Intraepithelial and Interstitial Deposition of Pathological Prion Protein in Kidneys of Scrapie-Affected Sheep

Prions have been documented in extra-neuronal and extra-lymphatic tissues of humans and various ruminants affected by Transmissible Spongiform Encephalopathy (TSE). The presence of prion infectivity detected in cervid and ovine blood tempted us to reason that kidney, the organ filtrating blood derived proteins, may accumulate disease associated PrPSc. We collected and screened kidneys of experimentally, naturally scrapie-affected and control sheep for renal deposition of PrPSc from distinct, geographically separated flocks. By performing Western blot, PET blot analysis and immunohistochemistry we found intraepithelial (cortex, medulla and papilla) and occasional interstitial (papilla) deposition of PrPSc in kidneys of scrapie-affected sheep. Interestingly, glomerula lacked detectable signals indicative of PrPSc. PrPSc was also detected in kidneys of subclinical sheep, but to significantly lower degree. Depending on the stage of the disease the incidence of PrPSc in kidney varied from approximately 27% (subclinical) to 73.6% (clinical) in naturally scrapie-affected sheep. Kidneys from flocks without scrapie outbreak were devoid of PrPSc. Here we demonstrate unexpectedly frequent deposition of high levels of PrPSc in ovine kidneys of various flocks. Renal deposition of PrPSc is likely to be a pre-requisite enabling prionuria, a possible co-factor of horizontal prion-transmission in sheep

Aerosols Transmit Prions to Immunocompetent and Immunodeficient Mice

Prions, the agents causing transmissible spongiform encephalopathies, colonize the brain of hosts after oral, parenteral, intralingual, or even transdermal uptake. However, prions are not generally considered to be airborne. Here we report that inbred and crossbred wild-type mice, as well as tga20 transgenic mice overexpressing PrPC, efficiently develop scrapie upon exposure to aerosolized prions. NSE-PrP transgenic mice, which express PrPC selectively in neurons, were also susceptible to airborne prions. Aerogenic infection occurred also in mice lacking B- and T-lymphocytes, NK-cells, follicular dendritic cells or complement components. Brains of diseased mice contained PrPSc and transmitted scrapie when inoculated into further mice. We conclude that aerogenic exposure to prions is very efficacious and can lead to direct invasion of neural pathways without an obligatory replicative phase in lymphoid organs. This previously unappreciated risk for airborne prion transmission may warrant re-thinking on prion biosafety guidelines in research and diagnostic laboratories

Correction: Aerosols transmit prions to immunocompetent and immunodeficient mice

<p>Correction: Aerosols transmit prions to immunocompetent and immunodeficient mice</p

PET blot analysis identifying PrP^Sc in the tubular structures of the cortex and medulla in naturally scrapie-sick Sarda sheep.

<p>(A) Brains of scrapie-sick (left) and healthy sheep (right) as a control for PET blot analysis procedure. Scale bar = 100 µm. (B) Low magnification of transverse paraffin sections from kidneys of a naturally scrapie-sick and a healthy sheep demonstrating PrP^Sc in the medullary and cortical regions of the kidney. Dotted lines indicate the locus of the kidney capsule or the edge of the tissue section. The small transparent box indicates the region shown at higher magnification and resolution in figure C. Scale bar = 1 cm. (C) PrP^Sc deposits and tubular structure fit together in the cortical part of the kidney in scrapie-sick sheep. Scale bar = 100 µm. (D) PrP^Sc is also found to a high degree in the collecting ducts of the medulla (arrowhead) as well as in the papillae (arrow) of kidneys derived from naturally scrapie infected sheep. Scale bar = 30 µm.</p

PrP^Sc detected by immunohistochemistry in scrapie-affected Sarda sheep.

<p>(A) Immunohistochemical analysis of the cortex, medulla and papillae of naturally scrapie-sick sheep and control sheep derived from scrapie-free flocks. Antibodies 2G11 and F99 were used. Scale bar = 30 µm. (B) Immunofluorescence using 2G11 identifies tubular deposits in the medullar-cortical junction of the kidney of a scrapie-sick sheep. Scrapie-free control is devoid of detectable positive signal.</p

Number (N), age, PrP genotype and clinical status of the sheep included in the study.

<p>Sheep are grouped as scrapie-affected sheep or negative control. The number (N) of the flocks from which the sheep were selected is also indicated.</p>*<p>Sheep from scrapie-affected flock</p

Western Blotting PrP^Sc deposition in scrapie-affected Sarda sheep.

<p>(A) Detection of PrP^Sc by conventional Western blot in the cortical and medullar part of kidneys. Brain and kidney homogenates were treated with (+) or without (−) proteinase K (PK), amounts of kidney and brain represented in each line are 1mg and 1g respectively. S.b. = brain derived from a naturally scrapie-sick sheep; h.b. = brain derived from a scrapie-free sheep control; h.k. = kidney derived from a scrapie-free sheep control, s.k. = kidney derived from a naturally scrapie-sick sheep. kD = kilo Dalton, M = medulla, C = cortex. (B) Detection of PrP^Sc by NaPTA Western blot in kidney homogenate derived from different Sarda sheep. Substantial individual variation of the Western blotting signal was observed among the sheep examined. Spike control = brain homogenate derived from a scrapie–sick sheep spiked into negative control kidney homogenate. h.b. = brain derived from a scrapie-free sheep control. 2000–4000 µg of renal homogenate were used as a starting material to perform NaPTA precipitation.</p