Saturday, January 05, 2013

Immunohistochemical Detection of Disease- Associated Prion Protein in the Peripheral Nervous System in Experimental H-Type Bovine Spongiform Encephalopathy

Immunohistochemical Detection of Disease- Associated Prion Protein in the Peripheral Nervous System in Experimental H-Type Bovine Spongiform Encephalopathy



H. Okada1, Y. Iwamaru1, T. Yokoyama1, and S. Mohri1



Abstract



H-type bovine spongiform encephalopathy (BSE) has been identified in aged cattle in Europe and North America. To determine the localization of disease-associated prion protein (PrPSc) in the peripheral nerve tissues of cattle affected with H-type BSE, we employed highly sensitive immunohistochemical and immunofluorescence techniques with the tyramide signal amplification (TSA) system. PrPSc deposition was detected in the inferior ganglia, sympathetic nerve trunk, vagus nerve, spinal nerves, cauda equina, and adrenal medulla, using this system. Notably, granular PrPSc deposits were present mainly in the Schwann cells and fibroblastlike cells and occasionally along certain nerve fibers at the surface of the axons. In the adrenal gland, PrPSc immunolabeling was observed within the sympathetic nerve fibers and nerve endings in the adrenal medulla. Although our results were limited to only 3 experimental cases, these results suggest that the TSA system, a highly sensitive immunohistochemical procedure, may help in elucidating the peripheral pathogenesis of H-type BSE.



Keywords



atypical bovine spongiform encephalopathy, H-type, prion, tyramide amplification, peripheral nervous system



snip...




Neuropil, but not neuronal, vacuolation was obvious in all brain areas, especially in the nuclei of the thalamus and brainstem. Immunolabeled PrPSc was widely distributed throughout the brain and spinal cord (Fig. 1). Eight patterns of PrPSc immunolabeling including intraneuronal, perineuronal, intraglial, linear, fine particulate, coarse granular, stellate, and plaques were identified in the brain. The most conspicuous pattern of PrPSc was that of fine particulate and coarse granular deposits in the cerebral and cerebellar cortices and the nuclei of the thalamus, brainstem, and cerebellum. Stellate-type PrPSc deposition was detected in the cerebral and cerebellar cortices, basal ganglia, thalamus, hypothalamus, and hippocampus. Intraglial-type PrPSc deposition was conspicuous throughout the white matter of the brain and spinal cord. Spherical PrPSc plaques were sparsely located in the thalamus, basal ganglia, midbrain, pons, deeper layers of the cerebral cortices, and subcortical white matter. Immunolabeled PrPSc was observed in the retina, neurohypophysis, and optic nerve. In the trigeminal and dorsal root ganglia, PrPSc was mainly found accumulated in both ganglionic and satellite cells, using the conventional polymer immunodetection method as described previously.9 In addition to the extracerebral tissues described above, positive PrPSc immunolabeling was detected in the adrenal gland, cauda equina, cervical spinal nerves, facial nerve, hypoglossal nerve, vagus nerve, sciatic nerve, and ganglia, such as the inferior ganglion of the vagus nerve, superior cervical ganglion, stellate ganglion, ganglia of the sympathetic trunk, and celiac and mesenteric ganglion complex, with the use of the TSA-biotin system. In these ganglia, not all neurons of ganglia exhibited intracytoplasmic labeling, but labeling was evident in the Schwann cells or fibroblast-like cells (Figs. 2, 3). In the cauda equina, dorsal roots of the cervical spinal nerves, facial nerve, hypoglossal nerve, vagus nerve, and sciatic nerve, granular PrPSc aggregate was mainly located in Schwann cells. In the adrenal gland, the PrPSc signal was evident at the intercellular fine processes of the nerve endings between the chromaffin cells of the adrenal medulla (Fig. 4). Immunolabeled PrPSc was not detected in the enteric nervous system such as the myenteric and submucosal plexi of the digestive tract. No specific immunolabeling was detectable in the lymphoid tissues with the TSA-biotin system.




Using the dual or triple immunofluorescence technique, localization of PrPSc was visible in the ganglionic and satellite cells of the ganglia in the merged images (Fig. 5). In addition, the merged image showed that PrPSc granules were rarely located at the periphery of axons or within the axons adjacent to the ganglia (Fig. 6). Moreover, granular PrPSc was observed outside S100-positive cells (Fig. 7). In the cauda equina and spinal nerves, PrPSc coexisted mostly in the periphery of Schwann cells labeled with MBP and S100 (Fig. 8). No PrPSc immunolabeling was detected in the compact layers of the myelin sheath. In addition, no positive signal was detected in the sections from uninfected controls incubated with mAb F99/97.6.1 by both TSA-biotin and TSA-fluorescence methods, and no background immunostaining was observed in any sections of H-type infected animals by both methods when non-immune mouse and rabbit IgG, or PBS, were applied to the sections instead of the primary PrP-antibody (Figs. 9, 10).





snip...see full text ;














snip...




Unfortunately, a detailed and all-encompassing analysis of neuropathology and topographical distribution of immunolabeled PrPSc in H-type BSE-affected cattle could not be performed, since only the obex region is routinely sampled for BSE surveillance testing and the remaining brain as well as the carcasses are not available in most countries [3,10,12,13,24-27]. Recently, clinical signs and biochemical properties of experimental German H-type BSE cases have been reported [20]. The primary objective of this study was to investigate the transmissibility of H-type BSE, using a field isolate detected in the active surveillance program in Canada [12]. The secondary objective was to extend the knowledge of the topographical distribution and deposition patterns of immunolabeled PrPSc in H-type BSE.




snip...






In addition, the present data will support risk assessments in some peripheral tissues derived from cattle affected with H-type BSE.











Friday, March 09, 2012



Experimental H-type and L-type bovine spongiform encephalopathy in cattle: observation of two clinical syndromes and diagnostic challenges Research article








Thursday, June 23, 2011


Experimental H-type bovine spongiform encephalopathy characterized by plaques and glial- and stellate-type prion protein deposits







P.4.23 Transmission of atypical BSE in humanized mouse models



Liuting Qing1, Wenquan Zou1, Cristina Casalone2, Martin Groschup3, Miroslaw Polak4, Maria Caramelli2, Pierluigi Gambetti1, Juergen Richt5, Qingzhong Kong1 1Case Western Reserve University, USA; 2Instituto Zooprofilattico Sperimentale, Italy; 3Friedrich-Loeffler-Institut, Germany; 4National Veterinary Research Institute, Poland; 5Kansas State University (Previously at USDA National Animal Disease Center), USA



Background: Classical BSE is a world-wide prion disease in cattle, and the classical BSE strain (BSE-C) has led to over 200 cases of clinical human infection (variant CJD). Atypical BSE cases have been discovered in three continents since 2004; they include the L-type (also named BASE), the H-type, and the first reported case of naturally occurring BSE with mutated bovine PRNP (termed BSE-M). The public health risks posed by atypical BSE were largely undefined.



Objectives: To investigate these atypical BSE types in terms of their transmissibility and phenotypes in humanized mice.



Methods: Transgenic mice expressing human PrP were inoculated with several classical (C-type) and atypical (L-, H-, or Mtype) BSE isolates, and the transmission rate, incubation time, characteristics and distribution of PrPSc, symptoms, and histopathology were or will be examined and compared.



Results: Sixty percent of BASE-inoculated humanized mice became infected with minimal spongiosis and an average incubation time of 20-22 months, whereas only one of the C-type BSE-inoculated mice developed prion disease after more than 2 years. Protease-resistant PrPSc in BASE-infected humanized Tg mouse brains was biochemically different from bovine BASE or sCJD. PrPSc was also detected in the spleen of 22% of BASE-infected humanized mice, but not in those infected with sCJD. Secondary transmission of BASE in the humanized mice led to a small reduction in incubation time.



*** The atypical BSE-H strain is also transmissible with distinct phenotypes in the humanized mice, but no BSE-M transmission has been observed so far.




Discussion: Our results demonstrate that BASE is more virulent than classical BSE, has a lymphotropic phenotype, and displays a modest transmission barrier in our humanized mice. BSE-H is also transmissible in our humanized Tg mice. The possibility of more than two atypical BSE strains will be discussed.




Supported by NINDS NS052319, NIA AG14359, and NIH AI 77774.









P26 TRANSMISSION OF ATYPICAL BOVINE SPONGIFORM ENCEPHALOPATHY (BSE) IN HUMANIZED MOUSE MODELS



Liuting Qing1, Fusong Chen1, Michael Payne1, Wenquan Zou1, Cristina Casalone2, Martin Groschup3, Miroslaw Polak4, Maria Caramelli2, Pierluigi Gambetti1, Juergen Richt5*, and Qingzhong Kong1 1Department of Pathology, Case Western Reserve University, Cleveland, OH 44106, USA; 2CEA, Istituto Zooprofilattico Sperimentale, Italy; 3Friedrich-Loeffler-Institut, Germany; 4National Veterinary Research Institute, Poland; 5Kansas State University, Diagnostic Medicine/Pathobiology Department, Manhattan, KS 66506, USA. *Previous address: USDA National Animal Disease Center, Ames, IA 50010, USA



Classical BSE is a world-wide prion disease in cattle, and the classical BSE strain (BSE-C) has led to over 200 cases of clinical human infection (variant CJD). Two atypical BSE strains, BSE-L (also named BASE) and BSE-H, have been discovered in three continents since 2004. The first case of naturally occurring BSE with mutated bovine PrP gene (termed BSE-M) was also found in 2006 in the USA. The transmissibility and phenotypes of these atypical BSE strains/isolates in humans were unknown. We have inoculated humanized transgenic mice with classical and atypical BSE strains (BSE-C, BSE-L, BSE-H) and the BSE-M isolate. We have found that the atypical BSE-L strain is much more virulent than the classical BSE-C.



*** The atypical BSE-H strain is also transmissible in the humanized transgenic mice with distinct phenotype, but no transmission has been observed for the BSE-M isolate so far.



III International Symposium on THE NEW PRION BIOLOGY: BASIC SCIENCE, DIAGNOSIS AND THERAPY 2 - 4 APRIL 2009, VENEZIA (ITALY)











I ask Professor Kong ;



Thursday, December 04, 2008 3:37 PM


Subject: RE: re--Chronic Wating Disease (CWD) and Bovine Spongiform Encephalopathies (BSE): Public Health Risk Assessment



''IS the h-BSE more virulent than typical BSE as well, or the same as cBSE, or less virulent than cBSE? just curious.....''



Professor Kong reply ;



.....snip



''As to the H-BSE, we do not have sufficient data to say one way or another, but we have found that H-BSE can infect humans. I hope we could publish these data once the study is complete. Thanks for your interest.'' Best regards, Qingzhong Kong, PhD Associate Professor Department of Pathology Case Western Reserve University Cleveland, OH 44106 USA




END...TSS




Thursday, December 04, 2008 2:37 PM


"we have found that H-BSE can infect humans." personal communication with Professor Kong. ...TSS


BSE-H is also transmissible in our humanized Tg mice.


The possibility of more than two atypical BSE strains will be discussed.


Supported by NINDS NS052319, NIA AG14359, and NIH AI 77774.








SNIP...




Friday, May 11, 2012


Experimental H-type bovine spongiform encephalopathy characterized by plaques and glial- and stellate-type prion protein deposits






Thursday, June 21, 2012


Clinical and Pathologic Features of H-Type Bovine Spongiform Encephalopathy Associated with E211K Prion Protein Polymorphism







Tuesday, November 02, 2010



IN CONFIDENCE



The information contained herein should not be disseminated further except on the basis of "NEED TO KNOW".



BSE - ATYPICAL LESION DISTRIBUTION (RBSE 92-21367) statutory (obex only) diagnostic criteria CVL 1992









2009 UPDATE ON ALABAMA AND TEXAS MAD COWS 2005 and 2006








let's take a closer look at this new prionpathy or prionopathy, and then let's look at the g-h-BSEalabama mad cow.




This new prionopathy in humans? the genetic makeup is IDENTICAL to the g-h-BSEalabama mad cow, the only _documented_ mad cow in the world to date like this, ......wait, it get's better. this new prionpathy is killing young and old humans, with LONG DURATION from onset of symptoms to death, and the symptoms are very similar to nvCJD victims, OH, and the plaques are very similar in some cases too, bbbut, it's not related to the g-h-BSEalabama cow, WAIT NOW, it gets even better, the new human prionpathy that they claim is a genetic TSE, has no relation to any gene mutation in that family. daaa, ya think it could be related to that mad cow with the same genetic make-up ??? there were literally tons and tons of banned mad cow protein in Alabama in commerce, and none of it transmitted to cows, and the cows to humans there from ??? r i g h t $$$




ALABAMA MAD COW g-h-BSEalabama




In this study, we identified a novel mutation in the bovine prion protein gene (Prnp), called E211K, of a confirmed BSE positive cow from Alabama, United States of America. This mutation is identical to the E200K pathogenic mutation found in humans with a genetic form of CJD. This finding represents the first report of a confirmed case of BSE with a potential pathogenic mutation within the bovine Prnp gene. We hypothesize that the bovine Prnp E211K mutation most likely has caused BSE in "the approximately 10-year-old cow" carrying the E221K mutation.














her healthy calf also carried the mutation (J. A. Richt and S. M. Hall PLoS Pathog. 4, e1000156; 2008).




This raises the possibility that the disease could occasionally be genetic in origin. Indeed, the report of the UK BSE Inquiry in 2000 suggested that the UK epidemic had most likely originated from such a mutation and argued against the scrapierelated assumption. Such rare potential pathogenic PRNP mutations could occur in countries at present considered to be free of BSE, such as Australia and New Zealand. So it is important to maintain strict surveillance for BSE in cattle, with rigorous enforcement of the ruminant feed ban (many countries still feed ruminant proteins to pigs). Removal of specified risk material, such as brain and spinal cord, from cattle at slaughter prevents infected material from entering the human food chain. Routine genetic screening of cattle for PRNP mutations, which is now available, could provide additional data on the risk to the public. Because the point mutation identified in the Alabama animals is identical to that responsible for the commonest type of familial (genetic) CJD in humans, it is possible that the resulting infective prion protein might cross the bovine–human species barrier more easily. Patients with vCJD continue to be identified. The fact that this is happening less often should not lead to relaxation of the controls necessary to prevent future outbreaks. Malcolm A. Ferguson-Smith Cambridge University Department of Veterinary Medicine, Madingley Road, Cambridge CB3 0ES, UK e-mail: maf12@cam.ac.uk Jürgen A. Richt College of Veterinary Medicine, Kansas State University, K224B Mosier Hall, Manhattan, Kansas 66506-5601, USA




NATURE|Vol 457|26 February 2009











Saturday, August 14, 2010



BSE Case Associated with Prion Protein Gene Mutation (g-h-BSEalabama) and VPSPr PRIONPATHY (see mad cow feed in COMMERCE IN ALABAMA...TSS)










Tuesday, November 6, 2012



***Transmission of New Bovine Prion to Mice, Atypical Scrapie, BSE, and Sporadic CJD, November-December 2012 update










Comments on technical aspects of the risk assessment were then submitted to FSIS.





Comments were received from Food and Water Watch, Food Animal Concerns Trust (FACT), Farm Sanctuary, R-CALF USA, Linda A Detwiler, and Terry S. Singeltary.


This document provides itemized replies to the public comments received on the 2005 updated Harvard BSE risk assessment. Please bear the following points in mind:









Owens, Julie


From: Terry S. Singeltary Sr. [flounder9@verizon.net]


Sent: Monday, July 24, 2006 1:09 PM


To: FSIS RegulationsComments



Subject: [Docket No. FSIS-2006-0011] FSIS Harvard Risk Assessment of Bovine Spongiform Encephalopathy (BSE) Page 1 of 98










FSIS, USDA, REPLY TO SINGELTARY










U.S.A. 50 STATE BSE MAD COW CONFERENCE CALL Jan. 9, 2001
















2012 atypical L-type BSE BASE California reports





SUMMARY REPORT CALIFORNIA BOVINE SPONGIFORM ENCEPHALOPATHY CASE INVESTIGATION JULY 2012



Summary Report BSE 2012



Executive Summary










Saturday, August 4, 2012



Update from APHIS Regarding Release of the Final Report on the BSE Epidemiological Investigation










Saturday, August 4, 2012



*** Final Feed Investigation Summary - California BSE Case - July 2012










Saturday, December 15, 2012




Bovine spongiform encephalopathy: the effect of oral exposure dose on attack rate and incubation period in cattle -- an update 5 December 2012












Thursday, December 20, 2012



OIE GROUP RECOMMENDS THAT SCRAPE PRION DISEASE BE DELISTED AND SAME OLD BSe WITH BOVINE MAD COW DISEASE











Tuesday, December 25, 2012



A Quantitative Assessment of the Amount of Prion Diverted to Category 1 Materials and Wastewater During Processing











2011 Monday, September 26, 2011



L-BSE BASE prion and atypical sporadic CJD











Tuesday, June 26, 2012



Creutzfeldt Jakob Disease Human TSE report update North America, Canada, Mexico, and USDA PRION UNIT as of May 18, 2012



type determination pending Creutzfeldt Jakob Disease (tdpCJD), is on the rise in Canada and the USA










Sunday, December 2, 2012



CANADA 19 cases of mad cow disease SCENARIO 4: ‘WE HAD OUR CHANCE AND WE BLEW IT’









Monday, July 23, 2012



The National Prion Disease Pathology Surveillance Center July 2012









Tuesday, December 25, 2012



CREUTZFELDT JAKOB TSE PRION DISEASE HUMANS END OF YEAR REVIEW DECEMBER 25, 2012










Monday, December 31, 2012



Creutzfeldt Jakob Disease and Human TSE Prion Disease in Washington State, 2006–2011-2012









TSS

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