"Imaging Molecular and Trace Element Changes in Scrapie-Infected Nervous Tissue with a Time Course Study"

Ariane Kretlow, Brookhaven National Laboratory

Scrapie is a fatal neurodegenerative disorder that involves the misfolding of a naturally occurring protein in the central and peripheral nervous system. During the pathogenesis, the α-helical rich cellular prion protein (PrPC) gets converted into its abnormal isoform (PrPSc), which has a predominant β-sheet content. It is believed that transition metals such as copper and zinc play an important role in the neurodegenerative process associated with prion infection. In this study, synchrotron x-ray fluorescence (XRF) microprobe was used to determine the concentration and distribution of Zn, Cu, Fe and Ca in dorsal root ganglia (DRG) of orally 263K scrapie infected hamster, while Fourier Transform InfraRed Microspectroscopy (FTIRM) determined changes in protein structure and distribution throughout pathogenesis. Results showed that early scrapie pathogenesis was characterized by excessive levels of extracellular zinc and copper, which are likely to be correlated to prion-prion interactions, internalization of PrPC and conformational changes of cellular prion protein towards the pathogenic form. Furthermore, increased total iron and calcium levels are indicative of the involvement of oxidative stress and apoptosis, respectively. FTIRM revealed a significant increase in protein expression, yet the intermolecular β-sheet protein content was significantly lower than for controls. We suggest that the pre-clinical stages of scrapie are characterized by an overexpression of proteins low in β-sheet content. As the disease progressed, intermolecularly bound β-sheet increased significantly. Immunostaining with a PrP-specific antibody, 3F4, confirmed that this increase was partly but not solely due to the formation of PrPSc in the tissue, indicating that the formation of other β-sheet rich proteins occurred as well. At the terminal stage of the disease, the relative protein expression declined significantly, possibly due to degeneration and death of neurons. Finally, our findings of significantly increased copper and zinc levels at terminal stage might be a result of disturbed metal homeostases at advanced scrapie stages, and once again prove their pivotal role in neurodegenerative diseases.