Neurogranin and Neurofilament Light Chain as Preclinical Biomarkers in Scrapie

Abstract

Prion diseases are diagnosed in the symptomatic stage, when the neuronal damage is spread throughout the central nervous system (CNS). The assessment of biological features that allow the detection of asymptomatic cases is needed, and, in this context, scrapie, where pre-symptomatic infected animals can be detected through rectal biopsy, becomes a good study model. Neurogranin (Ng) and neurofilament light chain (NfL) are proteins that reflect synaptic and axonal damage and have been studied as cerebrospinal fluid (CSF) biomarkers in different neurodegenerative disorders. In this study, we evaluated Ng and NfL both at the protein and transcript levels in the CNS of preclinical and clinical scrapie-affected sheep compared with healthy controls and assessed their levels in ovine CSF. The correlation between these proteins and the main neuropathological events in prion diseases, PrPSc deposition and spongiosis, was also assessed. The results show a decrease in Ng and NfL at the protein and gene expression levels as the disease progresses, and significant changes between the control and preclinical animals. On the contrary, the CSF levels of NfL increased throughout the progression of the disease. Negative correlations between neuropathological markers of prion disease and the concentration of the studied proteins were also found. Although further research is needed, these results suggest that Ng and NfL could act as biomarkers for neurodegeneration onset and intensity in preclinical cases of scrapie.

5-Methylcytosine and 5-Hydroxymethylcytosine in Scrapie-Infected Sheep and Mouse Brain Tissues

“…The scrapie characterization of the sheep used in this study was performed previously in an earlier publication from our group [ 34 ].…”

“…Sheep in the preclinical stage were identified by immunohistochemical analysis of rectal mucosa biopsies and sacrificed before clinical signs were detectable by pentobarbital overdose, whereas animals in the clinical stage were identified by the observation of scrapie-related symptoms. These animals correspond to those used previously in an assessment study of neurogranin and neurofilament light chain as preclinical biomarkers in scrapie [ 34 ].…”

“…PrP Sc was detected by immunohistochemistry using the monoclonal primary antibody L42 (R-Biopharm, Darmstadt, Germany) at 1:500 dilution for 30 min after formic acid treatment and proteinase K digestion, as previously described [ 77 ]. The histopathological evaluation of these sheep was performed in a previous study [ 34 ].…”

Endoplasmic reticulum stress and ubiquitin-proteasome system impairment in natural scrapie

“…Brain samples from sheep infected with a natural prion disease and control samples were obtained from the tissue bank of Centro de Encefalopatias y Enfermedades Transmisibles Emergentes of the University of Zaragoza. Sample collection and demographic details of animals were described in a previous study (Betancor et al, 2022). Thus, we used brain samples from 21 female Rasa Aragonesa sheep.…”

“…These samples were stained with hematoxylin and eosin to analyze spongiform lesions. Immunohistochemical detection of prion protein was performed using the monoclonal primary antibody L42 (1:500, R-Biopharm, Darmstadt, Germany) after formic acid treatment and proteinase K digestion, as described in a previous study (Betancor et al, 2022).…”