Pathological prion protein in muscles of hamsters and mice infected with rodent-adapted BSE or vCJD

Footnotes

Here, we report on PrP^TSE deposition in muscle tissue of hamsters and mice infected experimentally with BSE or variant CreutzfeldtJakob disease (vCJD) agents. We have examined muscles from terminally ill rodents inoculated intracerebrally with hamster-adapted BSE (isolate BSE-H; Thomzig et al., 2004b), mouse-adapted BSE (isolate 6PB1; Maignien et al., 1999) or mouse-adapted vCJD (Cervenakova et al., 2003) for the presence of PrP^TSE. The recipients inoculated with these TSE agents [50 or 30 µl 1 % (w/v) brain homogenate from terminally diseased hamsters or mice, respectively] showed incubation times of 287±28, 142±9 and 140±11 days until terminal disease (expressed as means±SD), respectively. Western blot and immunohistochemical detection of PrP^TSE were performed as described previously (Thomzig et al., 2004a), with the exception that, in murine samples, we used the monoclonal anti-PrP antibody ICSM-18 (D-Gen Ltd). The specificity of this antibody for the detection of PrP has been demonstrated previously (White et al., 2003). Our study revealed widespread accumulation of PrP^TSE in muscle tissue from animals in all three experimental models (Table 1).

Table 1. Detection of PrPTSE in muscles of rodents infected intracerebrally with BSE or vCJD agents Data represent the number of animals with PrPTSE in muscles/number of animals examined. Control, specimen from uninfected animals. ND, Not done.

Muscle extracts from terminally ill C57Bl mice infected intracerebrally with the 6PB1 BSE agent showed substantial PrP^TSE deposition in many of the skeletal muscles examined (Fig. 1b, lanes 13). We also observed similar deposits of PrP^TSE in muscles of C57Bl mice inoculated with the mouse-adapted vCJD agent (Fig. 1b, lanes 46). Positive muscle specimens from BSE- or vCJD-infected mice showed an approximately 500- to 1000-fold lower concentration of PrP^TSE than in the brain, similar to what has been reported previously for scrapie-infected mice (Bosque et al., 2002).

All of this special interest in muscle involvement derives from the fact that meat, and particularly beef, is a staple of the human diet and would thus constitute a public-health risk if it were found to be infectious in cattle or small ruminants like sheep and goats (European Food Safety Authority, 2005) with TSEs. Studies to investigate the involvement of muscle in the pathogenesis of TSEs date back to a report on scrapie infectivity in the muscle of an affected goat in 1962 (Pattison & Millson, 1962) that remained, for decades, the sole successful transmission attempt. However, increasingly sensitive methodologies developed within the past few years have documented the presence of PrP^TSE in rodents infected experimentally with various strains of TSE and, as noted above, in both natural and experimental scrapie infections of sheep (Table 2). Although the detection of PrP^TSE cannot be equated directly to the presence of infectious TSE agent, a consistent association between PrP^TSE and infectivity was found when these two parameters were assayed in muscles of scrapie-infected hamsters and mice (Bosque et al., 2002; Thomzig et al., 2004a).

Table 2. Reports on the detection of TSE infectivity or PrPTSE in muscles sCJD, Sporadic CreutzfeldtJakob disease; ND, not done.

It must be borne in mind that the data of this study have been obtained from intracerebrally infected rodents, whereas natural transmission of scrapie and BSE probably occurs via the oral route. However, the relevance of the reported findings is corroborated by the detection of PrP^TSE in muscles of sheep and hamsters after both oral and intracerebral infection with scrapie (Andréoletti et al., 2004; Thomzig et al., 2004a; A. Thomzig, unpublished data for intracerebrally infected hamsters). If rodents challenged intracerebrally with BSE or vCJD agents mirror the pathophysiology of muscle targeting in ovine BSE and human vCJD to a similar extent, this would point to muscles as reservoirs for infectivity in these diseases. Thus, our findings emphasize the need for further assessment of the risks for public health that may result from prions in skeletal muscle (Bosque et al., 2002), either by oral exposure to BSE in ruminant meat products or by surgical exposure to human vCJD.

This is the first report showing deposition of BSE- and vCJD-associated PrP^TSE in muscle tissue. Kinetic studies in intracerebrally and perorally challenged animals will be necessary to further reveal whether BSE- and vCJD-related agents target muscle tissue prior to the onset of clinical symptoms after either highly invasive inoculation or alimentary infection.