[Show abstract][Hide abstract] ABSTRACT: Neurodegenerative disease is a general designation for the disorders that are progressive loss of structure or function and final death of neurons, including Alzheimer's, Parkinson's, Huntington's, prion diseases, etc. In this study, we comparatively analyzed 21 individual microarray data sets of the cortex tissues from 11 sporadic Creutzfeldt-Jakob disease (sCJD), 3 fatal familial insomnia (FFI), 3 Alzheimer's disease (AD), and 4 normal controls. After normalization, a collection of 730 differently expressed sets (DESets) were obtained by comparison of the data of three diseases with their original controls. Principal component analysis (PCA) showed a background-related distribution within the groups of FFI, AD, and normal control, but two apparently different subgroups within the group of sCJD were observed. Review of the clinical materials of 11 sCJD patients identified the difference in brain PrP(Sc) deposits between two subgroups. Hierarchical cluster analysis illustrated the relatively independent clusters of normal controls, FFIs, six sCJD cases (subgroup 1) with more PrP(Sc) deposits, respectively, while an overlapped cluster of five cases of sCJD2 (subgroup 2) with less PrP(Sc) deposits and AD patients. Despite of the presence of special gene expressions, many common features were found among those neurodegenerative diseases. The most commonly changed biological processes (BPs) were signal transduction, synaptic transmission, and neuropeptide signaling pathway. The most commonly changed pathways were MAPK signaling pathway, Parkinson's disease, and oxidative phosphorylation. Our data here provide the similarity and difference in global gene expressions among the patients with sCJD, FFI, and AD, which may help to understand the common mechanism of neurodegenerative diseases.
[Show abstract][Hide abstract] ABSTRACT: Prion diseases are irreversible progressive neurodegenerative diseases characterized in the brain by PrP(Sc) deposits, neuronal degeneration, gliosis and by cognitive, behavioral and physical impairments, leading to severe incapacity and inevitable death. Proteins of the p21-activated kinase (PAK) family are noted for roles in gene transcription, cytoskeletal dynamics, cell cycle progression and survival signaling. In the present study, we aimed to identify the potential roles of PAKs during prion infection, utilizing the brains of scrapie agent-infected hamsters. Western blots and immunohistochemical assays showed that brain levels of PAK3 and PAK1, as well as their upstream activator Rac/cdc42 and downstream substrate Raf1, were remarkably reduced at terminal stage. Double-stained immunofluorescent assay demonstrated that PAK3 was expressed mainly in neurons. Dynamic analyses of the brain samples collected at the different time points during the incubation period illustrated successive decreases of PAK3, PAK1 and Raf1, especially phosphor Raf1, which correlated well with neuron loss. Rac/cdc42 in the brain tissues increased at early stage and reached to the top at mid-late stage, but diminished at final stage. Unlike the alteration of PAKs in vivo, PAK3 and PAK1, as well as Rac/cdc42 and Raf1 in the prion-infected cell line SMB-S15 remained unchanged compared with those of its normal cell line SMB-PS. Our data here indicate that the functions of PAKs and their associated signaling pathways are seriously affected in the brains of prion disease, which appear to associate closely with the extensive neuron loss.
[Show abstract][Hide abstract] ABSTRACT: It has been described that A disintegrin and metalloproteinase (ADAM10) may involve in the physiopathology of prion diseases, but the direct molecular basis still remains unsolved. In this study, we confirmed that ADAM10 was able to cleave recombinant human prion protein in vitro. Using immunoprecipitation tests (IP) and immunofluorescent assays (IFA), reliable molecular interaction between the native cellular form of PrP (PrP(C)) and ADAM10 was observed not only in various cultured neuronal cell lines but also in brain homogenates of healthy hamsters and mice. Only mature ADAM10 (after removal of its prodomain) molecules showed the binding activity with the native PrP(C). Remarkably more prion protein (PrP)-ADAM10 complexes were detected in the membrane fraction of cultured cells. In the scrapie-infected SMB cell model, the endogenous ADAM10 levels, especially the mature ADAM10, were significantly decreased in the fraction of cell membrane. IP and IFA tests of prion-infected SMB-S15 cells confirmed no detectable PrP-ADAM10 complex in the cellular lysates and PrP-ADAM10 co-localization on the cell surface. Furthermore, we demonstrated that the levels of ADAM10 in the brain homogenates of scrapie agent 263K-infected hamsters and agent ME7-infected mice were also almost diminished at the terminal stage, showing time-dependent decreases during the incubation period. Our data here provide the solid molecular basis for the endoproteolysis of ADAM10 on PrP molecules and interaction between ADAM10 and PrP(C). Obvious loss of ADAM10 during prion infection in vitro and in vivo highlights that ADAM10 may play essential pathophysiological roles in prion replication and accumulation.
[Show abstract][Hide abstract] ABSTRACT: Pathogenic mitochondrial DNA (mtDNA) mutations leading to mitochondrial dysfunction can cause a variety of chronic diseases in central nervous system (CNS). However, the role of mtDNA mutations in sporadic Creutzfeldt-Jakob disease (sCJD) has still been unknown. In this study, we comparatively analyzed complete mtDNA sequences of 31 Chinese sCJD patients and 32 controls. Using MITOMASTER and PhyloTree, we characterized 520 variants in sCJD patients and 507 variants in control by haplogroup and allele frequencies. We classified the mtDNAs into 40 sub-haplogroups of 5 haplogroups, most of them being Asian-specific haplogroups. Haplogroup U, an European-specific haplogroups mtDNA, was found only in sCJD. The analysis to control region (CR) revealed a 31% increase in the frequency of mtDNA CR mutations in sCJD versus controls. In functional elements of the mtDNA CR, six CR mutations were in conserved sequence blocks I (CSBI) in sCJD, while only one in control (P<0.05). More mutants in transfer ribonucleic acid-Leu (tRNA-Leu) were detected in sCJD. The frequencies of two synonymous amino-acid changes, m.11467A>G, p.(=) in NADH dehydrogenase subunit 4 (ND4) and m.12372G>A, p.(=) in NADH dehydrogenase subunit 5 (ND5), in sCJD patients were higher than that of controls. Our study, for the first time, screened the variations of mtDNA of Chinese sCJD patients and identified some potential disease-related mutations for further investigations.European Journal of Human Genetics advance online publication, 26 March 2014; doi:10.1038/ejhg.2014.52.
European journal of human genetics: EJHG 03/2014; · 3.56 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Shadoo (Sho) is an N-glycosylated glycophosphatidylinositol-anchored protein that is expressed in the brain and exhibits neuroprotective properties. Recently, research has shown that a reduction of Sho levels may reflect the presence of PrP(Sc) in the brain. However, the possible mechanism by which prion infection triggers down-regulation of Sho remains unclear. In the present study, Western blot and immunohistochemical assays revealed that Sho, especially glycosylated Sho, declined markedly in the brains of five scrapie agent-infected hamsters and mice at the terminal stages. Analyses of the down-regulation of Sho levels with the emergence of PrP(Sc) C2 proteolytic fragments did not identify close association in all tested scrapie-infected models. To further investigate the mechanism of depletion of Sho in prion disease, a Sho-expressing plasmid with HA tag was introduced into a scrapie-infected cell line, SMB-S15, and its normal cell line, SMB-PS. Western blot assay revealed dramatically decreased Sho in SMB-S15 cells, especially its glycosylated form. Proteasome inhibitor MG132 reversed the decrease of nonglycosylated Sho, but had little effect on glycosylated Sho. N-acetylglucosamine transferase inhibitor tunicamycin efficiently reduced the glycosylations of Sho and PrP(C) in SMB-PS cells, while two other endoplasmic reticulum stress inducers showed clear inhibition of diglycosylated PrP(C), but did not change the expression level and profile of Sho. Furthermore, immunoprecipitation of HA-Sho illustrated ubiquitination of Sho in SMB-S15 cells, but not in SMB-PS cells. We propose that the depletions of Sho in scrapie-infected cell lines due to inhibition of glycosylation mediate protein destabilization and subsequently proteasome degradation after modification by ubiquitination.
[Show abstract][Hide abstract] ABSTRACT: Proteins of the 14-3-3 family are universal participate in multiple cellular processes. However, their exact role in the pathogenesis of prion diseases remains unclear. In this study, we proposed that human PrP was able to form molecular complex with 14-3-3β. The domains responsible for the interactions between PrP and 14-3-3β were mapped at the segments of amino acid (aa) residues 106-126 within PrP and aa 1-38 within 14-3-3β. Homology modeling revealed that the key aa residues for molecular interaction were D22 and D23 in 14-3-3β as well as K110 in PrP. Mutations in these aa residues inhibited the interaction between the two proteins in vitro. Our results also showed that recombinant PrP encouraged 14-3-3β dimer formation, whereas PrP106-126 peptide inhibited it. Recombinant 14-3-3β disaggregated the mature PrP106-126 fibrils in vitro. Moreover, the PrP-14-3-3 protein complexes were observed in the brain tissues of normal and scrapie agent 263K infected hamsters. Colocalization of PrP and 14-3-3 was seen in the cytoplasm of human neuroblastoma cell line SH-SY5Y, as well as human cervical cancer cell line HeLa transiently expressing full-length human PrP. Our current data suggest the neuroprotection of PrPC and neuron damage caused by PrPSc may be associated with their functions of 14-3-3 dimerization regulation.
The international journal of biochemistry & cell biology 11/2013; · 4.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The protein of p62/sequestosome 1 (SQSTM1), a key cargo adaptor protein involved in autophagy-lysosome degradation, exhibits inclusion bodies structure in cytoplasm and plays a protective role in some models of neurodegenerative diseases. Some PrP mutants, such as PrP-CYTO and PrP-PG14, also form cytosolic inclusion bodies and trigger neuronal apoptosis either in cultured cells or in transgenic mice. Here, we demonstrated that the cellular p62/SQSTM1 incorporated into the inclusion bodies formed by expressing the abnormal PrP mutants, PrP-CYTO and PrP-PG14, in human embryonic kidney 293 cells. Overexpression of p62/SQSTM1 efficiently relieved the cytosolic aggregations and cell apoptosis induced by the abnormal PrPs. Autophagy-lysosome inhibitors instead of proteasome inhibitor sufficiently blocked the p62/SQSTM1-mediated degradations of abnormal PrPs. Overexpression of p62/SQSTM1 did not alter the levels of light chain 3 (LC3) in the cells expressing various PrPs. However, more complexes of p62/SQSTM1 with LC3 were detected in the cells expressing the misfolded PrPs. These data imply that p62/SQSTM1 plays an important role in the homeostasis of abnormal PrPs via autophagy-lysosome-dependent way.
Medical Microbiology and Immunology 11/2013; · 3.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Glucose transporters 1 (GLUT1) and 3 (GLUT3) belong to the solute carrier family 2 (SLC2, facilitated glucose transporter) and are the two most important glucose transporters (GLUTs) in brain tissue, and between them, GLUT3 is the primary one for neurons, which is responsible for glucose uptake. To obtain insights into the possible alterations of GLUT1 and GLUT3 in transmissible spongiform encephalopathies (TSEs), the protein levels of GLUT1 and GLUT3 in the brain tissues of agents 263K- and 139A-infected hamsters, as well as agents 139A- and ME7-infected mice, were evaluated. Western blots, immunofluorescent assay (IFA), and immunohistochemical (IHC) assays revealed that at the terminal stages of the infection, GLUT3 level in the brain tissues of scrapie-infected rodents was significantly downregulated, while GLUT1 level remained almost unchanged. The decline of GLUT3 level was closely related with prolonged incubation time. In line with these results in vivo, the GLUT3 level in a prion persistently infected cell line SMB-S15 was also lower than that of its normal cell line SMB-PS. Moreover, the level of hypoxia-inducible factor-1 alpha (HIF-1α), which positively regulated the expressions of GLUTs, was also markedly downregulated in the brains of several scrapie-infected animals. In vitro glucose uptake assays illustrated a markedly decreased 2-[N-(7-nitrobenze-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose uptake activity in SMB-S15 cells. Our data indicate that the reduction of GLUT3 is a common phenomenon in prion diseases, which occurs much earlier than the appearance of clinical symptoms. Defect in glucose uptake and metabolism of neurons, like in other neurodegenerative diseases, for example, Alzheimer's disease (AD), may be one of the essential processes in the pathogenesis of prion diseases.
[Show abstract][Hide abstract] ABSTRACT: The 14-3-3 proteins are a family of highly homologous and ubiquitously expressed isoforms that are involved in a wide variety of physiological processes. 14-3-3 have showed actively molecular interaction with PrP and positive 14-3-3 is frequently observed in the cerebrospinal fluid (CSF) samples of the patients with sporadic Creutzfeldt-Jakob disease (CJD). However, the alterations of 14-3-3 in the brain tissues of patients with prion diseases remain little addressed. To address the possible change of brain 14-3-3 during prion infection, we firstly tested the levels of 14-3-3 in the brain tissues of scrapie agent 263 K infected hamsters. Obviously decreased 14-3-3 were observed in the samples of the infected animals, showing time-dependent reduction in the incubation period, while the amounts of S-nitrosylated 14-3-3 were increased in the brains collected at the late stage. A low level of 14-3-3 was also observed in the scrapie infectious cell line SMB-S15, accompanied with up-regulated Bax and down-regulated Bcl-2. Moreover, we found that treatment of PrP106-126 on the cultured cells decreased the cellular 14-3-3 and caused translocations of cellular Bax to the membrane fractions. Knockdown of cellular 14-3-3 sensitized the cultured cells to the challenge of PrP106-126. These data illustrate that significant down-regulation of brain 14-3-3 levels during prion infection may not only be a scenario of the terminal consequence of interacting with abnormal PrP(Sc) but may also participate in the pathogenesis of neuronal damage.
[Show abstract][Hide abstract] ABSTRACT: To investigate the possible associations between mutations in TP53 and phenotypes of single-nucleotide polymorphisms (SNPs) in codon 72 (SNP72) with the expression profiles of p53 and human papillomavirus (HPV) infection, 93 pathologically diagnosed head and neck squamous cell carcinomas (HNSCCs) were included for study. Using PCR and direct sequencing, 45 TP53 mutations in 35 cases (37.6%) were confirmed out of the 93 HNSCCs. P53 immunohistochemistry (IHC) confirmed 34 (36.6%) cases with positive staining, including 22 cases with strong and 12 with weak positivity. IARC database and software analysis showed similar results that most of the mutated p53 proteins lost their normal function. Further statistical analysis found a negative correlation between p53 IHC and HPV IHC in the tissues from the group of other HNSCCs (of various sites other than the larynx) but not in the tissues from the laryngeal carcinomas. Analyses of SNP72 showed that the patients with the Arg phenotype had a significantly older age at disease onset when compared to patients with the Pro phenotype, particularly in the group of other HNSCCs. In addition, all cases with strong staining for p53 in the laryngeal carcinoma group had the Pro phenotype and all tumors with poor pathological differentiation in the group of other HNSCCs had the Pro phenotype. These data indicate that the profiles of TP53 mutations, SNP72 polymorphism, p53 IHC and HPV E6 IHC are distinct between the groups of laryngeal carcinoma and other HNSCCs.
[Show abstract][Hide abstract] ABSTRACT: The shotgun proteomic based on the approach of tandem mass tag (TMT) labeling has received increasing attention for neuroproteomics analysis and becomes an effective tool for the identification and quantification of a large number of proteins for the purpose of revealing key proteins involved in the neuronal dysfunction and an inflammatory response associated with neurodegenerative disorders. To assess the potential expression difference of proteins in cerebrospinal fluids (CSF) between Creutzfeldt-Jakob disease (CJD) and non-CJD patients, the pooled CSF samples from 39 Chinese probable sporadic CJD (sCJD) patients and from 52 non-CJD cases were comparably analyzed with the methodology of TMT labeling and RP-RP-UPLC-MS/MS. Totally, 437 possible proteins were identified in the tested CSF specimen, among them, 49 proteins with 95 % confidence interval. Differential assays showed among those 49 CSF proteins, 12 were upregulated and 13 were downregulated significantly in the sCJD compared to non-CJD. The most affected pathway of the differential expression proteins in CSF of sCJD was complement and coagulation cascade. Western blots for six selected changed proteins in the pooled CSF samples revealed the similar altering profiles in the groups of sCJD and non-CJD as proteomics. Furthermore, CSF samples from 24 CJD patients and 24 non-CJD patients were randomly selected and subjected individually into the Western blots of an increased protein (phosphoglycerate mutase 1) and a decreased one (alpha-1-antichymotrysin), which also confirmed the altering tendency of these identified proteins. Those data indicate that proteomic assay of CSF is a powerful technique not only for selection of the potential biomarkers for the development of diagnostic tool of CJD but also for supplement of useful scientific clues for understanding the CSF homeostasis during the pathogenesis of prion diseases.
[Show abstract][Hide abstract] ABSTRACT: αB-crystallin is a member of the small heat shock protein family constitutively presenting in brains at a relatively low level. To address the alteration of αB-crystallin in prion disease, the αB-crystallin levels in the brains of scrapie agent 263 K-infected hamsters were analyzed. The levels of αB-crystallin were remarkably increased in the brains of 263 K-infected hamsters, showing a time-dependent manner along with incubation time. Immunohistochemical (IHC) and immunofluorescent (IFA) assays illustrated more αB-crystallin-positive signals in the regions of the cortex and thalamus containing severe astrogliosis. Double-stained IFA verified that the αB-crystallin signals colocalized with the enlarged glial fibrillary acidic protein-positive astrocytes, but not with neuronal nuclei-positive cells. IHC and IFA of the serial brain sections of infected hamsters showed no colocalization and correlation between PrP(Sc) deposits and αB-crystallin increase. Moreover, increased αB-crystallin deposits were observed in the brain sections of parietal lobe of a sporadic Creutzfeldt-Jakob disease (sCJD) case, parietal lobe and thalamus of a G114V genetic CJD case, and thalamus of a fatal family insomnia (FFI) case, but not in a parietal lobe of FFI where only very mild astrogliosis was addressed. Additionally, the molecular interaction between αB-crystallin and PrP was only observed in the reactions of recombinant proteins purified from Escherichia coli, but not either in that of brain homogenates or in that of the cultured cell lysates expressing human PrP and αB-crystallin. Our data indicate that brain αB-crystallin is abnormally upregulated in various prion diseases, which is coincidental with astrogliosis. Direct interaction between αB-crystallin and PrP seems not to be essential during the pathogenesis of prion infection.
Journal of Molecular Neuroscience 07/2013; · 2.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Microglial activations have been described in different subtypes of human prion diseases such as sporadic Creutzfeldt-Jakob disease (CJD), variant CJD, Kuru and Gerstmann-Straussler-Scheinker disease (GSS). However, the situation of microglia in other genetic prion diseases such as fatal familial insomnia (FFI) and familial CJD remains less understood. The brain microglia was evaluated comparatively between the FFI, G114V and sCJD cases in the study.
Specific Western blots, immunohistochemical and immunofluorescent assays were used to detect the changes of microglia and ELISA tests were used for levels of inflammatory cytokines.
Western blots, immunohistochemical and immunofluorescent assays illustrated almost unchanged microglia in the temporal lobes of FFI and G114V gCJD, but obviously increased in those of sCJD. The Iba1-levels maintained comparable in six different brain regions of FFI and G114V cases, including thalamus, cingulate gyrus, frontal cortex, parietal cortex, occipital cortex and temporal cortex. ELISA tests for inflammatory cytokines revealed significantly up-regulated IL-1beta, IL-6 and TNF-alpha in the brain homogenates from sCJD, but not in those from FFI and G114V gCJD.
Data here demonstrates silent brain microglia in FFI and G114V gCJD but obviously increased in sCJD, which reflects various pathogenesis of different human prion diseases subtypes.
[Show abstract][Hide abstract] ABSTRACT: Here, we report a Chinese case of Creutzfeldt-Jakob disease (CJD) with a rare mutation in the prion protein gene (PRNP) leading to an exchange of amino acid from valine (Val) to isoleucine (I) at codon 203 (V203I). The 80-y-old male presented with sudden memory loss, rapid loss of vocabulary, inattention and slow responses, accompanied by dizziness, blurred vision and ataxia. Two weeks after admission, he exhibited tremor, myoclonus and bilateral Babinski signs. At the end of the clinical course, he developed severe akinetic mutism. The cerebrospinal fluid (CSF) was positive for 14-3-3 protein. Increased bilateral signal intensity in the frontal and parietal lobes was seen on diffusion-weighted imaging (DWI); periodic activity was recorded on an electroencephalogram (EEG). There was no family history of similar symptoms. The total clinical course was approximately two months.
[Show abstract][Hide abstract] ABSTRACT: Polo-like kinases (PLKs) consist of a family of kinases which play critical roles during multiple stages of cell cycle progression. Increase of PLK1 and decrease of PLK3 are associated with the developments and metastases of many types of human malignant tumors; however, the situations of PLKs in prion diseases are less understood. Using Western blots and immunohistochemical and immunofluorescent assays, marked increase of PLK1 and decrease of PLK3 were observed in the brains of scrapie strain 263K-infected hamsters, presenting obviously a time-dependent phenomenon along with disease progression. Similar alterations of PLKs were also detected in a scrapie infectious cell line SMB-S15. Both PLK1 and PLK3 were observed in neurons by confocal microscopy. Accompanying with the changes of PLKs in the brains of 263K-infected hamsters, Cdc25C and its phosphorylated forms (p-Cdc25C-Ser198 and p-Cdc25C-Ser216) were significantly down-regulated, whereas Cyclin B1 and PCNA were obviously up-regulated, while phospho-histone H3 remained almost unchanged. Moreover, exposure of the cytotoxic peptide PrP106-126 on the primary cultured cortical neuron cells induced similar changes of cellular PLKs and some cell cycle-related proteins, such as Cdc25C and its phosphorylated forms, phospho-histone H3. Those results illustrate obviously aberrant expressions of cell cycle regulatory proteins in the prion-infected neurons, which may lead to the cell cycle arrest at M phase. Possibly due to the ill-regulation of some key cell cycle events during prion infection, together with the fact that neurons are unable to complete mitosis, the cell cycle reentry in prion-infected neurons is definitely abortive, which may lead to neuron apoptosis and neuron degeneration.
[Show abstract][Hide abstract] ABSTRACT: Flotillins are membrane association proteins consisting of two homologous members, flotillin-1 (Flot-1) and flotillin-2 (Flot-2). They define a clathrin-independent endocytic pathway in mammal cells, which are also distinct from some other endocytosis mechanisms. The implicated cargoes of the flotillin-dependent pathway are mainly some GPI-anchored proteins, such as CD59 and Thy-1, which positionally colocalize with flotillins at the plasma membrane microdomains. To see whether flotillins are involved in the endocytosis of PrP(C), the potential molecular interaction between PrP(C) and flotillins in a neuroblastoma cell line SK-N-SH was analyzed. Co-immunoprecipitation assays did not reveal a detectable complex in the cell lysates of a normal feeding situation. After stimulation of Cu(2+), PrP(C) formed a clear complex with Flot-1, but not with Flot-2. Immunofluorescent assays illustrated that PrP(C) colocalized well with Flot-1, and the complexes of PrP(C)-Flot-1 shifted from the cell membrane to the cytoplasm along with the treatment of Cu(2+). Down-regulating the expression of Flot-1 in SK-N-SH cells by Flot-1-specific RNAi obviously abolished the Cu(2+)-stimulated endocytosis process of PrP(C). Moreover, we also found that in the cell line human embryonic kidney 293 (HEK293) without detectable PrP(C) expression, the distribution of cellular Flot-1 maintained almost unchanged during Cu(2+) treatment. Cu(2+)-induced PrP(C)-Flot-1 molecular interaction and endocytosis in HEK293 cells were obtained when expressing wild-type human PrP (PrP(PG5)), but not in the preparation expressing octarepeat-deleted PrP (PrP(PG0)). Our data here provide direct evidences for the molecular interaction and endocytosis of PrP(C) with Flot-1 in the presence of copper ions, and the octarepeat region of PrP(C) is critical for this process, which strongly indicates that the Flot-1-dependent endocytic pathway seems to mediate the endocytosis process of PrP(C) in the special situation.
[Show abstract][Hide abstract] ABSTRACT: Microglial cells are resident mononuclear phagocytes of the central nervous system (CNS). Active proliferation of microglia in the brain has been identified in neurodegenerative disorders, including some kinds of prion disease. However, the detailed regional distribution between microglia and PrP(Sc) deposition has not been presented, and investigation of fractalkine signaling which is involved in the regulation of activation of microglia in prion disease is not well documented. In this study, the disease phenomenon of microglial accumulation in the CNS was thoroughly analyzed using a scrapie-infected experimental model. Western blots of microglia-specific markers Iba1 and CD68, immunohistochemical and immunofluorescent assays demonstrated obviously activation of microglia in almost whole brain regions in the infected animals. Under the dynamic analysis on hallmarks of activation of microglia, a time-dependent increase of Iba1 and CD68 was detected, accompanied by accumulation of PrP(Sc) and progression of neurodegenerative symptoms. With serial brain sections and double staining of Iba1 and PrP(Sc), we observed that the microglia distributed around PrP(Sc) deposits in 263K-infected hamsters' brains, proposing PrP(Sc) phagocytosis. Flow cytometry assays with the single-cell suspensions prepared from the cortical region of the infected brains verified an activation of microglial population. ELISA assays of the cytokines in brain homogenates revealed significant upregulations of interleukin (IL)-1β, IL-6 and TNF-α when infected. Evaluation of fractalkine signaling in the infected hamsters' brains showed progressively downregulation of CX3CL1 during the incubation. Prion peptide PrP106-126 also disrupted fractalkine and evoked microglial activation in rat primary neuron-glia mixed cultures. Our data here demonstrate an activated status of microglia in CNS tissues of infectious prion disease, possibly through fractalkine signaling deficiency.
Journal of Molecular Neuroscience 03/2013; · 2.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: alphaB-crystallin is a member of the sHSP (Small heat shock protein) family, which plays an impor tant role in multiple neurodegeneration diseases. To give insight into the possible alternation and the role of aB-crystallin in prion disease, the alphaB-crystallin levels in the brain tissues of agent 263K-infected hamsters were evaluated. Western blots and IHC revealed that at the terminal stages of the diseases, the levels of alphaB-crystallin were increased up to 3-fold in the brain tissues of scrapie infected 263K hamsters compared with normal controls. Immunofluorescent assays revealed that the up-regulated alphaB-crystallin was mainly observed in astrocytes, but not in neurons. The co-localization between alphaB-crystallin and abnormal deposition of PrPsc in the brain tissues of the scrapie infected hamsters was not observed. The study may provide a foundation for further revealing the potential role of alphaB-crystallin in prion disease.
Bing du xue bao = Chinese journal of virology / [bian ji, Bing du xue bao bian ji wei yuan hui] 03/2013; 29(2):192-6.
[Show abstract][Hide abstract] ABSTRACT: Fatal familial insomnia (FFI) is a special subtype of genetic human prion diseases that is caused by the D178N mutation of the prion protein gene (PRNP). According to the surveillance data from 2006, FFI accounts for about half of all genetic prion disease cases in China. In this study, global expression patterns of the thalamus and parietal cortex from three patients with FFI were analyzed by Affymetrix Human Genome U133+ 2.0 chip. A total of 1,314 genes in the thalamus and 332 ones in the parietal lobe were determined to be differentially expressed genes (DEGs). The percentage of upregulated DEGs is much less in the thalamus (19.3 %) than that in the parietal lobe (42.8 %). Moreover, 255 of those DEGs showed the same altering tendencies in both tested regions, including 99 upregulated and 156 downregulated ones. The reliability of the results was confirmed by the real-time RT-PCR assays. There were 1,152 and 531 biological processes affected in the thalamus and the parietal lobe, respectively, as well as 391 overlapping ones in both regions. The most significantly changed molecular functions included transcription and DNA-dependent regulation of transcription, RNA splicing, mitochondrial electron transport, etc. The changed functions in the thalamus contained more numbers of DEGs than parietal lobe. According to KEGG classification, there were 167 and 115 different pathways changed in the thalamus and the parietal lobe, respectively, while 102 were changed in both. Interestingly, the top three changed pathways in the three groups mentioned above were Parkinson's disease, Alzheimer's disease, and oxidative phosphorylation. These results demonstrate the greater damage in the thalamus than in the parietal lobe during FFI pathogenesis, which is consistent with previous pathological observations. This study aims to describe the global expression profiles in various brain regions of FFI while proposing useful clues for understanding the pathogenesis of FFI and selecting potential biomarkers for diagnostic and therapeutic tools.
[Show abstract][Hide abstract] ABSTRACT: To analyze the infection of human papillomavirus in laryngeal squamous cell carcinoma patients.
The pathological samples of 64 clinical diagnosed laryngeal squamous cell carcinoma patients were collected. Lunimex and PCR techniques were used to detect the HPV gene infection and immunohistochemistry method was used to analyze the HPV protein expression in the samples.
In the 64 cases, 7 were positive for HPV infection by Luminex and PCR tests. 18 were positive for HPV16/18 E6 protein expression. The total positive rate was about 39. 1%.
The high HPV infection rate in laryngeal squamous cell carcinoma patients in the study indicated indirectly that the importance of the HPV infection in pathogenesis of the laryngeal squamous cell carcinoma.
Zhonghua shi yan he lin chuang bing du xue za zhi = Zhonghua shiyan he linchuang bingduxue zazhi = Chinese journal of experimental and clinical virology 02/2013; 27(1):22-4.