Differential overexpression of SERPINA3 in human prion diseases.

Prion diseases are fatal neurodegenerative disorders with sporadic, genetic or acquired etiologies. The molecular alterations leading to the onset and the spreading of these diseases are still unknown. In a previous work we identified a five-gene signature able to distinguish intracranially BSE-infected macaques from healthy ones, with SERPINA3 showing the most prominent dysregulation. We analyzed 128 suitable frontal cortex samples, from prion-affected patients (variant Creutzfeldt-Jakob disease (vCJD) n = 20, iatrogenic CJD (iCJD) n = 11, sporadic CJD (sCJD) n = 23, familial CJD (gCJD) n = 17, fatal familial insomnia (FFI) n = 9, Gerstmann-Sträussler-Scheinker syndrome (GSS)) n = 4), patients with Alzheimer disease (AD, n = 14) and age-matched controls (n = 30). Real Time-quantitative PCR was performed for SERPINA3 transcript, and ACTB, RPL19, GAPDH and B2M were used as reference genes. We report SERPINA3 to be strongly up-regulated in the brain of all human prion diseases, with only a mild up-regulation in AD. We show that this striking up-regulation, both at the mRNA and at the protein level, is present in all types of human prion diseases analyzed, although to a different extent for each specific disorder. Our data suggest that SERPINA3 may be involved in the pathogenesis and the progression of prion diseases, representing a valid tool for distinguishing different forms of these disorders in humans.

Postmortem findings in a case of variant Creutzfeldt-Jakob disease treated with intraventricular pentosan polysulfate.

BACKGROUND: A small number of patients with variant Creutzfeldt-Jakob disease (vCJD) have been treated with intraventicular pentosan polysulfate (iPPS) and extended survival has been reported in some cases. To date, there have been no reports on the findings of postmortem examination of the brain in treated patients and the reasons for the extended survival are uncertain. We report on the neuropathological findings in a case of vCJD treated with PPS. METHODS: Data on survival in vCJD is available from information held at the National CJD Research and Surveillance Unit and includes the duration of illness in 176 cases of vCJD, five of which were treated with iPPS. One of these individuals, who received iPPS for 8 years and lived for 105 months, underwent postmortem examination, including neuropathological examination of the brain. RESULTS: The mean survival in vCJD is 17 months, with 40 months the maximum survival in patients not treated with PPS. In the 5 patients treated with PPS survival was 16 months, 45 months, 84 months, 105 months and 114 months. The patient who survived 105 months underwent postmortem examination which confirmed the diagnosis of vCJD and showed severe, but typical, changes, including neuronal loss, astrocytic gliosis and extensive prion protein (PrP) deposition in the brain. The patient was also given PPS for a short period by peripheral infusion and there was limited PrP immunostaining in lymphoreticular tissues such as spleen and appendix. CONCLUSIONS: Treatment with iPPS did not reduce the overall neuropathological changes in the brain. The reduced peripheral immunostaining for PrP may reflect atrophy of these tissues in relation to chronic illness rather than a treatment effect. The reason for the long survival in patients treated with iPPS is unclear, but a treatment effect on the disease process cannot be excluded.

Neuropathology of the hippocampus in FTLD-Tau with Pick bodies: a study of the BrainNet Europe Consortium.

AIMS: Frontotemporal lobar degeneration with Pick bodies (Pick's disease) is characterized by the presence of tau immunoreactive spherical structures in the cytoplasm of neurones. In view of confusion about the molecular pathology of Pick's disease, we aimed to evaluate the spectrum of tau pathology and concomitant neurodegeneration-associated protein depositions in the characteristically affected hippocampus. METHODS: We evaluated immunoreactivity (IR) for tau (AT8, 3R, 4R), α-synuclein, TDP43, p62, and ubiquitin in the hippocampus, entorhinal and temporal cortex in 66 archival cases diagnosed neuropathologically as Pick's disease. RESULTS: Mean age at death was 68.2 years (range 49-96). Fifty-two (79%) brains showed 3R immunoreactive spherical inclusions in the granule cells of the dentate gyrus. These typical cases presented mainly with the behavioural variant of frontotemporal dementia, followed by progressive aphasia, mixed syndromes or early memory disturbance. α-Synuclein IR was seen only in occasional spherical tau-positive inclusions, TDP-43 IR was absent, and 4R IR was present only as neurofibrillary tangles in pyramidal neurones. Aß IR was observed in 16 cases; however, the overall level of Alzheimer's disease-related alterations was mainly low or intermediate (n = 3). Furthermore, we identified six cases with unclassifiable tauopathy. CONCLUSIONS: (i) Pick's disease may occur also in elderly patients and is characterized by a relatively uniform pathology with 3R tau inclusions particularly in the granule cells of dentate gyrus; (ii) even minor deviation from these morphological criteria suggests a different disorder; and (iii) immunohistological revision of archival cases expands the spectrum of tauopathies that require further classification.

Review: Creutzfeldt-Jakob disease: prion protein type, disease phenotype and agent strain.

The human transmissible spongiform encephalopathies or human prion diseases are one of the most intensively investigated groups of rare human neurodegenerative conditions. They are generally held to be unique in terms of their complex epidemiology and phenotypic variability, but they may also serve as a paradigm with which other more common protein misfolding disorders might be compared and contrasted. The clinico-pathological phenotype of human prion diseases appears to depend on a complex interaction between the prion protein genotype of the affected individual and the physico-chemical properties of the neurotoxic and transmissible agent, thought to comprise of misfolded prion protein. A major focus of research in recent years has been to define the phenotypic heterogeneity of the recognized human prion diseases, correlate this with molecular-genetic features and then determine whether this molecular-genetic classification of human prion disease defines the biological properties of the agent as determined by animal transmission studies. This review seeks to survey the field as it currently stands, summarize what has been learned, and explore what remains to be investigated in order to obtain a more complete scientific understanding of prion diseases and to protect public health.

RNA integrity in post mortem human variant Creutzfeldt-Jakob disease (vCJD) and control brain tissue.

AIMS: To determine premortem and post mortem factors affecting quality and yield of RNA isolated from the unique archived brain material in the UK National Creutzfeldt-Jakob Disease Surveillance Unit Brain and Tissue Bank and to compare this to control brain tissue with no neurological disease. METHODS: In parallel and in replicate, RNA was prepared from the frontal parasagittal or subfrontal cortex of samples dissected from half brains (frozen intact) or from brain samples snap frozen or placed in RNALater. A total of 350 RNA samples from 78 human autopsy cases, 21 variant Creutzfeldt-Jakob disease, 26 other neurological diseases and 31 non-neurological diseases were studied. RESULTS: There was no difference in the quality or yield of RNA isolated from variant Creutzfeldt-Jakob disease, other neurological disease and non-neurological disease brains. RNA preparations from archived frozen half brains or snap frozen autopsy samples were generally of poor quality (RNA integrity number<5). There was a highly significant negative correlation between the number of times frozen half brains had been sampled and the quality of RNA. Samples stored in RNALater provided higher-quality RNA (RNA integrity number>5). Age at death, gender, post mortem interval and freezer storage time had no effect on RNA quality. CONCLUSION: Reasonable-quality RNA can be isolated from samples dissected from archived frozen human half brains but repeated sampling results in RNA degradation. Better-quality RNA is obtained from samples placed in RNALater than from snap frozen samples. The quality and yield of RNA are not affected by age at death, gender, post mortem interval of >6 h or freezer storage time.

Variant Creutzfeldt-Jakob disease.

SUMMARY: Variant Creutzfeldt-Jakob disease (CJD) is an emerging form of human prion disease caused by oral exposure to the bovine spongiform encephalopathy agent. Most cases have occurred in the UK, but smaller numbers of cases have been identified in 10 other countries worldwide. All confirmed cases belong to a single genetic subgroup defined by methionine homozygosity at codon 129 in the prion protein gene. Variant CJD has a widespread distribution of infectivity in the body, involving lymphoid tissues during at least the latter part of the incubation period. This is unlike other forms of human prion disease, and raised concerns that the transmissible agent might also be present in blood. To date, four probable cases of variant CJD infection have been identified following transfusion of packed red blood cells from asymptomatic donors who subsequently died from variant CJD. Recently, one case of likely transmission of variant CJD infection by UK factor VIII (FVIII) concentrates has been reported in an elderly haemophilic patient in the UK, who had been treated with FVIII produced from pooled plasma to which a donor who subsequently died from variant CJD had contributed. The recipient showed no signs or symptoms of variant CJD during life, but evidence of variant CJD infection was detected in his spleen following a postmortem examination. Continued surveillance is required to investigate the prevalence of secondary variant CJD infection in other patients with bleeding disorders who have been treated with UK-sourced pooled plasma products.

The first case of protease-sensitive prionopathy (PSPr) in The Netherlands: a patient with an unusual GSS-like clinical phenotype.

An atypical case of prion disease is described in a 54-year-old Dutch man, homozygous for valine at codon 129 of the prion protein gene (PRNP). The clinical phenotype was characterised by progressive dementia, spastic paraplegia and sensorimotor polyneuropathy. The disease duration was 20 months. Genetic analysis of PRNP did not reveal any abnormalities. Neuropathologically, only mild spongiform change and a coarse granular immunohistochemical staining for the abnormal prion protein, PrP(Sc), was observed, with poorly formed plaques in the molecular layer of the cerebellar cortex. However, Western blotting showed low but detectable levels of proteinase K(PK)-resistant PrP(Sc) occurring in an unusual ladder-like profile. These features define a phenotype that corresponds to the recently described protease-sensitive prionopathy (PSPr). Our report on the first Dutch patient with PSPr further expands the spectrum of prionopathies and exemplifies the need to re-evaluate cases of atypical prion disease.

Risk reduction strategies for variant Creutzfeldt-Jakob disease transmission by UK plasma products and their impact on patients with inherited bleeding disorders.

SUMMARY: The appearance and rapid evolution of BSE in UK cattle in the mid 1980s, with compelling data supporting variant Creutzfeldt-Jakob disease (vCJD) as its human manifestation, pose a potentially severe threat to public health. Three clinical cases and one asymptomatic case of vCJD infection have been reported in UK recipients of non-leucodepleted red cell transfusions from donors subsequently diagnosed with vCJD. Plasma from both these and other donors who later developed vCJD has contributed towards plasma pools used to manufacture clotting factor concentrate. The United Kingdom Haemophilia Centre Doctors' Organisation (UKHCDO) Surveillance Study has detected asymptomatic vCJD postmortem in a haemophilic patient treated with UK plasma products including two batches of clotting factor linked to a donor who subsequently developed vCJD. Over 4000 bleeding disorder patients treated with UK plasma products are recorded on the UKHCDO National Haemophilia Database. The risk of vCJD transmission by plasma products is not known. However, public health precautions have been implemented since 2004 in all UK inherited bleeding disorder patients who received UK-sourced plasma products between 1980 and 2001 to minimize the possible risk of onward vCJD transmission. We evaluate vCJD surveillance and risk management measures taken for UK inherited bleeding disorder patients, report current data and discuss resultant challenges and future directions.

Variant CJD infection in the spleen of a neurologically asymptomatic UK adult patient with haemophilia.

SUMMARY: All UK patients with bleeding disorders treated with any UK-sourced pooled factor concentrates between 1980 and 2001 have been informed that they may be at an increased risk of infection with variant Creutzfeldt-Jakob disease (vCJD). We describe a study to detect disease-associated, protease-resistant prion protein (PrP(res)) in 17 neurologically aymptomatic patients with haemophilia considered to be at increased risk of vCJD. Materials from 11 autopsy and seven biopsy cases were analysed for PrP(res). The tissues available from each case were variable, ranging from a single biopsy sample to a wide range of autopsy tissues. A single specimen from the spleen of one autopsy case gave a strong positive result on repeated testing for PrP(res) by Western blot analysis. This tissue came from a 73-year-old male patient with no history of neurological disease, who was heterozygous (methionine/valine) at codon 129 in the prion protein gene. He had received over 9000 units of factor VIII concentrate prepared from plasma pools known to include donations from a vCJD-infected donor, and some 400,000 units not known to include donations from vCJD-infected donors. He had also received 14 units of red blood cells and had undergone several surgical and invasive endoscopic procedures. Estimates of the relative risks of exposure through diet, surgery, endoscopy, blood transfusion and receipt of UK-sourced plasma products suggest that by far the most likely route of infection in this patient was receipt of UK plasma products.

Primary postoperative chemotherapy without radiotherapy for treatment of brain tumours other than ependymoma in children under 3 years: results of the first UKCCSG/SIOP CNS 9204 trial.

BACKGROUND: Radiotherapy is an effective adjuvant treatment for brain tumours arising in very young children, but it has the potential to damage the child's developing nervous system at a crucial time - with a resultant reduction in IQ leading to cognitive impairment, associated endocrinopathy and risk of second malignancy. We aimed to assess the role of a primary chemotherapy strategy in avoiding or delaying radiotherapy in children younger than 3 years with malignant brain tumours other than ependymoma, the results of which have already been published. METHODS: Ninety-seven children were enrolled between March 1993 and July 2003 and, following diagnostic review, comprised: medulloblastoma (n=31), astrocytoma (26), choroid plexus carcinoma [CPC] (15), CNS PNET (11), atypical teratoid/rhabdoid tumours [AT/RT] (6) and ineligible (6). Following maximal surgical resection, chemotherapy was delivered every 14 d for 1 year or until disease progression. Radiotherapy was withheld in the absence of progression. FINDINGS: Over all diagnostic groups the cumulative progression rate was 80.9% at 5 years while the corresponding need-for-radiotherapy rate for progression was 54.6%, but both rates varied by tumour type. There was no clear relationship between chemotherapy dose intensity and outcome. Patients with medulloblastoma presented as a high-risk group, 83.9% having residual disease and/or metastases at diagnosis. For these patients, outcome was related to histology. The 5-year OS for desmoplastic/nodular medulloblastoma was 52.9% (95% confidence interval (CI): 27.6-73.0) and for classic medulloblastoma 33.3% (CI: 4.6-67.6); the 5-year EFS were 35.3% (CI: 14.5-57.0) and 33.3% (CI: 4.6-67.6), respectively. All children with large cell or anaplastic variants of medulloblastoma died within 2 years of diagnosis. The 5-year EFS for non-brainstem high-grade gliomas [HGGs] was 13.0% (CI: 2.2-33.4) and the OS was 30.9% (CI: 11.5-52.8). For CPC the 5-year OS was 26.67% (CI: 8.3-49.6) without RT. This treatment strategy was less effective for AT/RT with 3-year OS of 16.7% (CI: 0.8-51.7) and CNS PNET with 1-year OS of 9.1% (CI: 0.5-33.3). INTERPRETATION: The outcome for very young children with brain tumours is dictated by degree of surgical resection and histological tumour type and underlying biology as an indicator of treatment sensitivity. Overall, the median age at radiotherapy was 3 years and radiotherapy was avoided in 45% of patients. Desmoplastic/nodular sub-type of medulloblastoma has a better prognosis than classic histology, despite traditional adverse clinical features of metastatic disease and incomplete surgical resection. A subgroup with HGG and CPC are long-term survivors without RT. This study highlights the differing therapeutic challenges presented by the malignant brain tumours of early childhood, the importance of surgical approaches and the need to explore individualised brain sparing approaches to the range of malignant brain tumours that present in early childhood.

Panencephalopathic Creutzfeldt-Jakob disease in the Netherlands and the UK: clinical and pathological characteristics of nine patients.

AIMS: The panencephalopathic type of Creutzfeldt-Jakob disease (PECJD) has extensive abnormalities in cerebral white matter as well as the cortex. PECJD has rarely been described in Caucasians and debate continues on its classification and pathogenesis. We describe our experience of PECJD over a 14-year period of surveillance for CJD in the Netherlands and the UK. METHODS: Between 1993 and 2006, nine cases of PECJD were identified. Clinical, histological and biochemical characteristics of all patients were analysed and compared; all cases were classified clinically as sporadic CJD. RESULTS: The median age at onset was 57.8 years and median disease duration was 22 months. The average brain weight was 887 g. Most patients showed a two-stage clinical course with initial rapid deterioration to a state of akinetic mutism, which then persisted over a longer time scale. Neuropathological findings were characterized by severe global atrophy with status spongiosus. Cerebral white matter involvement tended to be associated with either disease duration or severity of cerebral cortical lesions. Five patients could be classified into the MM1 subtype of sporadic CJD, one patient into the MM2 subgroup and another into the MV2 subgroup. Two patients were heterozygous at codon 129 in the prion protein gene and contained both type 1 and type 2 PrP(res) isoforms in the brain. CONCLUSIONS: We believe that white matter pathology in PECJD represents an end-stage pattern that reflects secondary degeneration due to widespread cortical neuronal loss that occurs in the early part of the disease, rather than representing a primary lesion.

A quantitative study of the pathological changes in the cerebellum in 15 cases of variant Creutzfeldt-Jakob disease (vCJD).

AIMS: To determine in the cerebellum in variant Creutzfeldt-Jakob disease (vCJD): (i) whether the pathology affected all laminae; (ii) the spatial topography of the pathology along the folia; (iii) spatial correlations between the pathological changes; and (iv) whether the pathology was similar to that of the common methionine/methionine Type 1 subtype of sporadic CJD. METHODS: Sequential cerebellar sections of 15 cases of vCJD were stained with haematoxylin and eosin, or immunolabelled with monoclonal antibody 12F10 against prion protein (PrP) and studied using spatial pattern analysis. RESULTS: Loss of Purkinje cells was evident compared with control cases. Densities of the vacuolation and the protease-resistant form of prion protein (PrP(Sc)) (diffuse and florid plaques) were greater in the granule cell layer (GL) than the molecular layer (ML). In the ML, vacuoles and PrP(Sc) plaques occurred in clusters regularly distributed along the folia with larger clusters of vacuoles and diffuse plaques in the GL. There was a negative spatial correlation between the vacuoles and the surviving Purkinje cells in the ML. There was a positive spatial correlation between the vacuoles and diffuse PrP(Sc) plaques in the ML and GL. CONCLUSIONS: (i) all laminae were affected by the pathology, the GL more severely than the ML; (ii) the pathology was topographically distributed along the folia especially in the Purkinje cell layer and ML; (iii) pathological spread may occur in relation to the loop of anatomical connections involving the cerebellum, thalamus, cerebral cortex and pons; and (iv) there were pathological differences compared with methionine/methionine Type 1 sporadic CJD.

Risk of Creutzfeldt-Jakob disease transmission by ocular surgery and tissue transplantation.

Creutzfeldt-Jakob disease (CJD) is a rare, fatal neurodegenerative disease that occurs in sporadic, genetic, variant, and iatrogenic forms. The transformation of normal prion protein (PrP(C)) to the abnormal form (PrP(Sc)) is a key step in the pathogenesis of CJD and leads to the accumulation of amyloid and spongiform changes in the brain. The presence of PrP(Sc) in tissue is a surrogate marker for CJD infectivity. Sporadic CJD, whose cause is unknown, is by far the most frequent form with 1-2 cases per million population occurring every year-the genetic forms of CJD are rather rarer. The majority of variant CJD cases have occurred in the United Kingdom, where there have been four reports of transmission of vCJD by blood transfusion. The great majority of iatrogenic transmissions of CJD have resulted from the use of pituitary-derived hormones or dura mater with only a very few cases attributable to neurosurgical instruments or corneal transplants. In the absence of a validated test for CJD infectivity in eye donors, the application of appropriate donor selection criteria and the use of single-use instruments in eye banks are currently the most effective means of reducing the risk of CJD transmission. Onward transmission by reusable ophthalmic surgical instruments has not been reported, but the risk cannot be excluded. Use of appropriate cleaning and disinfection protocols and the ability to identify and quarantine instruments that may have been used on an infected patient are important safeguards.

Ultrastructural study of florid plaques in variant Creutzfeldt-Jakob disease: a comparison with amyloid plaques in kuru, sporadic Creutzfeldt-Jakob disease and Gerstmann-Sträussler-Scheinker disease.

BACKGROUND: Although the histological features of the amyloid plaques in variant Creutzfeldt-Jakob disease (vCJD) are distinct from those in other forms of prion disease [kuru, sporadic Creutzfeldt-Jakob disease (sCJD) and Gerstmann-Sträussler-Scheinker disease (GSS)], their ultrastructural features have only been described in a single case report. AIMS: To study vCJD plaques systematically and compare them with plaques in kuru, sCJD, GSS and Alzheimer disease (AD). METHODS: Amyloid plaques were studied by transmission electron microscopy and image analysis in five cases of vCJD, three cases of GSS, two cases of sCJD, one case of kuru and five cases of AD. Immunohistochemistry was performed on paraffin sections from one case of vCJD, two cases of GSS, one case of kuru and two cases of sCJD. RESULTS: The florid plaques in vCJD were either compact or more diffuse; in both forms, the radiating fibrils were organized into thick 'tongues', in contrast to kuru plaques. Dystrophic neurites (DNs) containing lysosomal electron-dense bodies or vesicles surrounded florid plaques. Microglial cells were found within florid plaques; occasional amyloid fibrils were identified in membrane-bound pockets of microglial cells. In vCJD, there was significant tau immunoreactivity in DNs around florid plaques while, in sCJD, GSS and kuru, minimal tau immunoreactivity was observed around plaques. CONCLUSIONS: The ultrastructure of the florid plaques and DNs in vCJD is more reminiscent of neuritic plaques in AD than kuru or multicentric plaques. These findings may reflect differences both in the strains of the transmissible agents responsible for these disorders and in host factors.

Sporadic Creutzfeldt-Jakob disease in two adolescents.

BACKGROUND: Sporadic Creutzfeldt-Jakob disease (CJD) is a condition predominantly affecting older age groups, with cases aged less than 45 years rare and an age at onset or death of less than 20 years exceptional. METHODS: Data from the systematic study of sporadic CJD in the UK are available from 1970 onwards. Clinical and pathological data are reviewed in order to identify atypical cases, including those at the extremes of the age range of sporadic CJD. Detailed analysis of atypical cases is undertaken, and in selected cases laboratory transmission studies are carried out in order to provide information on the characteristics of the infectious agent. RESULTS: In the UK, two cases of sporadic CJD in adolescents have been identified, dying at ages 16 and 20 years. The first case predated the epidemic of bovine spongiform encephalopathy and the characteristics of the second case, including laboratory transmission studies, are consistent with a diagnosis of sporadic rather than variant CJD. CONCLUSION: The cases in this report indicate that sporadic CJD can develop at a very young age, that variant CJD is not the only form of CJD occurring in this age group and that neuropathological examination is essential for accurate diagnosis of human prion disease.

Infective dementias.

A wide range of infectious diseases can result in dementia, although the identity and nature of these diseases has changed over time. Two of the most significant current groups in terms of scientific complexity are HIV/AIDS and prion diseases. In these disorders, dementia occurs either as a consequence of targeting the brain and selectively damaging neurones, or by an indirect effect of neuroinflammation. In prion diseases, both direct neurotoxicity and neuroinflammation may act to result in neuronal damage. In HIV encephalitis, the progression of the dementia is slower, perhaps reflecting indirect damage that appears to result from neuroinflammation as a main cause of neuronal death. An ever-increasing range of model systems is now available to study the neuronal damage in infectious dementias, ranging from cell culture systems to animal models, some of which, particularly in the case of prion diseases, are very well characterised and amenable to controlled manipulation in terms of both host and agent parameters. As valuable as these experimental models are, they do not allow a direct approach to an understanding of dementia, the complexities of which cannot readily be studied in vitro or in animal models, but they do allow studies of interventions and therapeutic strategies. This review summarises the current state of knowledge regarding the major infective dementias.