF) which are readily identified by negative stain EM. Their presence in extracts of BSE-affected brain was important in confirming the histological observations that BSE is a scrapie-like disease of cattle (Wells et al., 1987).
SAF are easily purified and much is known about them (Hope et al., 1988). They are derived from a normal (i.e. host-coded) membrane glycoprotein, known as PrP, which is present in many tissues, particularly the brain. In the course of scrapie infection, this normal protein undergoes an abnormal post-translational modification (in ways that are not yet understood) and acquires the ability to form fibrils. The modified protein is partially resistant to proteolytic enzymes so that it accumulates in brain, often to about ten times the concentration of the normal protein.
The fibrils from BSE-affected brain have been purified and studied in terms of size, protease resistance, immunoreactivity (with antibodies prepared against SAF), lectin binding and partial N-terrninal amino acid sequence. The results show conclusively that the fibrils from BSE-affected brain are made from bovine PrP (Hope et al., 1988).
There are three ways in which the fibril form of PrP can be detected:
SAF can be identified by their characteristic morphology when examined by EM (Wells et al., 1987; Hope et a/., 1988; Scott et al., 1 990b).
Purified or even crude preparations of SAF can be analysed by western blotting after polyacrylamide gel electrophoresis. This method does not detect SAF (which are dissolved prior to electrophoresis), but the constituent protein, modified PrP. The protease resistance of modified PrP allows it to be distinguished from normal PrP, which is much more sensitive to digestion with proteases (Hope et al., 19X8; Scott et al., 1 990b).
The modified form of PrP can be detected in sections of BSE-affected brain by immunocytochemical staining with antibodies to SAF. The primary amino acid sequence of normal PrP is highly conserved in different species so the antibodies do not necessarily have to be raised against SAF purified from affected cattle. Since anti-SAF antibodies recognize normal PrP, this has to be removed by treatment with proteases. Little work of this kind has been published on BSE but it has been shown that the amyloid plaques occasionally seen in brain are immunocytochemically positive for PrP (Wells, Wilesmith and McGill, 1991).
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