Elevated levels of serum S100 beta protein in scrapie hamsters.

Recently, immunoluminometric analysis revealed elevated levels of serum S100 beta protein in sporadic and genetic cases of Creutzfeldt-Jakob disease (CJD) as compared to demented and nondemented control patients (Otto et al, 1998). S100 beta protein is a well established marker for the activation of astrocytes and also shows enhanced histochemical presentation in Alzheimer's disease (Sheng et al, 1997). Thus, it will be necessary to precisely assess the specificity of the new test for human transmissible spongiform encephalopathies (TSE) in larger study populations. Notwithstanding the need for further case-control studies, the high homology of S100 beta protein in man and cattle (Jensen et al, 1985) suggests that the protein might be used for the diagnosis of bovine spongiform encephalopathy in clinically inconspicuous animals as a protective measure for human health. This requires to first establish whether abnormal concentrations of S100 beta protein can also be found in infected individuals, either in animals exposed to the transmissible agent or in humans suffering from new variant CreutzfeldtJakob disease (vCJD), a form of CJD most likely originating from bovine spongiform encephalopathy (BSE) (Bruce et al, 1997, Hill et al, 1997). Therefore, we investigated serum levels of S100 beta protein in scrapie-infected hamsters, a wellestablished animal model for transmissible spongiform encephalopathies. The analysis was performed in 36 Syrian golden hamsters having reached the terminal stage of disease after intracerebral or intraperitoneal infection (18 animals each) with scrapie strain 263K, and in 23 non-infected controls. All hamsters for the i.c.series were taken from bioassay experiments. These recipients had been inoculated with various amounts of infectivity and showed incubation times in the range of 83 ± 146 days. The i.p.-infected animals received a uniform dose of agent (2*10 LD50) and developed scrapie at 111+11 days post infection (d.p.i.). For euthanasia animals were exposed to CO2 and 2 ml of blood were removed by puncture of the right cardiac ventricle immediately after cessation of breathing. The blood was incubated for 15 min at 378C and centrifuged for 10 min at 10000 r.p.m. (Eppendorf centrifuge). The serum was separated from the blood clot and subsequently kept frozen until analysis. In order to prevent an artefactual increase in S100 beta protein concentration (data not shown) isolation and freezing of the serum was accomplished within 3 h after blood collection. The determination of serum S100 beta protein levels in blinded samples from scrapie-infected and normal hamsters according to the previously published method (Otto et al, 1998) revealed a significant difference between the infected and the control group (P50.0001; Mann-Whitney U-test). There was also a difference between i.p. and i.c. infected animals, although on a lower level of significance (P50.05; Mann-Whitney U-test). As shown in Figure 1 hamsters with terminal scrapie exhibited elevated levels of serum S100 beta protein in the range of 78 ± 1021 pg/ml (median: 226 pg/ml; mean: 304 pg/ml) after i.c.and 23 ± 451 pg/ml (median: 130 pg/ml; mean: 164 pg/ml) after i.p.-infection. Only 10 ± 107 pg/ml (median: 20 pg/ml; mean: 32 pg/ml) were found in the non-infected animals. The previous case-control study in humans revealed about fourfold higher median serum levels of S100 beta protein in CJD cases (median: 395 pg/ ml; mean: 387 pg/ml) than in patients with non-TSE diseases (median: 109 pg/ml; mean: 177 pg/ml) (Otto et al, 1998) . The median serum concentrations of the protein in i.c.and i.p.-infected scrapie hamsters were considerably lower, but corre-