Surveillance For Creutzfeldt-Jakob Disease In China And The Studies For Two Genetic Human Prion Diseases

Transmissible spongiform encephalopathies (TSEs) are rare degenerative neurological disorders that afflict human beings (Creutzfeldt-Jakob disease, CJD; Gerstmann-Straussler-Scheinker syndrome, GSS; Kuru; and fatal familial insomnia, FFI), sheep and goat (scrapie), cattle (bovine spongiform encephalopathy), elk (Chronic wasting disease, CWD), mink (Transmissible Mink encephalopathies, TME), Feline(Feline Spongiform encephalopathies, FSE) and other animals. Creutzfeldt-Jakob disease may have a sporadic, inherited or transmissible origin. This study contains three individual parts including the surveillance for Creutzfeldt-Jakob disease in China from 2006 to 2007, the pathological and pathogenic characteristics in various brain regions from a Chinese patient with G114V genetic CJD and mouse-adapted scrapie strains 139A and ME7 overcome species barrier to induce experimental scrapie in hamsters and changed their pathogenic features. Part I:Surveillance for Creutzfeldt-Jakob disease in China from 2006 to 2007 and two genetic CJD casesIn China, the CJD surveillance system was established under the framework of the surveillances for communicable diseases led by Chinese Center for Disease Control and Prevention (CCDC) since 2002 and became broader in the past sereval years. The paper collects the surveillance data from 2006 to 2007. The highest incidences were in the group of 60-69 year-old in both probable and possible sCJD cases. The male to female ratio was 35:16 in probable sCJD and 20:10 in possible sCJD. The resident places of probable and possible sCJD cases were diffused in 23 different provinces in China. The occupations of probable and possible sCJD cases were also various, including workers, farmers, teachers, officials, self-employees and house-wives. Additionally, the suspected CJD cases were reported around all year, without seasonal specificity.The familiar foremost symptoms of the reported sCJD cases was progressive dementia and followed by mental-related syndromes, cerebellum-related syndromes, pyramidal or extrapyramidal disfunction and slow progressive dementia and so on. The most common clinical symptom was pyramidal or extrapyramidal disfunction, followed by myoclonus, visual or cerebellar disturbance, akinetic mutism. The appearance of common symptoms between probable and possible sCJD had no statistical difference. The distributing frequency of the main clinical symptoms between probable and possible sCJD were quite similar, without statistical difference.Comparative analyses of the clinical manifestations showed that the patients with both 14-3-3 positive and EEG change seem to have more clinical symptoms. By statistical analyses, there was no difference in visual or cerebellar disturbance, pyramidal or extrapyramidal disfunction and akinetic mutism symptoms, but significant difference in myoclonus symptom among the three groups. Analyses of PRNP revealed again that most cases (131,97.0%) were methionine homozygous genotype at codon 129, four (3.0%) were methionine/valine heterozygosity and none was valine/valine homozygous.Under the framework of the China CJD surveillance system, we found a Chinese female with G114V mutation suffering from progressive dementia, tiredness, lethargy and mild difficulty in falling asleep for two years. EEG and MRI findings more like as sporadic CJD. Obviously PK-resistant PrP-specific signals were detected in midbrain, thalamus, cerebellum, frontal lobe, temporal lobe, parietal lobe and occipital lobe with predominance of monoglycosylated PrPSc which was similar to that of PrPSc typeⅠ. Severe and extensive vacuolization was frequently identified in the tested tissues and PrPSc immunoblots were widely distributed in the tested tissues. Information on the pedigree was collected by interviews with family members. One of them was hospitalized for progressive memory impairment at the age of 32-year-old. During hospitalization, he showed mild ataxia of limbs and tremor. Neurological examination showed capability decline of memory, calculation and comprehension, and left limb Babinski sign positive. He is still alive.In addition, we reported a 58 year-old Chinese female with mutation in codon 188 (T188K) of the PRNP gene, while the codon 129 was a methionine homozygous genotype. The patient displayed 4-year long slowly progressive sleeping disturbance and rapid exacerbation of neurological status after other neurological manifestations appeared. Cerebral spinal fluid (CSF) 14-3-3 protein was positive.Part II:The pathological and pathogenic characteristics in various brain regions from a Chinese patient with G114V genetic CJDPreviously we have reported a Chinese gCJD case with a substitution of valine (V) for glycine (G) at codon 114. To investigate the detailed neuropathological and pathogenic characteristics of G114V gCJD, ten different brain regions were thoroughly analyzed. Neuropathological tests revealed the typical vacuolations in brains, far more intense and severe in the cortex regions, characterized with larger vacuoles that often fused each other. Widespread granular PrPSc positive-signals were observed in the sections of the brain cortex regions, while only tiny PrPSc deposits in the area of cerebellum. Abundant GFAP-positive stained cells and signals were observed in all tested regions while the profiles of GFAP-staining in various regions seemed to be diversity.In line with the observations of total PrP, intensive PrPres signals were seen in all four cortex regions, while much weaker signals in midbrain, thalamus and, cerebellum. Analyses of the glycosylating profiles of total PrP in various brain regions revealed a predominance of diglycosylated PrP in the regions without PrPres or with mild deposit of PrPres and a predominance of monoglycosylated PrP in the regions with severe deposit of PrPres.Assays of the transcriptions of PrP specific mRNA by RT-PCR and real-time PCR showed comparable levels in ten brain regions and lightly higher in the four cortex areas. Using the comparative Ct formula 2 (ΔCt1 [minus]ΔCt2), the relative quantitative difference of wild-type and mutant PRNP mRNA was 22.78 times at this experimental condition. The values of the relative amounts of the mutated and wild-type PRNP cDNA were 1.00E+00 and 6.86E+00 respectively.Two-dimensional immunoblots showed that the large portions of the dots of the diglycosylated and monoglycosylated PrPSc appeared in the acidic region of the pH gradient and some in the neutral area, whereas the dots of the aglycosylated PrPSc mostly presented in the neutral and alkaline region of the pH gradient. This data suggests that the PrPSc in brains is a mixture of the molecules with different biochemical property, varying each other not only in glycosylating profiles and molecular masses, but also in the protein electric charges.Part III:Mouse-adapted scrapie strains 139A and ME7 overcome species barrier to induce experimental scrapie in hamsters and changed their pathogenic features To understand the potential pathogenic alterations, two mouse-adapted strains 139A and ME7 were cerebrally inoculated to hamsters. After long incubation times, animals infected with strain 139A and ME7 showed typical clinical symptoms. In line with the observation of agent 263K-infected hamsters, both SAFs and PrPSc were detected in the brain tissues earlier than the appearance of clinical symptoms.Brain samples of each strain at the terminal clinical stage identified many long, ramose and roughly 25 nm in diameter fibrils, whose surfaces were shaggy. But the lengths of SAFs in 139A-ha and ME7-ha infected hamsters seemed shorter than that of hamster-adapted scrpaie agent 236K. Typical spongiform degeneration was more intensive and severe in the cortex infected by the strains and rarely observed in cerebellum regions. Abundant large GFAP-positively stained astrocytes were detected in cortex regions, in which the gliosis in the agent ME7-ha infected brains were comparably severe as that in agent 263K-infected one, while gliosis in the agent 139A-ha infected brains was significantly mild.The PrPres from the hamster brains infected with strains 139A and ME7 showed identical electrophoretic and glycosylation profiles, in which the diglycosyl form of PrPres was the most predominant, followed by monoglycosyl and aglycosyl forms, showing similar profiles as that in the preparation of the hamsters infected with strain 263K and different with the characters of mouse-adapted scrapie strains. In addition, the PrPSc in hamsters'brains by infections of hamster-adapted agent 263K or mouse-adapted agents 139A and ME7 have similar PK-resistant features and conformational stabilities. The data conclude that mouse-adapted agent 139A and ME7 change their pathogenic characteristics during cross-species transmission on hamsters. The PrPSc formed in hamsters'brains obtain new molecular properties that show markedly hamster-specific.