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Serum amyloid A protein in amyloidosis, rheumatic, and neoplastic diseases  

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Serum levels of amyloid protein A (SAA) have been shown to be elevated in different types of amyloidosis and in rheumatic diseases by radioimmunoassay using 125 iodine labeled AA and anti-AA. SAA levels were elevated in both primary and secondary amyloidosis, but there were highly significant differences between these levels. In heredofamilial amyloid, SAA levels were within normal limits. While the mean SAA level was elevated in persons over 70 years, the fact that some persons in this age group had normal levels suggested that marked elevation after age 70 may be due to occult inflammatory or neoplastic disease. High SAA levels in patients with rheumatoid arthritis correlated, in most cases, with physician evaluation of disease activity and Westergren ESR. SAA levels in patients with systemic lupus erythematosus were lower than those in patients with rheumatoid arthritis, and most patients with degenerative joint disease had normal levels. Very high levels of SAA were found in patients with neoplastic diseases. Patients with carcinoma of the lung and bowel had much higher levels than patients with carcinoma of the breast. Determination of SAA levels may be of value in evaluating different forms of systemic amyloidosis, assessing the activity of rheumatic disease, and screening for occult inflammatory or neoplastic disease

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Identification of Human Serum Proteins Which Interact With Alzheimer`s Amyloid ?A4 Protein  

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Full Text Available Alzheimer`s amyloid ?A4 protein fused with glutathione S-transferase (GST was highly expressed using a strong prokaryotic expression system in Escherichia coli. The expressed protein had expected molecular mass on SDS-PAGE and appeared exclusively immunoreactive with antibody specific for ?A4 epitope. This recombinant protein was purified with a combination of urea solubilization and ion exchange chromatography. To identify the human serum proteins which interact with ?A4, affinity columns were prepared by immobilizing GST- ?A4 and GST respectively. Using the affinity columns and human serum, we have observed an interaction of ?A4 with serum proteins. Two proteins of Mr 45 and 15 kDa were identified on SDS-PAGE to be involved in the interaction. Our demonstration of the ability of ?A4 to interact with serum protein strongly support the notion that such an interaction may underlie with the biological function of ?A4 in vivo.

Golam Sadik

2000-01-01

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Comparison of C-Reactive Protein and Serum Amyloid A Protein in Septic Shock Patients  

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Full Text Available Septic shock is a severe inflammatory state caused by an infectious agent. Our purpose was to investigate serum amyloid A (SAA protein and C-reactive protein (CRP as inflammatory markers of septic shock patients. Here we evaluate 29 patients in postoperative period, with septic shock, in a prospective study developed in a surgical intensive care unit. All eligible patients were monitored over a 7-day period by sequential organ failure assessment (SOFA score, daily CRP, SAA, and lactate measurements. CRP and SAA strongly correlated up to the fifth day of observation but were not good predictors of mortality in septic shock.

Fábio Ely Martins Benseñor

2008-03-01

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Serum amyloid A in geese; cloning and expression of recombinant protein.  

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We defined the nucleotide-sequence of the full-length goose serum amyloid A and compared it to SAA sequences of the duck. The aim of this work was to clone and express recombinant goose SAA and to produce antibody against this protein: Total RNA was isolated from goose liver and used to synthesise first strand cDNA. The coding region of the goose SAA cDNA was amplified by PCR using primers corresponding to the appropriate conservative regions of duck SAA mRNA. The product was subcloned into pET-15b expression vector to result in a His*Tag fusion protein expression. The protein was purified by affinity chromatography. Rabbits were then immunized against the recombinant purified goose SAA protein. The anti-SAA serum was tested by Western blotting. Full-length goose SAA mRNA sequence has been obtained and sequenced. PMID:16011987

Kovács, Beáta Marianna; Szilágyi, László; Janan, Janbaz; Rudas, Péter

2005-06-01

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Serum amyloid A protein (SAA) from mink, horse, and man: a comparative study  

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Serum amyloid A protein (SAA) was isolated from mink, horse, and human serum by ultracentrifugation and gel filtration and characterized by two-dimensional gel electrophoresis, Western blotting followed by autoradiography and N-terminal amino acid analysis. SAA was found in similar quantities in the high density lipoprotein (HDL) fraction of serum from a patient suffering from systemic juvenile rheumatoid arthritis (JRA) and mink stimulated with lipopolysaccharide (LPS), and in somewhat smaller quantities in serum from horses stimulated with Escherichia coli cultures. Only very small quantities were present in normal human controls and not detectable in normal mink and horse. Striking similarities were found between human and mink SAA with respect to molecular weight, isolectric point and degree of heterogeneity, while the molecular weight, isolectric point and degree of heterogeneity, while the molecular weight of horse SAA seemed to be somewhat lower, and no obvious heterogeneity could be demonstrated in this protein using two-dimensional gel electrophoresis. Immunologic cross-reactivity between SAA from the three species was not found. In contrast to human and horse HDL, mink HDL was found not to contain apoA-II and only minute amounts of apoC proteins. Normal horse HDL also contained additional apoproteins not present in HDL from the other species. N-terminal amino acids analysis of SAA from mink and horse demonstrated the same similarity with the corresponding AA protein as previously reported for human SAA/AA

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Medicated rat serum containing Gengnianchun decoction reduces apoptosis of pheochromocytoma cells insulted by amyloid beta protein  

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Full Text Available Objective: To investigate the effects of medicated rat serum containing Gengnianchun (GNC decoction and its protection to pheochromocytoma cells (PC12 cells from amyloid beta (A?25-35-insulted apoptosis and to find the possible mechanism.Methods: Medicated rat serum was prepared by administering ovariectomized Sprague-Dawley (SD rats with GNC decoction. The effects of medicated rat serum on viability of PC12 cells were evaluated by cell counting kit-8 (CCK-8 assay. The PC12 cells were cultured with different doses of A?25-35 to induce a model of Alzheimer's disease in vitro. Then, the protective effects of medicated rat serum on A?25-35-insulted PC12 cells were evaluated by using CCK-8 assay to detect the cell viability, using Annexin ?-fluorescein isothiocyanate (FITC/propidium iodide (PI flow cytometry to detect cell apoptosis rate and using Western blotting assay to analyze the expressions of Bcl-2, Bax and active caspase-3 proteins.Results: PC12 cells cultured with 20% medicated rat serum containing GNC decoction for 24 h or 48 h had higher viability than those cultured with normal culture medium (P<0.05. After 24- or 48-hour treatment of different concentrations of A?25-35, cell viabilities were all decreased as compared with normal medium (P<0.05. Cells underwent apoptosis, which showed the neurotoxicity of A?25-35. The cell apoptosis induced by A?25-35 was significantly decreased in PC12 cells which were pretreated with 20% medicated rat serum or nerve growth factor (NGF according to CCK-8 assay and Annexin ?-FITC/PI flow cytometry (P<0.05. The ratio of Bax expression to Bcl-2 expression and the expression of active caspase-3 were decreased in the cells treated with medicated serum or NGF as compared with the cells cultured with A?25-35 only.Conclusion: The GNC-medicated rat serum at concentration of 20% can promote viability of A?25-35-insulted PC12 cells and decrease the cell apoptosis by regulating the expressions of Bcl-2, Bax and active caspase 3.

Jun LI

2010-05-01

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Isolation and characterization of pharmaceutical grade human pentraxins, serum amyloid P component and C-reactive protein, for clinical use.  

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The human pentraxin proteins, serum amyloid P component (SAP) and C-reactive protein (CRP) are important in routine clinical diagnosis, SAP for systemic amyloidosis and CRP for monitoring the non-specific acute phase response. They are also targets for novel therapies currently in development but their roles in health and disease are controversial. Thus, both for clinical use and to rigorously elucidate their functions, structurally and functionally intact, pharmaceutical grade preparations o...

Pepys, M. B.; Gallimore, J. R.; Lloyd, J.; Li, Z.; Graham, D.; Taylor, G. W.; Ellmerich, S.; Mangione, P. P.; Tennent, G. A.; Hutchinson, W. L.; Millar, D. J.; Bennett, G.; More, J.; Evans, D.; Mistry, Y.

2012-01-01

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Evaluation of Sialic Acid and Acute Phase Proteins (Haptoglobin and Serum Amyloid A) in Clinical and Subclinical Bovine Mastitis  

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The present study was conducted to evaluate the concentrations of sialic acids (total, lipid bound and protein bound) and their correlation with acute phase proteins (haptoglobin and serum amyloid A) in clinical and subclinical mastitis of cattle. Thirty subclinical mastitic cows with positive California mastitis test (CMT) test and no clinical signs of mastitis, 10 clinical mastitic cows and 10 healthy cows with negative CMT test and normal somatic cell count were selected. Milk and blood sa...

S Nazifi, M. Haghkhah

2011-01-01

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Developing column material for the separation of serum amyloid P and C reactive protein from biological sources.  

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In this study, we have investigated the isolation of serum amyloid P (SAP) and C-reactive protein (CRP) from rainbow trout. It has recently been found that SAP is deposited in atherosclerotic lesions or neurofibrillary tangles, which are related to aging process and Alzheimer's disease. Given the importance of CRP, the CRP level in blood is becoming recognized as a potential means of monitoring cardiovascular risk. These two proteins, members of the pentraxin family of oligomeric serum proteins, were isolated from rainbow trout using N-methacryloyl-phosphoserine (MA-pSer) immobilized poly (2-hydroxy ethylmethacrylate) (PHEMA) cryogels as a column material in a fast protein liquid chromatography system. The separation process was verified in two steps. First, SAP and CRP proteins were isolated together from serum sample of rainbow trout using MA-pSer/PHEMA cryogel columns. Second, SAP protein was separated chromatographically from CRP protein using the Ca(2+) ion immobilized PHEMA cryogel column. According to the data, a new and effective technique has been developed for the isolation of SAP and CRP proteins from a biological source, rainbow trout. Finally, purified SAP and CRP were loaded using sodium dodecyl sulfate-polyacrylamide gel and western blot analysis to investigate the purity of chromatographically isolated SAP and CRP compared with commertial SAP and CRP. PMID:24827758

Ersöz, Arzu; Ünlüer, Özlem Biçen; Dönmez, Gülnur; Hür, Deniz; Say, R Dvan

2014-10-01

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Evaluation of Sialic Acid and Acute Phase Proteins (Haptoglobin and Serum Amyloid A in Clinical and Subclinical Bovine Mastitis  

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Full Text Available The present study was conducted to evaluate the concentrations of sialic acids (total, lipid bound and protein bound and their correlation with acute phase proteins (haptoglobin and serum amyloid A in clinical and subclinical mastitis of cattle. Thirty subclinical mastitic cows with positive California mastitis test (CMT test and no clinical signs of mastitis, 10 clinical mastitic cows and 10 healthy cows with negative CMT test and normal somatic cell count were selected. Milk and blood samples were collected after confirmation of clinical and subclinical mastitis by somatic cell count and bacterial identification. Serum haptoglobin (Hp, serum amyloid A (SAA, total sialic acid (TSA, lipid bound sialic acid (LBSA and protein bound sialic acid (PBSA were measured by validated standard methods. Haptoglobin and SAA increased significantly in both types of mastitis compared with control group (P<0.001. However, the ratio of HP/SAA was significantly different from the control group only in clinical mastitis. The results showed that TSA and LBSA were significantly different in control group compared with clinical and subclinical mastitis (P<0.001. Protein bound sialic acid did not change in subclinical mastitis in comparison with control group (P=0.86. There was positive correlation between LBSA and PBSA in clinical mastitis (r=0.72, P=0.02 whereas significant negative correlation was observed between LBSA and PBSA in subclinical mastitis (r=-0.62, P<0.001. Results also showed no correlation between Hp and SAA with each other or with any other parameters in study groups.

S. Nazifi*, M. Haghkhah1, Z. Asadi, M. Ansari-Lari2, M. R. Tabandeh3, Z. Esmailnezhad and M. Aghamiri

2011-01-01

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SERUM ANALYSIS OF AMYLOID BETA-PROTEIN 1-40 IN HEALTHY SUBJECTS, AUTISTIC CHILDREN AND ALZHEIMER’S PATIENTS  

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Full Text Available Amyloid beta-protein1-40 (AP40 is a low molecu­lar weight peptide produced throughout life during normal cell metabolism and neurodegenerative diseases. Owing to its neurotrophic and neurotoxic effects, the present study was conducted to evalu­ate serum levels of AP40 in healthy subjects, au­tistic children and Alzheimer’s disease patients. Serum AP40 was measured by enzyme-linked im­munosorbent assay (ELISA. AP40 was signifi­cantly higher in normal children compared to nor­mal older controls, in normal children compared to autistic children, and in autistic children compared to Alzheimer’s patients (p value was less than 0.05 for all groups. This finding suggests an age-re­lated decline of serum AP40 in normal aging, as well as in autism and Alzheimer’s disease. This decline may result from abnormal processing of amyloid beta-protein precursor (APP during nor­mal aging and age-related diseases such as autism in children and Alzheimer’s disease in elderly. Possible explanations for this decline may include age-related increased interactions of AP40 with cytoskeletal proteins for brain tissue deposition, increased serine proteases for APP metabolism or hyperimmune reaction (antibodies to AP40 for removal of circulating AP40. To conclude, the AP40 metabolism declines with normal aging and in addition to its role in Alzheimer’s disease this protein might also be a contributing factor in au­tism.

Vijendra K. SINGH

2008-06-01

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Relationship between urinary sialylated saccharides, serum amyloid A protein, and C-reactive protein in rheumatoid arthritis and systemic lupus erythematosus.  

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The urinary excretion of sialic-acid-containing oligosaccharides, total sialic acid, serum amyloid A protein (SAA), and C-reactive protein (CRP) has been studied in 48 patients with rheumatoid arthritis (RA) and in 17 patients with systemic lupus erythematosus (SLE). Linear regression analysis revealed a close positive correlation between serum SAA and CRP levels in both RA (r = 0.71, p less than 0.001) and SLE (r = 0.86, p less than 0.001). The urinary excretion of sialyl lactose showed a po...

Maury, C. P.; Teppo, A. M.; Wegelius, O.

1982-01-01

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Nanophotonics of protein amyloids  

Science.gov (United States)

Technological breakthroughs in the super-resolution optical imaging techniques have enriched our current understanding of a range of biological systems and biomolecular processes at the nanoscopic spatial resolution. Protein amyloids are an important class of ordered protein assemblies consisting of misfolded proteins that are implicated in a wide range of devastating human diseases. In order to decipher the structural basis of the supramolecular protein assembly in amyloids and their detrimental interactions with the cell membranes, it is important to employ high-resolution optical imaging techniques. Additionally, amyloids could serve as novel biological nanomaterials for a variety of potential applications. In this review, we summarize a few examples of the utility of near-field scanning optical imaging methodologies to obtain a wealth of structural information into the nanoscale amyloid assembly. Although the near-field technologies were developed several decades ago, it is only recently that these methodologies are being applied and adapted for amyloid research to yield novel information pertaining to the exciting nanoscopic world of protein aggregates. We believe that the account on the nanophotonics of amyloids described in this review will be useful for the future studies on the biophysics of amyloids.

Bhattacharya, Mily; Mukhopadhyay, Samrat

2014-04-01

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Comparison of serum amyloid A and C-reactive protein as diagnostic markers of systemic inflammation in dogs  

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The diagnostic performance of canine serum amyloid A (SAA) was compared with that of C-reactive protein (CRP) in the detection of systemic inflammation in dogs. Sera from 500 dogs were retrospectively included in the study. C-reactive protein and SAA were measured using validated automated assays. The overlap performance, clinical decision limits, overall diagnostic performance, correlations, and agreement in the clinical classification between these 2 diagnostic markers were compared. Significantly higher concentrations of both proteins were detected in dogs with systemic inflammation (SAA range: 48.75 to > 2700 mg/L; CRP range: 0.4 to 907.4 mg/L) compared to dogs without systemic inflammation (SAA range: 1.06 to 56.4 mg/L; CRP range: 0.07 to 24.7 mg/L). Both proteins were shown to be sensitive and specific markers of systemic inflammation in dogs. Significant correlations and excellent diagnostic agreement were observed between the 2 markers. However, SAA showed a wider range of concentrations and a significantly superior overall diagnostic performance compared with CRP.

Christensen, Michelle B; Langhorn, Rebecca

2014-01-01

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Antibodies to human serum amyloid P component eliminate visceral amyloid deposits.  

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Accumulation of amyloid fibrils in the viscera and connective tissues causes systemic amyloidosis, which is responsible for about one in a thousand deaths in developed countries. Localized amyloid can also have serious consequences; for example, cerebral amyloid angiopathy is an important cause of haemorrhagic stroke. The clinical presentations of amyloidosis are extremely diverse and the diagnosis is rarely made before significant organ damage is present. There is therefore a major unmet need for therapy that safely promotes the clearance of established amyloid deposits. Over 20 different amyloid fibril proteins are responsible for different forms of clinically significant amyloidosis and treatments that substantially reduce the abundance of the respective amyloid fibril precursor proteins can arrest amyloid accumulation. Unfortunately, control of fibril-protein production is not possible in some forms of amyloidosis and in others it is often slow and hazardous. There is no therapy that directly targets amyloid deposits for enhanced clearance. However, all amyloid deposits contain the normal, non-fibrillar plasma glycoprotein, serum amyloid P component (SAP). Here we show that administration of anti-human-SAP antibodies to mice with amyloid deposits containing human SAP triggers a potent, complement-dependent, macrophage-derived giant cell reaction that swiftly removes massive visceral amyloid deposits without adverse effects. Anti-SAP-antibody treatment is clinically feasible because circulating human SAP can be depleted in patients by the bis-d-proline compound CPHPC, thereby enabling injected anti-SAP antibodies to reach residual SAP in the amyloid deposits. The unprecedented capacity of this novel combined therapy to eliminate amyloid deposits should be applicable to all forms of systemic and local amyloidosis. PMID:20962779

Bodin, Karl; Ellmerich, Stephan; Kahan, Melvyn C; Tennent, Glenys A; Loesch, Andrzej; Gilbertson, Janet A; Hutchinson, Winston L; Mangione, Palma P; Gallimore, J Ruth; Millar, David J; Minogue, Shane; Dhillon, Amar P; Taylor, Graham W; Bradwell, Arthur R; Petrie, Aviva; Gillmore, Julian D; Bellotti, Vittorio; Botto, Marina; Hawkins, Philip N; Pepys, Mark B

2010-11-01

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The diagnostic role of human epididymis protein 4 and serum amyloid-A in early-stage endometrial cancer patients.  

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The aim of this study was to evaluate the prognostic and predictive efficacy of the human epididymis secretory protein 4 (HE4) and serum amyloid-A (S-AA) together with the other tumor markers (CA 125, CA 15-3, CEA, and CA 19-9) in endometrial cancer patients. The study group consisted of 64 patients with defined stage and grade of endometrial cancer and 60 women with benign uterine diseases. Thirty-four healthy women were defined as the control group. Fasting blood samples were collected prior to surgery and tumor marker levels were determined in blood samples by E170 autoanalyzer. S-AA concentrations were measured by particle-enhanced immunonephelometry. Preoperative serum HE4 and S-AA levels were significantly higher in endometrial cancer patients than in controls, whereas the other measured parameters were not significantly different. Serum levels of HE4 were related to both the stage and grade of tumor. The best cutoff point for HE4 was determined to be 59.7 pmol/L; with 75 % sensitivity and 65.5 % specificity. For S-AA, the cutoff point was 8.8 U/mL, with 68.7 % sensitivity and 58.6 % specificity. The combination of HE4, CA 125, CEA, and S-AA raised the sensitivity to 84 %. Preoperative measurement of serum HE4 and S-AA may be of help in early detection of endometrial cancer. Preoperative screening with these markers may provide important information about the patient's outcome and prognosis. PMID:23640061

Omer, Beyhan; Genc, Sema; Takmaz, Ozguc; Dirican, Ahmet; Kusku-Kiraz, Zeynep; Berkman, Sinan; Gurdol, Figen

2013-10-01

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Diagnostic utility and limitations of glutamine synthetase and serum amyloid-associated protein immunohistochemistry in the distinction of focal nodular hyperplasia and inflammatory hepatocellular adenoma.  

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Inflammatory hepatocellular adenoma can show overlapping histological features with focal nodular hyperplasia, including inflammation, fibrous stroma, and ductular reaction. Expression of serum amyloid-associated protein in inflammatory hepatocellular adenoma and map-like pattern of glutamine synthetase in focal nodular hyperplasia can be helpful in this distinction, but the pitfalls and limitations of these markers have not been established. Morphology and immunohistochemistry were analyzed in 54 inflammatory hepatocellular adenomas, 40 focal nodular hyperplasia, and 3 indeterminate lesions. Morphological analysis demonstrated that nodularity, fibrous stroma, dystrophic blood vessels, and ductular reaction were more common in focal nodular hyperplasia, while telangiectasia, hemorrhage, and steatosis were more common in inflammatory hepatocellular adenoma, but there was frequent overlap of morphological features. The majority of inflammatory hepatocellular adenomas demonstrated perivascular and/or patchy glutamine synthetase staining (73.6%), while the remaining cases had diffuse (7.5%), negative (3.8%), or patchy pattern of staining (15%) that showed subtle differences from the classic map-like staining pattern and was designated as pseudo map-like staining. Positive staining for serum amyloid-associated protein was seen in the majority of inflammatory hepatocellular adenomas (92.6%) and in the minority of focal nodular hyperplasia (17.5%). The glutamine synthetase staining pattern was map-like in 90% of focal nodular hyperplasia cases, with the remaining 10% of cases showing pseudo map-like staining. Three cases were labeled as indeterminate and showed focal nodular hyperplasia-like morphology but lacked map-like glutamine synthetase staining pattern; these cases demonstrated a patchy pseudo map-like glutamine synthetase pattern along with the expression of serum amyloid-associated protein. Our results highlight the diagnostic errors that can be caused by variant patterns of staining with glutamine synthetase and serum amyloid-associated protein in inflammatory hepatocellular adenoma and focal nodular hyperplasia. PMID:23807780

Joseph, Nancy M; Ferrell, Linda D; Jain, Dhanpat; Torbenson, Michael S; Wu, Tsung-Teh; Yeh, Matthew M; Kakar, Sanjay

2014-01-01

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Human serum amyloid genes--molecular characterization  

International Nuclear Information System (INIS)

Three clones containing human genes for serum amyloid A protein (SAA) have been isolated and characterized. Each of two clones, GSAA 1 and 2 (of 12.8 and 15.9 kilobases, respectively), contains two exons, accouting for amino acids 12-58 and 58-103 of mature SAA; the extreme 5' termini and 5' untranslated regions have not yet been defined but are anticipated to be close based on studies of murine SAA genes. Initial amino acid sequence comparisons show 78/89 identical residues. At 4 of the 11 discrepant residues, the amino acid specified by the codon is the same as the corresponding residue in murine SAA. Identification of regions containing coding regions has permitted use of selected subclones for blot hybridization studies of larger human SAA chromosomal gene organization. The third clone, GSAA 3 also contains SAA coding information by DNA sequence analysis but has a different organization which has not yet been fully described. We have reported the isolation of clones of human DNA hybridizing with pRS48 - a plasmid containing a complementary DNA (cDNA) clone for murine serum amyloid A (SAA; 1, 2). We now present more detailed data confirming the identity and defining some of the organizational features of these clones

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Serum amyloid a in clinical practice  

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Full Text Available Serum amyloid A (SAA is an acute phase first class protein discovered a quarter of the century ago. Its concentration depends on clinical findings of the patient, illness activity and the therapy applied. SAA increases moderately to markedly (100-1000 mg/l in bacterial and fungal infections, invasive malignant diseases, tissue injuries in the acute myocardial infarction and autoimmune diseases such as rheumatoid arthritis and vasculitis. Mild elevation (10-100 mg/l is often seen in viral infections, systemic lupus erythematosus and localized inflammation or tissue injuries in cystitis and cerebral infarction. SAA as sensitive, non-invasive parameter is used in organ transplantation where early and correct diagnosis is needed as well as where prompt therapy is required. Besides acute kidney allograft rejection, SAA is used in the diagnosis of rejection after liver transplantation, simultaneous pancreas and kidney transplantation and also in bone marrow transplantation (acute „graft vs. host disease". Simultaneous determination of C-reactive protein (CRP and SAA may point to acute kidney allograft rejection. Standard immunosuppressive therapy with cyclosporine A and prednisolone significantly suppresses the acute phase CRP reaction both in operation itself and acute rejection, but not in infection. On the other hand, SAA rejection in operation, acute allograft rejection and infection is present in spite of cyclosporine A and steroids therapy. Different reaction of SAA and CRP in transplant patients to cyclosporine A therapy helps in differentiation between the infection and rejection. Although CRP and SAA are sensitive and acute phase reactants, their serum concentrations cannot be valued as prognostic and diagnostic criteria without creatinine serum concentration and clinical findings. In addition, they offer important information for clinical diagnosis as well as the kind of therapy.

Jovanovi? Dijana B.

2004-01-01

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Kinetics of human serum amyloid A  

International Nuclear Information System (INIS)

In order to better understand the pathogenetic role of serum amyloid A (SAA) we studied the kinetics of 131I radiolabelled pure SAA, extracted from 400 ml serum of a human volunteer. 50 microCi of 131I SAA and 15 microCi 125I labelled sodium iodide were administered i.v. on two occasions at 6 month intervals. Serum and plasma samples were collected at 10-20 min intervals x 10, then once daily x 10; lymphocytes were separated from monocytes and granulocytes. Counts per minute of 131I and 125I were measured in each sample in the serum, in serum precipitates resulting after addition of a rabbit anti-SAA antibody and of TCA and in various cell subpopulations as well as in the whole urine and TCA precipitated urine from each micturition. The 131I disappearance curves from the plasma and serum precipitates were semilogarithmically plotted; cumulative 131I cpm in plasma, cells and urine at various intervals were determined. Body scanning was performed at 2, 16, and 48 h. The results of the two experiments were very similar. The curve of 131I SAA in plasma TCA precipitates indicated the existence of 4 compartments likely due to uptake of 131I SAA by some plasma proteins, circulating cells and other tissues; later release from tissues started at 6 h. The 131I SAA half-life time in these compartments was found to be 35, 170, 255, and 550 min, respectively. Tissue bindin, and 550 min, respectively. Tissue binding of 131I was also suggested by a rising of the 125I:131I ratio with time and by a 26% release of 131I in the urine at 15 h which could not account for its plasma disappearance. Scanning, except for 131I uptake in the spleen at 2 h likely due to blood activity, showed no organ concentration. 92% of the injected 131I was found in the urine but only 6.2% of 131I SAA was accounted for in urine precicipitates

 
 
 
 
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Serum amyloid P component scintigraphy in familial amyloid polyneuropathy: regression of visceral amyloid following liver transplantation  

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Familial amyloid polyneuropathy (FAP) associated with transthyretin (TTR) mutations is the commonest type of hereditary amyloidosis. Plasma TTR is produced almost exclusively in the liver and orthotopic liver transplantation is the only available treatment, although the clinical outcome varies. Serum amyloid P component (SAP) scintigraphy is a method for identifying and quantitatively monitoring amyloid deposits in vivo, but it has not previously been used to study the outcome of visceral amyloid deposits in FAP following liver transplantation. Whole body scintigraphy following injection of iodine-123 labelled SAP was performed in 17 patients with FAP associated with TTR Met30 and in five asymptomatic gene carriers. Follow-up studies were performed in ten patients, eight of whom had undergone orthotopic liver transplantation 1-5 years beforehand. There was abnormal uptake of {sup 123}I-SAP in all FAP patients, including the kidneys in each case, the spleen in five cases and the adrenal glands in three cases. Renal amyloid deposits were also present in three of the asymptomatic carriers. Follow-up studies 1-5 years after liver transplantation showed that there had been substantial regression of the visceral amyloid deposits in two patients and modest improvement in three cases. The amyloid deposits were unchanged in two patients. In conclusion, {sup 123}I-SAP scintigraphy identified unsuspected visceral amyloid in each patient with FAP due to TTR Met30. The universal presence of renal amyloid probably underlies the high frequency of renal failure that occurs in FAP following liver transplantation. The variable capacity of patients to mobilise amyloid deposits following liver transplantation may contribute to their long-term clinical outcome. (orig.) With 2 figs., 2 tabs., 22 refs.

Rydh, A.; Hietala, S.O.; Aahlstroem, K.R. [Department of Diagnostic Radiology, University Hospital of Northern Sweden, Umeaa (Sweden); Suhr, O. [Department of Internal Medicine, University Hospital of Northern Sweden, Umeaa (Sweden); Pepys, M.B.; Hawkins, P.N. [Immunological Medicine Unit, Department of Medicine, Imperial College School of Medicine, London (United Kingdom)

1998-07-01

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Serum amyloid P component scintigraphy in familial amyloid polyneuropathy: regression of visceral amyloid following liver transplantation  

International Nuclear Information System (INIS)

Familial amyloid polyneuropathy (FAP) associated with transthyretin (TTR) mutations is the commonest type of hereditary amyloidosis. Plasma TTR is produced almost exclusively in the liver and orthotopic liver transplantation is the only available treatment, although the clinical outcome varies. Serum amyloid P component (SAP) scintigraphy is a method for identifying and quantitatively monitoring amyloid deposits in vivo, but it has not previously been used to study the outcome of visceral amyloid deposits in FAP following liver transplantation. Whole body scintigraphy following injection of iodine-123 labelled SAP was performed in 17 patients with FAP associated with TTR Met30 and in five asymptomatic gene carriers. Follow-up studies were performed in ten patients, eight of whom had undergone orthotopic liver transplantation 1-5 years beforehand. There was abnormal uptake of 123I-SAP in all FAP patients, including the kidneys in each case, the spleen in five cases and the adrenal glands in three cases. Renal amyloid deposits were also present in three of the asymptomatic carriers. Follow-up studies 1-5 years after liver transplantation showed that there had been substantial regression of the visceral amyloid deposits in two patients and modest improvement in three cases. The amyloid deposits were unchanged in two patients. In conclusion, 123I-SAP scintigraphy identified unsuspected visceral amyloid in each patient with FAP due to TTR Met30. The uh patient with FAP due to TTR Met30. The universal presence of renal amyloid probably underlies the high frequency of renal failure that occurs in FAP following liver transplantation. The variable capacity of patients to mobilise amyloid deposits following liver transplantation may contribute to their long-term clinical outcome. (orig.)

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Binding of complement proteins C1q and C4bp to serum amyloid P component (SAP) in solid contra liquid phase.  

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Serum amyloid P component (SAP), a member of the conserved pentraxin family of plasma proteins, binds calcium dependently to its ligands. The authors investigated SAPs interaction with the complement proteins C4b binding protein (C4bp) and C1q by ELISA, immunoelectrophoresis and electron microscopy. Binding of these proteins to SAP was demonstrated when SAP was immobilized using F(ab')2 anti-SAP, but not when SAP reacted with these proteins in liquid phase; thus the binding to human SAP was markedly phase state dependent. Presaturation of solid phase SAP with heparin, which binds SAP with high affinity, did not interfere with the subsequent binding of C4bp or C1q to SAP. In contrast, collagen I and IV showed partial competition with the binding of C1q to SAP. Using fresh serum, immobilized native SAP bound C4bp whereas binding of C1q/C1 could not be demonstrated. Altogether the results indicate that firm binding of C1q and C4bp to SAP requires that SAP is presented on a solid phase, that C1q and C4bp react with sites distinct from the heparin binding site, and that C1q and collagen I share binding sites on SAP. Immobilized native SAP, aggregated SAP and SAP-heparansulphate complexes induced no detectable complement activation.

SØrensen, Inge Juul; Nielsen, EH

1996-01-01

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Native human serum amyloid P component is a single pentamer  

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Serum amyloid P component (SAP) and C-reactive protein (CRP) are members of the pentraxin protein family. SAP is the precursor protein to amyloid P component present in all forms of amyloidosis. The prevailing notion is that SAP in circulation has the form of a double pentameric molecule (decamer) whereas CRP is a single pentameric molecule. We have investigated by gel permeation chromatography the M(r) of SAP in freshly collected human serum and of SAP purified by carbohydrate affinity chromatography and anion exchange chromatography. SAP was monitored by quantitative immunoelectrophoresis and ELISA, and SAP peak fractions were analysed by use of SDS-PAGE, Western blotting, and electron microscopy. The results indicate that native SAP circulates as a single pentamer, a part of which forms complexes with C4b-binding protein. The properties of SAP changed during purification as indicated by rocket immunoelectrophoresis and electron microscopy. Thus, electron micrographs of purified SAP showed a predominance of decamers. However, the decamer form of SAP reversed to single pentamers when purified SAP was incorporated into SAP-depleted serum.

SØrensen, Inge Juul; Andersen, Ove

1995-01-01

25

Search for Amyloid-Binding Proteins by Affinity Chromatography  

Science.gov (United States)

Amyloid binging proteins’ is a generic term used to designate proteins that interact with different forms of amyloidogenic peptides or proteins and that, as a result, may modulate their physiological and pathological functions by altering solubility, transport, clearance, degradation, and fibril formation. We describe a simple affinity chromatography protocol to isolate and characterize amyloid-binding proteins based on the use of sequential elution steps that may provide further information on the type of binding interaction. As an example, we depict the application of this protocol to the study of Alzheimer’s amyloid ? (A?) peptide-binding proteins derived from human plasma. Biochemical analysis of the proteins eluted under different conditions identified serum amyloid P component (SAP) and apolipoprotein J (clusterin) as the main plasma A?-binding proteins while various apolipoproteins (apoA-IV, apoE, and apoA-I), as well as albumin (HSA) and fibulin were identified as minor contributors. PMID:22528093

Calero, Miguel; Rostagno, Agueda; Ghiso, Jorge

2013-01-01

26

The presence of a novel protein in calf serum that recognizes beta amyloid in the formalin-fixed section.  

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Here we report on a monoclonal antibody, H6-33, that labels various beta-amyloid plaques, including diffuse plaques in the formalin-fixed, paraffin-embedded section from the brain affected with Alzheimer's disease (AD), without formic acid pretreatment. H6-33 also labels some neurofibrillary tangles and all kuru plaques in Gerstmann-Sträussler-Scheinker disease. In sharp contrast, H6-33 did not stain beta amyloid in the leptomeningeal vessel. For specific staining, H6-33 required the presenc...

Kanemaru, K.; Hasegawa, M.; Shimada, H.; Ihara, Y.

1990-01-01

27

Protein electrophoresis - serum  

Science.gov (United States)

... measures the types of protein in the fluid (serum) part of a blood sample. Other electrophoresis tests that measure proteins in the serum include: Immunoelectrophoresis Immunofixation Globulin electrophoresis

28

Amyloid Aggregation and Membrane Disruption by Amyloid Proteins  

Science.gov (United States)

Amyloidogenesis has been the focus of intense basic and clinical research, as an increasing number of amyloidogenic proteins have been linked to common and incurable degenerative diseases including Alzheimer's, type II diabetes, and Parkinson's. Recent studies suggest that the cell toxicity is mainly due to intermediates generated during the assembly process of amyloid fibers, which have been proposed to attack cells in a variety of ways. Disruption of cell membranes is believed to be one of the key components of amyloid toxicity. However, the mechanism by which this occurs is not fully understood. Our research in this area is focused on the investigation of the early events in the aggregation and membrane disruption of amyloid proteins, Islet amyloid polypeptide protein (IAPP, also known as amylin) and amyloid-beta peptide, on the molecular level. Structural insights into the mechanisms of membrane disruption by these amyloid proteins and the role of membrane components on the membrane disruption will be presented.[4pt] References:[0pt] [1] Sciacca et al., Biophys. J. 2012, 103, 702-10.[0pt] [2] Sciacca et al., Biochemistry. 2012, 51, 7676-84[0pt] [3] Brender et al., Acc. Chem. Res. 2012, 45, 454-62.[0pt] [4] Nanga et al., Biochim. Biophys. Acta 2011, 1808, 2337-42.[0pt] [5] Brender et al., Biophys J. 2011, 100, 685-92.

Ramamoorthy, Ayyalusamy

2013-03-01

29

C-reactive protein and serum amyloid A as early-phase and prognostic indicators of acute radiation exposure in nonhuman primate total-body irradiation model  

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Terrorist radiological attacks or nuclear accidents could expose large numbers of people to ionizing radiation. In mass-casualty radiological incidents early medical-management requires triage tools for first-responders to quantitatively identify individuals exposed to life-threatening radiation doses and for early initiation (i.e., within one day after radiation exposure) of cytokine therapy for treatment of bone marrow acute radiation syndrome. Herein, we present results from 30 rhesus macaques total-body irradiated (TBI) to a broad dose range of 1-8.5 Gy with {sup 60}Co {gamma}-rays (0.55 Gy min{sup -1}) and demonstrate dose- and time-dependent changes in blood of C-reactive protein (CRP), serum amyloid A (SAA), and interleukin 6 (IL-6) measured by enzyme linked immunosorbent assay (ELISA). CRP and SAA dose-response results are consistent with {approx}1 Gy and {approx}0.2 Gy thresholds for photon-exposure at 24 h after TBI, respectively. Highly significant elevations of CRP and SAA (p = 0.00017 and p = 0.0024, respectively) were found in animal plasma at 6 h after all TBI doses suggesting their potential use as early-phase biodosimeters. Results also show that the dynamics and content of CRP and SAA levels reflect the course and severity of the acute radiation sickness (ARS) and may function as prognostic indicators of ARS outcome. These results demonstrate proof-of-concept that these radiation-responsive proteins show promise as a complementary approach to conventional biodosimetry for early assessment of radiation exposures and may also contribute as diagnostic indices in the medical management of radiation accidents.

Ossetrova, N.I., E-mail: ossetrova@afrri.usuhs.mil [Armed Forces Radiobiology Research Institute, 8901 Wisconsin Avenue, Bldg. 42, Bethesda, MD 20889-5603 (United States); Sandgren, D.J.; Blakely, W.F. [Armed Forces Radiobiology Research Institute, 8901 Wisconsin Avenue, Bldg. 42, Bethesda, MD 20889-5603 (United States)

2011-09-15

30

Purification, subunit characterization and ultrastructure of three soluble bovine lectins: conglutinin, mannose-binding protein and the pentraxin serum amyloid P-component.  

DEFF Research Database (Denmark)

Conglutinin and mannose-binding protein (MBP) are members of the C-type lectins which are widely present in mammalian plasma. Serum amyloid P-component (SAP) is a member of the pentraxin family with lectin properties. A scheme for the partial purification of all three lectins by carbohydrate affinity chromatography and selective elution was developed. The purification was monitored by SDS-PAGE, Western blotting and electron microscopy. Binding of the lectins to Sephadex-iC3b, their collagenase sensitivity, and the size and antibody reactivity of their subunits was investigated. The demonstration, by SDS-PAGE, of 25-kDa subunits, which were unaffected by collagenase treatment but bound to Sephadex-iC3b and antibodies to human SAP, indicated the existence of bovine SAP. Bovine conglutinin (BK) also showed calcium-dependent binding to Sephadex-iC3b, whereas bovine MBP did not. The binding of BK was inhibitable with GlcNAc. A 3000-fold increase in BK activity (ELISA) was obtained in eluates from Sephadex-iC3b. SDS-PAGE analyses of BK and MBP revealed subunits with an Mr of 43 kDa and 30 kDa, respectively. These subunits were sensitive to collagenase treatment which reduced the Mr to 20 kDa. Electron micrographs revealed a prominent flexible tetramer molecule (diameter 96 nm) in the BK preparations, a predominantly hexameric structure (diameter 30 nm) in the MBP preparations, and single annular pentameric disc-like molecules (diameter 11 nm) in the SAP preparations.

Andersen, Ove; Friis, P

1992-01-01

31

Autoantibodies against protective molecules--C1q, C-reactive protein, serum amyloid P, mannose-binding lectin, and apolipoprotein A1: prevalence in systemic lupus erythematosus.  

Science.gov (United States)

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of several autoantibodies. Among the multiple factors involved in SLE development, apoptotic defects and impaired clearance of cellular debris have gained considerable interest, as they contribute to autoantigen overload. Several molecules of the innate immunity, also participate in the removal of damaged and apoptotic cells. Among them are C1q, C-reactive protein (CRP), serum amyloid P protein (SAP), mannose-binding lectin (MBL), and apolipoprotein A1 (APO A1). To evaluate the prevalence of autoantibodies against CRP, SAP, MBL, APO A1, and C1q among SLE patients, and their relationship with disease activity, a total of 150 SLE patients were screened for the presence of elevated antibody titers against C1q, CRP, SAP, MBL, and APO A1, utilizing the enzyme-linked immunosorbent assay (ELISA) method. Disease activity was assessed using the ECLAM or SLEDAI scores. The study population comprised two groups of patients: 100 patients with quiescent disease (median ECLAM score 2) comprised the first group, and 50 patients with active disease (median SLEDAI score 16) comprised group 2. Elevated titers of anti-CRP antibodies were significantly elevated only in group 1 (10% versus 4% of controls). Antibodies against SAP were evaluated only among patients in group 1, and were found at a significant high prevalence (20%). Elevated titers of anti-MBL antibodies were significantly elevated only in group 1 (15% versus 3.6%); and antibodies directed against APO A1 were significantly elevated in 21% of group 1, and 50% of group 2 patients. Elevated titers of anti-C1q were evaluated only in group 2, and were found at a significant prevalence of 66%. Significant correlation with disease activity was found only for anti-APO A1 antibodies, and only in group 1. Several patients harbored more than one of the autoantibodies tested. In patients with SLE, autoantibodies directed against protective molecules, that is, acute-phase proteins involved in the disposal of cellular and nuclear debris, can be detected. These autoantibodies may play a pathogenic role in the development or perpetuation of autoimmunity in SLE. PMID:17899624

Shoenfeld, Yehuda; Szyper-Kravitz, Martine; Witte, Torsten; Doria, Andrea; Tsutsumi, Akito; Tatsuya, Abe; Dayer, Jean-Michel; Roux-Lombard, Pascale; Fontao, Lionel; Kallenberg, Cees G M; Bijl, Marc; Matthias, Torsten; Fraser, Abigail; Zandman-Goddard, Gisele; Blank, Miri; Gilburd, Boris; Meroni, Pier Luigi

2007-06-01

32

Does serum degrade amyloid fibrils? Failure to confirm enzymatic degradation of amyloid A fibrils as the basis of the so-called ''amyloid degrading activity'' of serum  

International Nuclear Information System (INIS)

Several reports from different laboratories on the capacity of serum to degrade amyloid A fibrils have been published since the 1979 Symposium on Amyloidosis. These are based on the observation that whole serum or isolated serum albumin causes increased translucency in turbid agarose gels containing AA fibrils. We report here a critical study of the phenomenon itself both in the gel and in solution. The clearing capacity of serum samples correlated well with albumin concentration and as previously shown by others was manifested by isolated albumin preparations. Clearing did not involve enzymatic degradation of amyloid fibrils. Serum albumin is known to improve the optical clarity of agarose gels. We propose that optical clearing without degradation is the basis of the so-called amyloid degrading activity of human serum. This activity does not seem to be of in vivo importance in amyloidogenesis

33

Purification of amyloid protein AA subspecies from amyloid-rich human tissues.  

Science.gov (United States)

Protein AA, the major amyloid fibril protein in reactive (secondary) systemic amyloidosis is derived from the acute phase reactant liver-produced apolipoprotein serum AA (SAA) by proteolytic cleavage, usually in the C-terminal half of the 104 amino acid residues long precursor. The cleavage points in SAA vary between patients and the deposited protein AA is often quite heterogeneous. In this chapter, we describe methods to extract amyloid fibrils and to purify protein AA by sequential gel filtration. Further purification of subspecies of protein AA is best achieved by the use of differences in charge and chromatofocusing is described as the method of choice. Analytic methods include sodium dodecylsulfate polyacrylamide gel electrophoresis and analytic isoelectric focusing. PMID:15980608

Westermark, Gunilla T; Westermark, Per

2005-01-01

34

Serum amyloid A isoforms in serum and synovial fluid from spontaneously diseased dogs with joint diseases or other conditions  

DEFF Research Database (Denmark)

Serum amyloid A (SAA) is a major acute phase protein in dogs. However, knowledge of qualitative properties of canine SAA and extent of its synthesis in extrahepatic tissues is limited. The aim of the study was to investigate expression of different SAA isoforms in serum and synovial fluid in samples obtained from dogs (n = 16) suffering from different inflammatory or non-inflammatory conditions, which were either related or unrelated to joints. Expression of SAA isoforms was visualized by denaturing isoelectric focusing and Western blotting. Serum amyloid A was present in serum from all dogs with systemic inflammatory activity, and up to four major isoforms with apparent isoelectric points between 6.1 and 7.9 were identified. In synovial fluid from inflamed joints one or more highly alkaline SAA isoforms (with apparent isoelectric points above 9.3) were identified, with data suggesting local production of these isoforms in the canine inflamed joint.

Kjelgaard-Hansen, Mads Jens; Christensen, Michelle B.

2007-01-01

35

Transthyretin sequesters amyloid beta protein and prevents amyloid formation  

DEFF Research Database (Denmark)

The cardinal pathological features of Alzheimer disease are depositions of aggregated amyloid beta protein (A beta) in the brain and cerebrovasculature. However, the A beta is found in a soluble form in cerebrospinal fluid in healthy individuals and patients with Alzheimer disease. We postulate that sequestration of A beta precludes amyloid formation. Failure to sequester A beta in Alzheimer disease may result in amyloidosis. When we added A beta to cerebrospinal fluid of patients and controls it was rapidly sequestered into stable complexes with transthyretin. Complexes with apolipoprotein E, which has been shown to bind A beta in vitro, were not observed in cerebrospinal fluid. Additional in vitro studies showed that both purified transthyretin and apolipoprotein E prevent amyloid formation.

Schwarzman, A L; Gregori, L

1994-01-01

36

Protein chemistry: Catalytic amyloid fibrils  

Science.gov (United States)

Amyloid fibrils are formed from polypeptide chains assembled into an organized fibrillar structure. Now, it has been shown that such fibrillar structures can also bind metal ions and catalyse chemical reactions.

Aumüller, Tobias; Fändrich, Marcus

2014-04-01

37

C-reactive protein, haptoglobin, serum amyloid A and pig major acute phase protein response in pigs simultaneously infected with H1N1 swine influenza virus and Pasteurella multocida  

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Full Text Available Abstract Background Swine influenza (SI is an acute respiratory disease caused by swine influenza virus (SIV. Swine influenza is generally characterized by acute onset of fever and respiratory symptoms. The most frequent complications of influenza are secondary bacterial pneumonia. The objective of this work was to study the acute phase proteins (APP responses after coinfection of piglets with H1N1 swine influenza virus (SwH1N1 and Pasteurella multocida (Pm in order to identify whether the individual APP response correlate with disease severity and whether APP could be used as markers of the health status of coinfected pigs. Results In all coinfected pigs clinical sings, including fever, coughing and dyspnea, were seen. Viral shedding was observed from 2 to 7 dpi. The mean level of antibodies against Pm dermonecrotoxin in infected piglets increase significantly from 7 dpi. Anti-SwH1N1 antibodies in the serum were detected from 7 dpi. The concentration of C-reactive protein (CRP increased significantly at 1 dpi as compared to control pigs, and remained significantly higher to 3 dpi. Level of serum amyloid A (SAA was significantly higher from 2 to 3 dpi. Haptoglobin (Hp was significantly elevated from 3 dpi to the end of study, while pig major acute phase protein (Pig-MAP from 3 to 7 dpi. The concentrations of CRP, Hp and SAA significantly increased before specific antibodies were detected. Positive correlations were found between serum concentration of Hp and SAA and lung scores, and between clinical score and concentrations of Pig-MAP and SAA. Conclusions The results of current study confirmed that monitoring of APP may revealed ongoing infection, and in this way may be useful in selecting clinically healthy pigs (i.e. before integration into an uninfected herd. Present results corroborated our previous findings that SAA could be a potentially useful indicator in experimental infection studies (e.g. vaccine efficiency investigations or as a marker for disease severity, because of correlation observed between its concentration in serum and disease severity (lung scores, clinical scores.

Pomorska-Mól Ma?gorzata

2013-01-01

38

Determination of serum amyloid P component in seminal plasma and correlations with serum hormone levels in young, healthy men.  

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Abstract Serum amyloid P component (SAP) belongs to the pentraxin family of proteins. SAP is evolutionary conserved, and involved in amyloidosis, innate immunity, inflammation, and apoptosis. We have previously described SAP in the male reproductive tract, where it occurs in seminal fluid, on spermatozoa, and in epididymal, seminal vesicle, and prostate tissue. In the present investigation, our aim was to characterize SAP in male reproduction. In short, we developed and evaluated an immunoass...

Sonesson, Annika; Hillarp, Andreas; Giwercman, Aleksander; Malm, Johan

2011-01-01

39

Ligand-binding sites in human serum amyloid P component  

DEFF Research Database (Denmark)

Amyloid P component (AP) is a naturally occurring glycoprotein that is found in serum and basement membranes, AP is also a component of all types of amyloid, including that found in individuals who suffer from Alzheimer's disease and Down's syndrome. Because AP has been found to bind strongly and specifically to certain glycosaminoglycans that are components of amyloid deposits, AP may play an important role in the maintenance of amyloid. In the present work, we isolated and identified two proteolytic fragments of AP that are responsible for its heparin-binding activity. Neither fragment corresponds to published heparin-binding sequences. The structural requirements for activity of the peptides (amino acid residues 27-38 and 192-203 of AP) were examined by means of solid-phase inhibition assays with synthetic peptides, AP-(192-203)-peptide inhibits the Ca2+-dependent binding of AP to heparin with an IC50 of 25 mu M, while the IC50 of AP-(27-38)-peptide and AP-(33-38)-peptide are 10 mu M and 2 mu M, respectively, The understanding of the structure and function of active AP peptides will be useful for development of amyloid-targeted diagnostics and therapeutics.

Heegaard, Peter M. H.

1996-01-01

40

Evaluation of goose serum amyloid a acute phase response by enzyme-linked immunosorbent assay.  

Science.gov (United States)

Serum amyloid A (SAA) is of interest as the circulating precursor of amyloid A protein, the fibrillar component of AA (secondary) amyloid deposits, and also as an extremely sensitive and rapid major acute phase protein. Serum concentrations of acute phase proteins (APPs) provide valuable information about the diagnosis and prognosis of various diseases, and thus the relevance of APPs for monitoring the health status of domestic animals is widely accepted. More importantly, the measurement of SAA concentration assists in assessing the prognosis in secondary amyloidosis, which is a common disease of geese, affecting an increasing number of animals. In the present study we introduce a highly sensitive goose-specific ELISA method for measuring SAA concentration in goose serum or plasma samples. Samples were taken from geese of the Landes Grey and Hungarian White breeds, which were stimulated for an acute phase reaction by administration of a commercially available fowl cholera vaccine containing inactivated Pasteurella multocida. Strong and characteristically rapid acute phase responses were measured in both breeds, peaking at approximately 24 h after inoculation. The maximum SAA concentration was 1200 microg/ml. At 72 h postinoculation, the concentrations returned to pre-inoculation values. There was significantly (p = 0.004) less intense response in the control groups; however, a very mild increase of SAA levels was detected due to the stress inevitably caused by the sampling procedure. PMID:17867462

Kovács, Beáta Marianna; Toussaint, Mathilda J M; Gruys, E; Fábián, Ibolya B; Szilágyi, L; Janan, J; Rudas, P

2007-09-01

 
 
 
 
41

Evaluation of systemic amyloidosis by scintigraphy with 123I-labeled serum amyloid P component  

International Nuclear Information System (INIS)

In systemic amyloidosis the distribution and progression of disease have been difficult to monitor, because they can be demonstrated only by biopsy. Serum amyloid P component (SAP) is a normal circulating plasma protein that is deposited on amyloid fibrils because of its specific binding affinity for them. We investigated whether labeled SAP could be used to locate amyloid deposits. Purified human SAP labeled with iodine-123 was given intravenously to 50 patients with biopsy-proved systemic amyloidosis--25 with the AL (primary) type and 25 with the AA (secondary) type--and to 26 control patients with disease and 10 healthy subjects. Whole-body images and regional views were obtained after 24 hours and read in a blinded fashion. In the patients with amyloidosis the 123I-SAP was localized rapidly and specifically in amyloid deposits. The scintigraphic images obtained were characteristic and appeared to identify the extent of amyloid deposition in all 50 patients. There was no uptake of the 123I-SAP by the control patients and the healthy subjects. In all patients with AA amyloidosis the spleen was affected, whereas the scans showed uptake in the heart, skin, carpal region, and bone marrow only in patients with the AL type. Positive images were seen in six patients in whom biopsies had been negative or unsuccessful; in all six, amyloid was subsequently found on biopsy or at autopsy. Progressive amyloid deposition was observed in 9 of 11 patients studied serially. Scintigraphy after the injection of 123I-SAP can be used for diagnosing, locating, and monitoring the extent of systemic amyloidosis

42

Establishment of a Transgenic Mouse Model Specifically Expressing Human Serum Amyloid A in Adipose Tissue  

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Obesity and obesity co-morbidities are associated with a low grade inflammation and elevated serum levels of acute phase proteins, including serum amyloid A (SAA). In the non-acute phase in humans, adipocytes are major producers of SAA but the function of adipocyte-derived SAA is unknown. To clarify the role of adipocyte-derived SAA, a transgenic mouse model expressing human SAA1 (hSAA) in adipocytes was established. hSAA expression was analysed using real-time PCR analysis. Male animals were...

Olsson, Maja; Ahlin, Sofie; Olsson, Bob; Svensson, Per-arne; Sta?hlman, Marcus; Bore?n, Jan; Carlsson, Lena M. S.; Sjo?holm, Kajsa

2011-01-01

43

Processing of the amyloid precursor protein and its paralogues amyloid precursor-like proteins 1 and 2  

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Alzheimer’s disease (AD) is a neurodegenerative disorder which is histopathologically characterised by amyloid plaques and neurofibrillary tangles. Amyloid plaques consist of the amyloid ?-peptide (A?) that can form aggregates in the brain. A? is generated from the amyloid precursor protein (APP) through proteolytic cleavage. APP belongs to a conserved protein family that also includes the two paralogues, APP-like proteins 1 and 2 (APLP1 and APLP2). Despite the immense amount of research...

Adlerz, Linda

2007-01-01

44

Expression of serum amyloid a in equine wounds  

DEFF Research Database (Denmark)

OBJECTIVES Aberrant wound healing with formation of exuberant granulation tissue (EGT) occurs frequently in horses and may affect their athletic career and quality of life. The objective of the study was to determine mRNA expression levels of serum amyloid A (SAA) in normal and aberrant wound healing in horses. METHODS Experimental wounds were made in six horses on both metatarsi and on regio brachii. One limb was bandaged to provoke formation of EGT. Biopsies were collected on day 21 and were divided in three groups: body wounds (regio brachii), unbandaged limb wounds (normal healing), and bandaged limb wounds (aberrant healing with formation of EGT). All biopsies were examined for the relative mRNA expression level of SAA using qRT-PCR. Differences in SAA expression levels between the three groups were analyzed by Kruskal-Wallis test and Dunns test. RESULTS SAA mRNA level was significantly higher (P < 0.01) in limb wounds healing with EGT formation than in body and limb wounds with normal healing. In body wounds and limb wounds with normal healing SAA expression was very low, in EGT SAA expression levels varied from low to very high. CONCLUSIONS SAA is a major equine acute phase protein, which is produced in the liver and several extrahepatic tissues during inflammatory conditions. This study shows that cells in EGT derived from horses produce SAA. This may be related to the length of the inflammatory phase of the wound healing, which is short (approximately 3 days) in wounds with normal healing, but protracted in wounds developing EGT. Chronic inflammation might facilitate binding of SAA-containing high density lipoproteins (HDL) to extracellular vascular proteoglycans, which would favour retention and modification of HDL by the vascular matrix. Such changes could be the pathophysiological reason underlying endothelial cell dysfunction and subsequently hypoxia, which has been implicated in EGT formation.

SØrensen, Mette Aamand; Jacobsen, Stine

45

Protective molecules--C-reactive protein (CRP), serum amyloid P (SAP), pentraxin3 (PTX3), mannose-binding lectin (MBL), and apolipoprotein A1 (Apo A1), and their autoantibodies: prevalence and clinical significance in autoimmunity.  

Science.gov (United States)

Apoptotic defects and impaired clearance of cellular debris are considered key events in the development of autoimmunity, as they can contribute to autoantigen overload, and may initiate an autoimmune response. The pentraxins are a group of highly conserved proteins including the short pentraxins, C-reactive protein (CRP) and serum amyloid-P (SAP), and the long pentraxin-3 (PTX3), which are all involved in innate immunity and in acute-phase responses. Mannan-binding lectin (MBL) is an activator of the complement system, and Apolipoprotein A-1 (Apo A-1) is pivotal in the cholesterol homeostasis and has anti-inflammatory properties. In addition to their role in innate immunity and inflammation, each of these five proteins participates in the removal of damaged and apoptotic cells. In this review, we discuss the clinical significance of different levels of these proteins, their role in the induction or protection from autoimmunity, and the presence of specific autoantibodies against them in the different autoimmune diseases. PMID:16380821

Kravitz, Martine Szyper; Pitashny, Milena; Shoenfeld, Yehuda

2005-11-01

46

Multiple isoforms of the human pentraxin serum amyloid P component.  

DEFF Research Database (Denmark)

Human serum amyloid P component (SAP) isolated from 20 healthy individuals was analyzed by anion exchange chromatography and isoelectric focusing (IEF) in order to investigate the existence of multiple forms of SAP and interindividual structural differences. Anion exchange chromatography showed one major and several minor subpopulations of SAP. IEF of all SAP isolates showed a previously unreported degree of heterogeneity with six isoelectric forms (pKi range 5.5-6.1) and with minor interindividual differences in respect of isoelectric points. Total enzymatic deglycosylation of SAP reduced the number of bands in IEF to two indicating the existence of two types of polypeptide chains.

SØrensen, Inge Juul; Andersen, Ove

1995-01-01

47

Serum amyloid A-related inflammation is lowered by increased fruit and vegetable intake, while high-sensitive C-reactive protein, IL-6 and E-selectin remain unresponsive.  

Science.gov (United States)

The present study assessed whether increased fruit and vegetable (F&V) intake reduced the concentrations of the inflammatory marker serum amyloid A (SAA) in serum, HDL2 and HDL3 and whether the latter reduction influenced any of the functional properties of these HDL subfractions. The present study utilised samples from two previous studies: (1) the FAVRIT (Fruit and Vegetable Randomised Intervention Trial) study - hypertensive subjects (systolic blood pressure (BP) range 140-190 mmHg; diastolic BP range 90-110 mmHg) were randomised to receive a 1-, 3- or 6-portion F&V/d intervention for 8 weeks, and (2) the ADIT (Ageing and Dietary Intervention Trial) study - older subjects (65-85 years) were randomised to receive a 2- or 5-portion F&V/d intervention for 16 weeks. HDL2 and HDL3 were isolated by rapid ultracentrifugation. Measurements included the following: serum high-sensitive C-reactive protein (hsCRP) by an immunoturbidimetric assay; serum IL-6 and E-selectin and serum-, HDL2- and HDL3-SAA by ELISA procedures; serum-, HDL2- and HDL3-cholesterol ester transfer protein (CETP) activity by a fluorometric assay. Although the concentrations of hsCRP, IL-6 and E-selectin were unaffected by increasing F&V intake in both studies (P>0·05 for all comparisons), those of SAA in HDL3 decreased in the FAVRIT cohort (P= 0·049) and those in HDL2 and HDL3 decreased in the ADIT cohort (P= 0·035 and 0·032), which was accompanied by a decrease in the activity of CETP in HDL3 in the FAVRIT cohort (P= 0·010) and in HDL2 in the ADIT cohort (P= 0·030). These results indicate that SAA responds to increased F&V intake, while other inflammatory markers remain unresponsive, and this leads to changes in HDL2 and HDL3, which may influence their antiatherogenic potential. Overall, the present study provides tangible evidence of the effectiveness of increased F&V intake, which may be of use to health policy makers and the general public. PMID:25141100

Nadeem, Nida; Woodside, Jayne V; Neville, Charlotte E; McCall, Damian O; McCance, David; Edgar, David; Young, Ian S; McEneny, Jane

2014-10-01

48

Serum amyloid-A levels in neonatal necrotizing enterocolitis.  

Science.gov (United States)

We aimed to evaluate serum levels of serum amyloid-A (SAA) both in the diagnosis and monitoring the treatment response of necrotizing enterocolitis (NEC). Forty-five preterm neonates were enrolled in the study, including 15 infants with NEC, 15 with sepsis, and 15 healthy preterm infants. Pre- and posttreatment serum SAA levels were measured. Among patients with NEC, 11 had stage 1 and 4 had stage 2 disease according to the modified Bell's staging criteria. Baseline SAA levels of the infants with NEC were significantly higher than controls (P=0.013) and were significantly lower than those with sepsis (P=0.004). When infants with stage 1 and stage 2 NEC were analyzed separately, baseline SAA levels of the infants with stage 2 NEC were significantly higher than controls (P=0.027) than those with stage 1 NEC (P=0.018), but similar to those with sepsis. There was a trend that baseline SAA levels were also correlated with the Bell stage (r=0.501, P=0.057). Posttreatment SAA levels significantly decreased in infants with sepsis (P=0.002). Pre- and posttreatment SAA levels were similar in patients with stage 1 and 2 NEC. In conclusion, SAA rises in early stages of NEC and may aid in diagnosis as a serum marker. PMID:21786324

Eras, Zeynep; O?uz, Suna; Dizdar, Evrim Alyamac; Sari, Fatma Nur; Dilmen, U?ur

2011-01-01

49

Imaging of experimental amyloidosis with /sup 131/I-labeled serum amyloid P component  

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/sup 131/I-labeled human serum amyloid P component, which was injected into mice with experimentally induced systemic AA amyloidosis and into controls, became specifically localized and was retained in amyloidotic organs. In comparison, it was rapidly and completely eliminated from unaffected tissues and from control animals. Distinctive images of this amyloid-specific deposition of labeled serum amyloid P component were derived from whole body scanning, in vivo, of amyloidotic mice. These findings suggest that such imaging may have applications for the diagnosis and quantitation of amyloid deposits in humans.

Caspi, D.; Zalzman, S.; Baratz, M.; Teitelbaum, Z.; Yaron, M.; Pras, M.; Baltz, M.L.; Pepys, M.B.

1987-11-01

50

Imaging of experimental amyloidosis with 131I-labeled serum amyloid P component  

International Nuclear Information System (INIS)

131I-labeled human serum amyloid P component, which was injected into mice with experimentally induced systemic AA amyloidosis and into controls, became specifically localized and was retained in amyloidotic organs. In comparison, it was rapidly and completely eliminated from unaffected tissues and from control animals. Distinctive images of this amyloid-specific deposition of labeled serum amyloid P component were derived from whole body scanning, in vivo, of amyloidotic mice. These findings suggest that such imaging may have applications for the diagnosis and quantitation of amyloid deposits in humans

51

Imaging of experimental amyloidosis with 131I-labeled serum amyloid P component.  

Science.gov (United States)

131I-labeled human serum amyloid P component, which was injected into mice with experimentally induced systemic AA amyloidosis and into controls, became specifically localized and was retained in amyloidotic organs. In comparison, it was rapidly and completely eliminated from unaffected tissues and from control animals. Distinctive images of this amyloid-specific deposition of labeled serum amyloid P component were derived from whole body scanning, in vivo, of amyloidotic mice. These findings suggest that such imaging may have applications for the diagnosis and quantitation of amyloid deposits in humans. PMID:3689465

Caspi, D; Zalzman, S; Baratz, M; Teitelbaum, Z; Yaron, M; Pras, M; Baltz, M L; Pepys, M B

1987-11-01

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Iodine-123-labelled serum amyloid P component scintigraphy in amyloidosis  

International Nuclear Information System (INIS)

This study describes the results of scintigraphy with iodine-123-labelled serum amyloid P component (SAP) as a means of establishing the distribution of organ involvement in amyloidosis. The significance of 123I-SAP scans obtained in 15 patients with biopsy-proven AA or AL amyloidosis is discussed. Biopsy-proven amyloidosis was typically confirmed by scintigraphy, though such confirmation was not obtained in the kidneys in six patients with histological proof of extensive renal amyloid deposition. This lack of uptake may have been due to the accumulation of a major part of the 123I-SAP in the spleen and/or liver. Twenty-four hour whole-body retention of 123I-SAP was higher in patients with amyloidosis than in controls. Twenty-four hour tracer accumulation of the radioactivity in the extravascular compartment was notably greater in patients than in controls and appeared to be a good diagnostic criterion. We conclude that 123I-SAP scintigraphy may be helpful for the evaluation of organ involvement not only in patients with biopsy-proven amyloidosis but also when a biopsy cannot be performed or when a strong suspicion of amyloidosis exists in spite of repeated negative biopsises. (orig.)

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Serum concentrations of haptoglobin and serum amyloid A in water buffaloes (Bubalus bubalis with abomasal ulcer  

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Full Text Available To evaluate the serum concentrations of haptoglobin (Hp and serum amyloid A (SAA in water buffaloes with abomasal ulcers, the abomasums of 100 randomly selected water buffaloes were examined after slaughter. Type I abomasal ulcers were found in 56 out of 100 buffaloes. Serum concentrations of Hp and SAA were measured. There was no significant difference between affected and non-affected buffaloes in the serum concentrations of Hp and SAA. The serum concentrations of Hp and SAA had no significant correlation with age and the serum SAA revealed no significant correlation with the number of abomasal ulcers. A significant correlation was found between the serum Hp and the number of abomasal ulcers (r =0.29, p = 0.04. There was no significant difference in the serum concentrations of Hp and SAA between buffaloes with different ulcer locations in the abomasums. Although more work on a larger number of animals is required in this area, it seems that the measurement of the serum Hp can be used to predict the abundance of type I abomasal ulcers.

Javad Tajik

2012-09-01

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Alzheimer's disease and the amyloid ?-protein.  

Science.gov (United States)

Alzheimer's disease is a devastating disorder that is estimated to affect more than 25 million people worldwide and for which there are no preventive, disease-modifying, or curative therapies. Substantial evidence indicates that the amyloid ?-protein (A?) is a seminal factor in disease causation and may be a tractable therapeutic target. The ability of A? to self-associate to form oligomeric assemblies appears to underlie the early toxic events that lead to memory impairment and subsequent neurodegeneration. We review here research on A? folding, self-assembly, and toxicity, highlighting areas critical for the development of efficacious A?-directed therapeutics. PMID:22482449

Walsh, Dominic M; Teplow, David B

2012-01-01

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Novel mediators of amyloid precursor protein signaling.  

Science.gov (United States)

Multiple recent reports implicate amyloid precursor protein (APP) signaling in the pathogenesis of Alzheimer's disease, but the APP-dependent signaling network involved has not been defined. Here, we report a novel consensus sequence for interaction with the PDZ-1 and PDZ-2 domains of the APP-interacting proteins Mint1, Mint2, and Mint3 (X11alpha, X11beta, and X11gamma), and multiple novel interactors for these proteins, with the finding that transcriptional coactivators are highly represented among these interactors. Furthermore, we show that Mint3 interaction with a set of the transcriptional coactivators leads to nuclear localization and transactivation, whereas interaction of the same set with Mint1 or Mint2 prevents nuclear localization and transactivation. These results define new mediators of the signal transduction network mediated by APP. PMID:20016085

Swistowski, Andrzej; Zhang, Qiang; Orcholski, Mark E; Crippen, Danielle; Vitelli, Cathy; Kurakin, Alexei; Bredesen, Dale E

2009-12-16

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Exogenous amyloidogenic proteins function as seeds in amyloid ?-protein aggregation.  

Science.gov (United States)

Amyloid ?-protein (A?) aggregation is considered to be a critical step in the neurodegeneration of Alzheimer's disease (AD). In addition to A?, many proteins aggregate into the amyloid state, in which they form elongated fibers with spines comprising stranded ?-sheets. However, the cross-seeding effects of other protein aggregates on A? aggregation pathways are not completely clear. To investigate the cross-seeding effects of exogenous and human non-CNS amyloidogenic proteins on A? aggregation pathways, we examined whether and how sonicated fibrils of casein, fibroin, sericin, actin, and islet amyloid polypeptide affected A?40 and A?42 aggregation pathways using the thioflavin T assay and electron microscopy. Interestingly, the fibrillar seeds of all amyloidogenic proteins functioned as seeds. The cross-seeding effect of actin was stronger but that of fibroin was weaker than that of other proteins. Furthermore, our nuclear magnetic resonance spectroscopic studies identified the binding sites of A? with the amyloidogenic proteins. Our results indicate that the amyloidogenic proteins, including those contained in foods and cosmetics, contribute to A? aggregation by binding to A?, suggesting their possible roles in the propagation of A? amyloidosis. PMID:24440525

Ono, Kenjiro; Takahashi, Ryoichi; Ikeda, Tokuhei; Mizuguchi, Mineyuki; Hamaguchi, Tsuyoshi; Yamada, Masahito

2014-04-01

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Beta-amyloid precursor protein cleavage by a membrane-bound protease.  

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The principal component of amyloid plaques in Alzheimer disease is beta-amyloid protein, an approximately 4-kDa peptide derived from amyloid precursor proteins. Previous studies have established that amyloid precursor proteins are secreted after proteolytic cleavage within the beta-amyloid peptide. The present investigation documents that, in cultured cells, amyloid precursor protein is cleaved on the plasma membrane by a membrane-bound endoprotease and that the specificity of peptide bond hy...

Sisodia, S. S.

1992-01-01

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Validación analítica de técnicas comerciales para la determinación de haptoglobina, proteína C reactiva y amiloide A sérico en caninos Analytical validation of commercial assays for the determination of haptoglobin, C-reactive protein and serum amyloid A in dogs  

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Full Text Available Los métodos analíticos deben ser validados antes de emplearlos de forma rutinaria en un laboratorio. El objetivo de este trabajo fue validar tres métodos analíticos comerciales usados en nuestro laboratorio para la determinación de haptoglobina (Hp, proteína C reactiva (CRP y amiloide A sérico (SAA en muestras de la especie canina con concentraciones bajas y altas de proteínas de fase aguda (PFAs. Los parámetros que se evaluaron para la validación fueron: (1 Precisión, determinada mediante el cálculo de los coeficientes de variación (CVs intra e interdeterminación. (2 Exactitud, evaluada indirectamente comprobando la linealidad bajo dilución. (3 Límite de detección, determinado como la mínima concentración que puede distinguirse de una muestra de valor 0. Todos los CVs intradeterminación fueron All laboratory tests must be validated before being introduced for patient testing. The objective of this work was to perform the analytical validation of three commercial assays that are being used at our laboratory for the determination of haptoglobin (Hp, C reactive protein (CRP and serum amyloid A (SAA in canine samples with low and high concentrations of these acute phase proteins (APPs. The parameters evaluated for the validation of the methods were: (1 Precision, assessed by determination of the within and between-run coefficients of variation (CVs. (2 Inaccuracy, evaluated indirectly by investigating linearity under dilution. (3 Limit of detection, determined as the lowest concentration of the APPs which could be distinguished from a zero sample. All within-run CVs were lower than 10%, however between-run CVs were lower than 10% only for Hp. Dilution of a serum sample with high concentrations of the different APPs resulted in a linear regression equation with correlation coefficient R²0.98 in all cases; so all methods showed a good accuracy. The detection limit of each assay was 0.02 g/L for Hp, 0.15 mg/L for CRP and 0.79 mg/L for SAA. Additionally they differentiate animals with inflammatory or infectious diseases from healthy subjects. Overall results of validation showed that the assays tested can be suitable for the routine measurement of APPs in canine samples, although it would be desirable to reduce the between run imprecision found for CRP and SAA assays.

S. Martínez-Subiela

2005-01-01

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Search for Amyloid-Binding Proteins by Affinity Chromatography  

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Amyloid binging proteins’ is a generic term used to designate proteins that interact with different forms of amyloidogenic peptides or proteins and that, as a result, may modulate their physiological and pathological functions by altering solubility, transport, clearance, degradation, and fibril formation. We describe a simple affinity chromatography protocol to isolate and characterize amyloid-binding proteins based on the use of sequential elution steps that may provide further informa...

Calero, Miguel; Rostagno, Agueda; Ghiso, Jorge

2012-01-01

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Formation of soluble amyloid oligomers and amyloid fibrils by the multifunctional protein vitronectin  

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Full Text Available Abstract Background The multifunctional protein vitronectin is present within the deposits associated with Alzheimer disease (AD, age-related macular degeneration (AMD, atherosclerosis, systemic amyloidoses, and glomerulonephritis. The extent to which vitronectin contributes to amyloid formation within these plaques, which contain misfolded, amyloidogenic proteins, and the role of vitronectin in the pathophysiology of the aforementioned diseases is currently unknown. The investigation of vitronectin aggregation is significant since the formation of oligomeric and fibrillar structures are common features of amyloid proteins. Results We observed vitronectin immunoreactivity in senile plaques of AD brain, which exhibited overlap with the amyloid fibril-specific OC antibody, suggesting that vitronectin is deposited at sites of amyloid formation. Of particular interest is the growing body of evidence indicating that soluble nonfibrillar oligomers may be responsible for the development and progression of amyloid diseases. In this study we demonstrate that both plasma-purified and recombinant human vitronectin readily form spherical oligomers and typical amyloid fibrils. Vitronectin oligomers are toxic to cultured neuroblastoma and retinal pigment epithelium (RPE cells, possibly via a membrane-dependent mechanism, as they cause leakage of synthetic vesicles. Oligomer toxicity was attenuated in RPE cells by the anti-oligomer A11 antibody. Vitronectin fibrils contain a C-terminal protease-resistant fragment, which may approximate the core region of residues essential to amyloid formation. Conclusion These data reveal the propensity of vitronectin to behave as an amyloid protein and put forth the possibilities that accumulation of misfolded vitronectin may contribute to aggregate formation seen in age-related amyloid diseases.

Langen Ralf

2008-10-01

 
 
 
 
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Regulation of Serum Amyloid A3 (SAA3) in Mouse Colonic Epithelium and Adipose Tissue by the Intestinal Microbiota  

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The gut microbiota has been proposed as an environmental factor that affects the development of metabolic and inflammatory diseases in mammals. Recent reports indicate that gut bacteria-derived lipopolysaccharide (LPS) can initiate obesity and insulin resistance in mice; however, the molecular interactions responsible for microbial regulation of host metabolism and mediators of inflammation have not been studied in detail. Hepatic serum amyloid A (SAA) proteins are markers and proposed mediat...

Reigstad, Christopher S.; Lunde?n, Gunnel O?stergren; Felin, Jenny; Ba?ckhed, Fredrik

2009-01-01

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Proteolytic processing of Alzheimer's ?-amyloid precursor protein.  

Science.gov (United States)

?-Amyloid precursor protein (APP) is a critical factor in the pathogenesis of Alzheimer's disease (AD). APP undergoes post-translational proteolysis/processing to generate the hydrophobic ?-amyloid (A?) peptides. Deposition of A? in the brain, forming oligomeric A? and plaques, is identified as one of the key pathological hallmarks of AD. The processing of APP to generate A? is executed by ?- and ?-secretase and is highly regulated. A? toxicity can lead to synaptic dysfunction, neuronal cell death, impaired learning/memory and abnormal behaviors in AD models in vitro and in vivo. Aside from A?, proteolytic cleavages of APP can also give rise to the APP intracellular domain, reportedly involved in multiple types of cellular events such as gene transcription and apoptotic cell death. In addition to amyloidogenic processing, APP can also be cleaved by ?-secretase to form a soluble or secreted APP ectodomain (sAPP-?) that has been shown to be mostly neuro-protective. In this review, we describe the mechanisms involved in APP metabolism and the likely functions of its various proteolytic products to give a better understanding of the patho/physiological functions of APP. PMID:22122372

Zhang, Han; Ma, Qilin; Zhang, Yun-wu; Xu, Huaxi

2012-01-01

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Soluble aggregates of the amyloid-? peptide are trapped by serum albumin to enhance amyloid-? activation of endothelial cells  

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Full Text Available Abstract Background Self-assembly of the amyloid-? peptide (A? has been implicated in the pathogenesis of Alzheimer's disease (AD. As a result, synthetic molecules capable of inhibiting A? self-assembly could serve as therapeutic agents and endogenous molecules that modulate A? self-assembly may influence disease progression. However, increasing evidence implicating a principal pathogenic role for small soluble A? aggregates warns that inhibition at intermediate stages of A? self-assembly may prove detrimental. Here, we explore the inhibition of A?1–40 self-assembly by serum albumin, the most abundant plasma protein, and the influence of this inhibition on A?1–40 activation of endothelial cells for monocyte adhesion. Results It is demonstrated that serum albumin is capable of inhibiting in a dose-dependent manner both the formation of A?1–40 aggregates from monomeric peptide and the ongoing growth of A?1–40 fibrils. Inhibition of fibrillar A?1–40 aggregate growth is observed at substoichiometric concentrations, suggesting that serum albumin recognizes aggregated forms of the peptide to prevent monomer addition. Inhibition of A?1–40 monomer aggregation is observed down to stoichiometric ratios with partial inhibition leading to an increase in the population of small soluble aggregates. Such partial inhibition of A?1–40 aggregation leads to an increase in the ability of resulting aggregates to activate endothelial cells for adhesion of monocytes. In contrast, A?1–40 activation of endothelial cells for monocyte adhesion is reduced when more complete inhibition is observed. Conclusion These results demonstrate that inhibitors of A? self-assembly have the potential to trap small soluble aggregates resulting in an elevation rather than a reduction of cellular responses. These findings provide further support that small soluble aggregates possess high levels of physiological activity and underscore the importance of resolving the effect of A? aggregation inhibitors on aggregate size.

Gonzalez-Velasquez Francisco J

2009-04-01

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Statistical Mechanical Treatments of Protein Amyloid Formation  

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Full Text Available Protein aggregation is an important field of investigation because it is closely related to the problem of neurodegenerative diseases, to the development of biomaterials, and to the growth of cellular structures such as cyto-skeleton. Self-aggregation of protein amyloids, for example, is a complicated process involving many species and levels of structures. This complexity, however, can be dealt with using statistical mechanical tools, such as free energies, partition functions, and transfer matrices. In this article, we review general strategies for studying protein aggregation using statistical mechanical approaches and show that canonical and grand canonical ensembles can be used in such approaches. The grand canonical approach is particularly convenient since competing pathways of assembly and dis-assembly can be considered simultaneously. Another advantage of using statistical mechanics is that numerically exact solutions can be obtained for all of the thermodynamic properties of fibrils, such as the amount of fibrils formed, as a function of initial protein concentration. Furthermore, statistical mechanics models can be used to fit experimental data when they are available for comparison.

John S. Schreck

2013-08-01

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Genetic Dissection of the Amyloid Precursor Protein in Developmental Function and Amyloid Pathogenesis*?  

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Proteolytic processing of the amyloid precursor protein (APP) generates large soluble APP derivatives, ?-amyloid (A?) peptides, and APP intracellular domain. Expression of the extracellular sequences of APP or its Caenorhabditis elegans counterpart has been shown to be sufficient in partially rescuing the CNS phenotypes of the APP-deficient mice and the lethality of the apl-1 null C. elegans, respectively, leaving open the question as what is the role of the highly conserved APP intracellul...

Li, Hongmei; Wang, Zilai; Wang, Baiping; Guo, Qinxi; Dolios, Georgia; Tabuchi, Katsuhiko; Hammer, Robert E.; Su?dhof, Thomas C.; Wang, Rong; Zheng, Hui

2010-01-01

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Modeling of chemical inhibition from amyloid protein aggregation kinetics  

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Backgrounds The process of amyloid proteins aggregation causes several human neuropathologies. In some cases, e.g. fibrillar deposits of insulin, the problems are generated in the processes of production and purification of protein and in the pump devices or injectable preparations for diabetics. Experimental kinetics and adequate modelling of chemical inhibition from amyloid aggregation are of practical importance in order to study the viable processing, formulation and storage as well as to...

Va?zquez, Jose? Antonio

2014-01-01

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Regulation of amyloid-? production by the prion protein  

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Alzheimer disease (AD) is characterized by the amyloidogenic processing of the amyloid precursor protein (APP), culminating in the accumulation of amyloid-? peptides in the brain. The enzymatic action of the ?-secretase, BACE1 is the rate-limiting step in this amyloidogenic processing of APP. BACE1 cleavage of wild-type APP (APPWT) is inhibited by the cellular prion protein (PrPC). Our recent study has revealed the molecular and cellular mechanisms behind this observation by showing that Pr...

Griffiths, Heledd H.; Whitehouse, Isobel J.; Hooper, Nigel M.

2012-01-01

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Examining serum amyloid P component microheterogeneity using capillary isoelectric focusing and MALDI-MS  

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Serum amyloid P component (SAP) is a glycoprotein of interest due to its presence in amyloid plaque formations. As with most glycoproteins, SAP can possibly vary greatly in its isoforms, which can be an important factor towards understanding the role of SAP. Interestingly, previous characterizations suggest varying degrees of microheterogeneity, some of which are in conflict. In this work, we provide new information to clarify SAP’s microheterogeneity profile using capillary isoelectric foc...

Weiss, Noah G.; Jarvis, Jason W.; Nelson, Randall W.; Hayes, Mark A.

2011-01-01

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Amyloid diseases of yeast: prions are proteins acting as genes.  

Science.gov (United States)

The unusual genetic properties of the non-chromosomal genetic elements [URE3] and [PSI+] led to them being identified as prions (infectious proteins) of Ure2p and Sup35p respectively. Ure2p and Sup35p, and now several other proteins, can form amyloid, a linear ordered polymer of protein monomers, with a part of each molecule, the prion domain, forming the core of this ?-sheet structure. Amyloid filaments passed to a new cell seed the conversion of the normal form of the protein into the same amyloid form. The cell's phenotype is affected, usually from the deficiency of the normal form of the protein. Solid-state NMR studies indicate that the yeast prion amyloids are in-register parallel ?-sheet structures, in which each residue (e.g. Asn35) forms a row along the filament long axis. The favourable interactions possible for aligned identical hydrophilic and hydrophobic residues are believed to be the mechanism for propagation of amyloid conformation. Thus, just as DNA mediates inheritance by templating its own sequence, these proteins act as genes by templating their conformation. Distinct isolates of a given prion have different biological properties, presumably determined by differences between the amyloid structures. Many lines of evidence indicate that the Saccharomyces cerevisiae prions are pathological disease agents, although the example of the [Het-s] prion of Podospora anserina shows that a prion can have beneficial aspects. PMID:25131596

Wickner, Reed B; Edskes, Herman K; Bateman, David A; Kelly, Amy C; Gorkovskiy, Anton; Dayani, Yaron; Zhou, Albert

2014-01-01

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Cytotoxic Aggregation and Amyloid Formation by the Myostatin Precursor Protein  

Science.gov (United States)

Myostatin, a negative regulator of muscle growth, has been implicated in sporadic inclusion body myositis (sIBM). sIBM is the most common age-related muscle-wastage disease with a pathogenesis similar to that of amyloid disorders such as Alzheimer's and Parkinson's diseases. Myostatin precursor protein (MstnPP) has been shown to associate with large molecular weight filamentous inclusions containing the Alzheimer's amyloid beta peptide in sIBM tissue, and MstnPP is upregulated following ER stress. The mechanism for how MstnPP contributes to disease pathogenesis is unknown. Here, we show for the first time that MstnPP is capable of forming amyloid fibrils in vitro. When MstnPP-containing Escherichia coli inclusion bodies are refolded and purified, a proportion of MstnPP spontaneously misfolds into amyloid-like aggregates as characterised by electron microscopy and binding of the amyloid-specific dye thioflavin T. When subjected to a slightly acidic pH and elevated temperature, the aggregates form straight and unbranched amyloid fibrils 15 nm in diameter and also exhibit higher order amyloid structures. Circular dichroism spectroscopy reveals that the amyloid fibrils are dominated by ?-sheet and that their formation occurs via a conformational change that occurs at a physiologically relevant temperature. Importantly, MstnPP aggregates and protofibrils have a negative effect on the viability of myoblasts. These novel results show that the myostatin precursor protein is capable of forming amyloid structures in vitro with implications for a role in sIBM pathogenesis. PMID:20161792

Starck, Carlene S.; Sutherland-Smith, Andrew J.

2010-01-01

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Serum Amyloid A as a Predictive Marker for Radiation Pneumonitis in Lung Cancer Patients  

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Purpose: To investigate serum markers associated with radiation pneumonitis (RP) grade ?3 in patients with lung cancer who were treated with radiation therapy. Methods and Materials: Pretreatment serum samples from patients with stage Ib-IV lung cancer who developed RP within 1 year after radiation therapy were analyzed to identify a proteome marker able to stratify patients prone to develop severe RP by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Dosimetric parameters and 3 biological factors were compared. Results: Serum samples from 16 patients (28%) with severe RP (grade 3-4) and 42 patients (72%) with no or mild RP (grade 0-2) were collected for analysis. All patients received a median of 54 Gy (range, 42-70 Gy) of three-dimensional conformal radiation therapy with a mean lung dose (MLD) of 1502 cGy (range, 700-2794 cGy). An m/z peak of 11,480 Da was identified by SELDI-TOF-MS, and serum amyloid A (SAA) was the primary splitter serum marker. The receiver operating characteristic area under the curve of SAA (0.94; 95% confidence interval [CI], 0.87-1.00) was higher than those of C-reactive protein (0.83; 95% CI, 0.72-0.94), interleukin-6 (0.79; 95% CI, 0.65-0.94), and MLD (0.57; 95% CI, 0.37-0.77). The best sensitivity and specificity of combined SAA and MLD for predicting RP were 88.9% and 96.0%, respectively. Conclusions: Baseline SAA could be used as an auxiliary marker for predicting severe RP. Extreme care should be taken to limit the lung irradiation dose in patients with high SAA.

Wang, Yu-Shan [Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Department of Animal Science, National Ilan University, Ilan, Taiwan (China); Chang, Heng-Jui [Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Chang, Yue-Cune [Department of Mathematics, Tamkang University, Taipei, Taiwan (China); Huang, Su-Chen; Ko, Hui-Ling; Chang, Chih-Chia [Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Yeh, Yu-Wung; Jiang, Jiunn-Song [Department of Chest Medicine, Shin-Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Lee, Cheng-Yen; Chi, Mau-Shin [Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Chi, Kwan-Hwa, E-mail: M006565@ms.skh.org.tw [Department of Radiation Therapy and Oncology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan (China); Institute of Radiation Science and School of Medicine, National Yang-Ming University, Taipei, Taiwan (China)

2013-03-01

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Netrin-1 interacts with amyloid precursor protein and regulates amyloid-beta production.  

Science.gov (United States)

The beta-amyloid precursor protein (APP) is an orphan transmembrane receptor whose physiological role is largely unknown. APP is cleaved by proteases generating amyloid-beta (Abeta) peptide, the main component of the amyloid plaques that are associated with Alzheimer's disease. Here, we show that APP binds netrin-1, a multifunctional guidance and trophic factor. Netrin-1 binding modulates APP signaling triggering APP intracellular domain (AICD)-dependent gene transcription. Furthermore, netrin-1 binding suppresses Abeta peptide production in brain slices from Alzheimer model transgenic mice. In this mouse model, decreased netrin-1 expression is associated with increased Abeta concentration, thus supporting netrin-1 as a key regulator of Abeta production. Finally, we show that netrin-1 brain administration in Alzheimer model transgenic mice may be associated with an amelioration of the Alzheimer's phenotype. PMID:19148186

Lourenço, F C; Galvan, V; Fombonne, J; Corset, V; Llambi, F; Müller, U; Bredesen, D E; Mehlen, P

2009-05-01

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Netrin-1 interacts with amyloid precursor protein and regulates amyloid-? production  

Science.gov (United States)

The ?-amyloid precursor protein (APP) is an orphan transmembrane receptor whose physiological role is largely unknown. APP is cleaved by proteases generating amyloid-? (A?) peptide, the main component of the amyloid plaques that are associated with Alzheimer’s disease. Here, we show that APP binds netrin-1, a multifunctional guidance and trophic factor. Netrin-1 binding modulates APP signaling triggering APP intracellular domain (AICD)-dependent gene transcription. Furthermore, netrin-1 binding suppresses A? peptide production in brain slices from Alzheimer model transgenic mice. In this mouse model, decreased netrin-1 expression is associated with increased A? concentration, thus supporting netrin-1 as a key regulator of A? production. Finally, we show that netrin-1 brain administration in Alzheimer model transgenic mice may be associated with an amelioration of the Alzheimer’s phenotype. PMID:19148186

Lourenco, FC; Galvan, V; Fombonne, J; Corset, V; Llambi, F; Muller, U; Bredesen, DE; Mehlen, P

2009-01-01

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Cellular Prion Protein Expression Is Not Regulated by the Alzheimer's Amyloid Precursor Protein Intracellular Domain  

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There is increasing evidence of molecular and cellular links between Alzheimer's disease (AD) and prion diseases. The cellular prion protein, PrPC, modulates the post-translational processing of the AD amyloid precursor protein (APP), through its inhibition of the ?-secretase BACE1, and oligomers of amyloid-? bind to PrPC which may mediate amyloid-? neurotoxicity. In addition, the APP intracellular domain (AICD), which acts as a transcriptional regulator, has been reported to control the e...

Lewis, Victoria; Whitehouse, Isobel J.; Baybutt, Herbert; Manson, Jean C.; Collins, Steven J.; Hooper, Nigel M.

2012-01-01

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Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk  

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The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically pruduced SAA isoforms in plasma and milk fra cows with mastitis.

Jacobsen, Stine; Niewold, T.A.

2005-01-01

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Haptoglobin and serum amyloid a in subacute ruminal acidosis in goats  

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Full Text Available Ruminal acidosis is a frequent disorder that occurs in goats as a consequence of feedingmistakes in animals not adapted to a diet of easily fermentable carbohydrates. The subacuteform of the disease is difficult to diagnose because no apparent signs are shownand the acid-base parameters may remain within the normal range. The present studyaimed at testing the hypothesis that haptoglobin (Hp and serum amyloid A (SAA,the two major acute phase proteins in ruminants, may be useful as markers of subacuteacidosis in goats.A subacute acidosis was induced in six Murciano-Granadina goats through a diet of60% mixed feed-40% alfalfa hay offered during 5 days to goats not adapted to eatmixed feed. Two goats were rumen-fistulated to investigate the effect of feeding onruminal pH. Sampling of blood and urine of all animals was done before the inductionof the acidosis, during 5 days after the onset of induction and for 18 days after theinduction (recovery period.Ruminal pH in the fistulated goats dropped to less than 5.5 during the inductionperiod, and half of the goats had diarrhea on the third day after the induction of acidosis.Acid-base parameters showed that the acid-base compensatory mechanisms wereefficient in maintaining the equilibrium. Serum Hp had a moderate increase duringthe induction period, while SAA did not change. These results suggest that Hp mightbe a potential marker for ruminal acidosis in goats.

F.H.D. González

2010-12-01

77

Complement activation by the amyloid proteins A beta peptide and beta 2-microglobulin.  

Science.gov (United States)

Complement activation (CA) has been reported to play a role in the pathogenesis of Alzheimer's disease (AD). To investigate whether CA may contribute to amyloidogenesis in general, the CA potential of different amyloid fibril proteins was tested. CA induced by A beta preparations containing soluble protein, protofilaments and some fibrils or only fibrils in a solid phase system (ELISA) was modest with a slow kinetics compared to the positive delta IgG control. Soluble A beta induced no detectable CA in a liquid phase system (complement consumption assay) while fibrillar A beta caused CA at 200 mg/ml and higher concentrations. Soluble beta 2-microglobulin (beta 2M) purified from peritoneal dialysates was found to be as potent a complement activator as A beta in both solid and liquid phase systems while beta 2M purified from urine exhibited lower activity, a difference which may be explained by differences observed in SDS-resistant oligomers and isoforms. Soluble Amyloid A-protein caused no significant CA. A beta and beta 2M activated complement via the classical pathway. The modifying influence by amyloid-associated molecules on A beta-induced CA was also investigated, but neither serum amyloid P component nor heparan sulfate did significantly alter the A beta-induced CA. The results indicate that not only fibrillar A beta but also oligomers of, in particular, beta 2M from patients with dialysis-associated amyloidosis are capable of inducing CA at supra-physiological concentrations. PMID:10611947

Nybo, M; Nielsen, E H; Svehag, S E

1999-12-01

78

Serum amyloid A and haptoglobin concentrations in serum and peritoneal fluid of healthy horses and horses with acute abdominal pain  

DEFF Research Database (Denmark)

BACKGROUND: Peritoneal fluid (PF) analysis is a valuable diagnostic tool in equine medicine. Markers such as serum amyloid A (SAA) and haptoglobin (Hp) could facilitate the diagnosis of inflammatory abdominal conditions. OBJECTIVES: The objectives were to (1) establish reference intervals (RI) for SAA and Hp in serum and PF in healthy horses, (2) compare SAA and Hp concentrations between healthy horses and horses with colic, and (3) to assess the correlation between serum and PF concentrations. METHODS: Serum amyloid A and Hp concentrations were determined by automated assays in prospectively enrolled healthy reference horses and horses with colic. RIs were calculated, group concentrations were compared by Student's t-test, and Pearson's correlation for serum and PF concentrations were determined. RESULTS: In healthy horses (n = 62) the measurements for SAA were below the detection limit (0.5 mg/L) in 94% of serum samples and 98% of PF samples. Horses with colic (n = 61) had statistically significantly increased SAA concentrations in serum (P 

Pihl, Tina Holberg; Andersen, Pia Haubro

2013-01-01

79

Amyloid formation and inhibition of an all-beta protein: A study on fungal polygalacturonase  

Science.gov (United States)

Theoretically, all proteins can adopt the nanofibrillar structures known as amyloid, which contain cross-beta structures. The all-beta folded proteins are particularly interesting in this regard, since they appear to be naturally more predisposed toward this structural arrangement. In this study, methanol has been used to drive the beta-helix protein polygalacturonase (PG), toward amyloid fibril formation. Congo red absorbance, thioflavin T fluorescence, circular dichroism (CD) and transmission electron microscopy have been used to characterize this process. Similar to other all-beta proteins, PG shows a non-cooperative fibrillation mechanism, but the structural changes that are monitored by CD indicate a different pattern. Furthermore, several compounds containing aromatic components were tested as potential inhibitors of amyloid formation. Another protein predominantly composed of alpha-helices (human serum albumin) was also targeted by these ligands, in order to get an insight into their potential anti-aggregation property toward structurally different proteins. Among tested compounds, silibinin and chlorpropamide were able to considerably affect both proteins fibrillation process.

Chinisaz, Maryam; Ghasemi, Atiyeh; Larijani, Bagher; Ebrahim-Habibi, Azadeh

2014-02-01

80

Tensile deformation and failure of amyloid and amyloid-like protein fibrils  

Science.gov (United States)

Here we report a series of full atomistic molecular dynamics simulations of six amyloid or amyloid-like protein fibrils in order to systematically understand the effect of different secondary structure motifs on the mechanical tensile and failure response of cross-\\beta protein fibrils. We find a similar failure behavior across the six structures; an initial failure event occurs at small strains involving cooperative rupture of a group of hydrogen bonds, followed by a slow one-by-one hydrogen bond rupture process as the remaining \\beta -sheets peel off with very low applied stress. We also find that the ultimate tensile strength of the protein fibrils investigated scales directly with the number of hydrogen bonds per unit area which break in the initial rupture event. Our results provide insights into structure-property relationships in protein fibrils important for disease and engineering applications and lay the groundwork for the development of materials selection criteria for the design of de novo amyloid-based functional biomaterials.

Solar, Max; Buehler, Markus J.

2014-03-01

 
 
 
 
81

The Hofmeister effect on amyloid formation using yeast prion protein  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A variety of proteins are capable of converting from their soluble forms into highly ordered fibrous cross-? aggregates (amyloids). This conversion is associated with certain pathological conditions in mammals, such as Alzheimer disease, and provides a basis for the infectious or hereditary protein isoforms (prions), causing neurodegenerative disorders in mammals and controlling heritable phenotypes in yeast. The N-proximal region of the yeast prion protein Sup35 (Sup35NM) is frequently used...

Yeh, Victor; Broering, James M.; Romanyuk, Andrey; Chen, Buxin; Chernoff, Yury O.; Bommarius, Andreas S.

2010-01-01

82

Characterization of rat serum amyloid A4 (SAA4): A novel member of the SAA superfamily  

Science.gov (United States)

The serum amyloid A (SAA) family of proteins is encoded by multiple genes, which display allelic variation and a high degree of homology in mammals. The SAA1/2 genes code for non-glycosylated acute-phase SAA1/2 proteins, that may increase up to 1000-fold during inflammation. The SAA4 gene, well characterized in humans (hSAA4) and mice (mSaa4) codes for a SAA4 protein that is glycosylated only in humans. We here report on a previously uncharacterized SAA4 gene (rSAA4) and its product in Rattus norvegicus, the only mammalian species known not to express acute-phase SAA. The exon/intron organization of rSAA4 is similar to that reported for hSAA4 and mSaa4. By performing 5?- and 3?RACE, we identified a 1830-bases containing rSAA4 mRNA (including a GA-dinucleotide tandem repeat). Highest rSAA4 mRNA expression was detected in rat liver. In McA-RH7777 rat hepatoma cells, rSAA4 transcription was significantly upregulated in response to LPS and IL-6 while IL-1?/? and TNF? were without effect. Luciferase assays with promoter-truncation constructs identified three proximal C/EBP-elements that mediate expression of rSAA4 in McA-RH7777 cells. In line with sequence prediction a 14-kDa non-glycosylated SAA4 protein is abundantly expressed in rat liver. Fluorescence microscopy revealed predominant localization of rSAA4-GFP-tagged fusion protein in the ER. PMID:25044109

Rossmann, Christine; Windpassinger, Christian; Brunner, Daniela; Kovacevic, Alenka; Schweighofer, Natascha; Malli, Roland; Schuligoi, Rufina; Prokesch, Andreas; Kluve-Beckerman, Barbara; Graier, Wolfgang F.; Kratky, Dagmar; Sattler, Wolfgang; Malle, Ernst

2014-01-01

83

Characterization of rat serum amyloid A4 (SAA4): a novel member of the SAA superfamily.  

Science.gov (United States)

The serum amyloid A (SAA) family of proteins is encoded by multiple genes, which display allelic variation and a high degree of homology in mammals. The SAA1/2 genes code for non-glycosylated acute-phase SAA1/2 proteins, that may increase up to 1000-fold during inflammation. The SAA4 gene, well characterized in humans (hSAA4) and mice (mSaa4) codes for a SAA4 protein that is glycosylated only in humans. We here report on a previously uncharacterized SAA4 gene (rSAA4) and its product in Rattus norvegicus, the only mammalian species known not to express acute-phase SAA. The exon/intron organization of rSAA4 is similar to that reported for hSAA4 and mSaa4. By performing 5'- and 3'RACE, we identified a 1830-bases containing rSAA4 mRNA (including a GA-dinucleotide tandem repeat). Highest rSAA4 mRNA expression was detected in rat liver. In McA-RH7777 rat hepatoma cells, rSAA4 transcription was significantly upregulated in response to LPS and IL-6 while IL-1?/? and TNF? were without effect. Luciferase assays with promoter-truncation constructs identified three proximal C/EBP-elements that mediate expression of rSAA4 in McA-RH7777 cells. In line with sequence prediction a 14-kDa non-glycosylated SAA4 protein is abundantly expressed in rat liver. Fluorescence microscopy revealed predominant localization of rSAA4-GFP-tagged fusion protein in the ER. PMID:25044109

Rossmann, Christine; Windpassinger, Christian; Brunner, Daniela; Kovacevic, Alenka; Schweighofer, Natascha; Malli, Roland; Schuligoi, Rufina; Prokesch, Andreas; Kluve-Beckerman, Barbara; Graier, Wolfgang F; Kratky, Dagmar; Sattler, Wolfgang; Malle, Ernst

2014-08-01

84

How amyloid precursor protein protects itself from cleavage.  

Science.gov (United States)

In this issue of Structure, Tang and colleagues probe how the Flemish mutation in amyloid precursor protein (APP) affects its conformation and cleavage by ?-secretase. They provide molecular insight into how an extracellular inhibitory element and cholesterol interactions affect the generation of A? peptides. PMID:24607141

Lin, Hsiang-Kai; van der Wel, Patrick C A

2014-03-01

85

Establishment of a transgenic mouse model specifically expressing human serum amyloid A in adipose tissue.  

Science.gov (United States)

Obesity and obesity co-morbidities are associated with a low grade inflammation and elevated serum levels of acute phase proteins, including serum amyloid A (SAA). In the non-acute phase in humans, adipocytes are major producers of SAA but the function of adipocyte-derived SAA is unknown. To clarify the role of adipocyte-derived SAA, a transgenic mouse model expressing human SAA1 (hSAA) in adipocytes was established. hSAA expression was analysed using real-time PCR analysis. Male animals were challenged with a high fat (HF) diet. Plasma samples were subjected to fast protein liquid chromatography (FPLC) separation. hSAA, cholesterol and triglyceride content were measured in plasma and in FPLC fractions. Real-time PCR analysis confirmed an adipose tissue-specific hSAA gene expression. Moreover, the hSAA gene expression was not influenced by HF diet. However, hSAA plasma levels in HF fed animals (37.7±4.0 µg/mL, n?=?7) were increased compared to those in normal chow fed animals (4.8±0.5 µg/mL, n?=?10; p<0.001), and plasma levels in the two groups were in the same ranges as in obese and lean human subjects, respectively. In FPLC separated plasma samples, the concentration of hSAA peaked in high-density lipoprotein (HDL) containing fractions. In addition, cholesterol distribution over the different lipoprotein subfractions as assessed by FPLC analysis was similar within the two experimental groups. The established transgenic mouse model demonstrates that adipose tissue produced hSAA enters the circulation, resulting in elevated plasma levels of hSAA. This new model will enable further studies of metabolic effects of adipose tissue-derived SAA. PMID:21611116

Olsson, Maja; Ahlin, Sofie; Olsson, Bob; Svensson, Per-Arne; Ståhlman, Marcus; Borén, Jan; Carlsson, Lena M S; Sjöholm, Kajsa

2011-01-01

86

AMYPdb: A database dedicated to amyloid precursor proteins  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Misfolding and aggregation of proteins into ordered fibrillar structures is associated with a number of severe pathologies, including Alzheimer's disease, prion diseases, and type II diabetes. The rapid accumulation of knowledge about the sequences and structures of these proteins allows using of in silico methods to investigate the molecular mechanisms of their abnormal conformational changes and assembly. However, such an approach requires the collection of accurate data, which are inconveniently dispersed among several generalist databases. Results We therefore created a free online knowledge database (AMYPdb dedicated to amyloid precursor proteins and we have performed large scale sequence analysis of the included data. Currently, AMYPdb integrates data on 31 families, including 1,705 proteins from nearly 600 organisms. It displays links to more than 2,300 bibliographic references and 1,200 3D-structures. A Wiki system is available to insert data into the database, providing a sharing and collaboration environment. We generated and analyzed 3,621 amino acid sequence patterns, reporting highly specific patterns for each amyloid family, along with patterns likely to be involved in protein misfolding and aggregation. Conclusion AMYPdb is a comprehensive online database aiming at the centralization of bioinformatic data regarding all amyloid proteins and their precursors. Our sequence pattern discovery and analysis approach unveiled protein regions of significant interest. AMYPdb is freely accessible 1.

Delamarche Christian

2008-06-01

87

Characterization of serum amyloid A (SAA) in rainbow trout using a new monoclonal antibody  

DEFF Research Database (Denmark)

Serum amyloid A (SAA) is an integral part of the innate immune response in mammals and considered to be important during the acute phase response. The present study was undertaken to elucidate the role of SAA protein in the innate immune response of rainbow trout. A monoclonal antibody raised against a recombinant peptide of rainbow trout SAA was characterized using Western blot, dot blot, ELISA and immunohistochemistry. SAA association with high density lipoprotein (HDL) complicated band identification in Western blot, but delipidization of the SAA-HDL isolate highly increased the quality of reaction in the western blot. Rainbow trout fry (87 days post hatch) infected with Yersinia ruckeri showed a significant up-regulation of the SAA gene at 72 h post infection with an increase until 96 h post infection. Non-significant up-regulations were seen at earlier time points i.e. 4 and 24 h. The expression pattern of SAA significantly correlated to the immunohistochemical analysis of the infected fry. A weak staining was seen in liver tissue at 4 h post infection which increased in intensity during the course of infection i.e. 24, 72 and 96 h post infection.

Kania, Per Walter; Chettri, Jiwan K

2014-01-01

88

Serum amyloid P ameliorates radiation-induced oral mucositis and fibrosis  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Purpose To evaluate the effect of the anti-fibrotic protein serum amyloid P (SAP on radiation-induced oral mucositis (OM and fibrosis in a hamster cheek-pouch model. Experimental Design Hamsters received a single dose of radiation (40 Gy to the left everted cheek pouch to induce significant OM. The protective therapeutic potential of SAP was evaluated using varying dosing regimens. The extent of OM was measured using a validated six-point scoring scheme ranging from 0 (normal tissue, no mucositis to 5 (complete ulceration. Fibrotic remodeling was also visualized histologically and quantified at later time points using collagen gene expression. Results SAP treatment attenuated the profile of radiation-induced oral mucositis by delaying the time of onset, reducing the peak value, and enhancing the resolution of injury. The peak mucositis score was reduced by approximately 0.5 grade in SAP-treated animals. The number of animal days with a score of ? 3 was reduced by 48% in the SAP-treated group, compared with the saline control group (P Conclusions SAP treatment significantly attenuated radiation-induced injury. In particular, SAP attenuated the severity of OM and inhibited pathogenic remodeling. This suggests that SAP may be a useful therapy for the palliation of side effects observed during treatment for head and neck cancer.

Murray Lynne A

2010-07-01

89

Enhanced degradation of amyloid AA proteins by enzyme activation: a possible model for a therapeutic approach  

International Nuclear Information System (INIS)

The capacity of plasminogen from human serum to degrade amyloid AA protein was tested using radiolabelled protein AA coupled to cyanogen bromide activated Sepharose 6 MB as substrate. Protein AA degrading activity was determined in fractions of normal human serum separated by Sephadex G 150. Each fraction was tested in the presence and absence of the plasminogen activator streptokinase. The AA degrading activity was markedly increased in fractions in which plasminogen activation had occurred. These fractions were also the same as those showing the presence of plasminogen as demonstrated by reaction with a specific anti-plasminogen antiserum. Moreover, the increase in AA degrading activity could be inhibited with antibodies to plasminogen. AA degrading activity could also be enhanced in whole human plasma by streptokinase activation

90

Serum amyloid A promotes osteosarcoma invasion via upregulating ?v?3 integrin.  

Science.gov (United States)

Serum amyloid A (SAA) is regarded as an important acute phase protein involved in tumor progression and metastasis. However, at present there is no evidence of its involvement in osteosarcoma. The present study aimed to investigate the effect of SAA on the invasion of osteosarcoma cells. The effects of SAA on the migration and invasion of osteosarcoma cells were detected using scratch wound healing and transwell assays, respectively. The expression of ?v?3 integrin was detected at the protein and mRNA levels in U2OS cells. Agonists, inhibitors or siRNA of formyl peptide receptor like?1 (FPRL?1), mitogen?activated protein kinases and ?v?3 integrin were used to investigate the mechanism underlying the effects of SAA on the regulation of U2OS cell migration and invasion. The present study revealed that SAA promoted osteosarcoma cell migration and invasion. SAA upregulated the expression of ?v?3 integrin in a concentration? and time?dependent manner. When inhibiting ?v?3 integrin with its antagonist, the migration and invasion abilities of the U2OS cells were markedly inhibited. SAA?induced ?v?3 integrin production was significantly downregulated by inhibiting FPRL?1 with siRNA and inhibitors. The present study also found that extracellular signal?regulated kinase (ERK) 1/2, but not c?Jun N?terminal kinase or p38, was important in this process. These findings demonstrated that SAA regulated osteosarcoma cell migration and invasion via the FPRL?1/ERK/?v?3 integrin pathway. PMID:25323768

Ren, Peng; Sun, Deshun; Xin, Dajiang; Ma, Wanli; Chen, Peng; Gao, Hongwei; Zhang, Shouqiang; Gong, Mingzhi

2014-12-01

91

The effect of corticosteroids on amyloid beta precursor protein/amyloid precursor-like protein expression and processing in vivo.  

Science.gov (United States)

In this study, we have investigated the effect of altered corticosteroid levels on the expression and processing of the amyloid beta precursor protein (A betaPP) and its amyloid precursor-like protein (APLP) homologue in rat brain. Four groups of animals were used in the study: sham operated, adrenalectomised, and adrenalectomised treated with either dexamethasone or aldosterone, with the A betaPP/APLP expression being determined by western blot analysis. While there were no changes in the levels of A betaPP/APLP following adrenalectomy, treatment with dexamethasone, but not aldosterone, resulted in a marked increase in protein expression levels with the level of increase varying between the brain regions examined. Corticosteroids had a more marked effect on the particulate rather than the soluble form of the protein, thus suggesting that elevated glucocorticoids may also be adversely influencing A betaPP/APLP processing. PMID:10586975

Budas, G; Coughlan, C M; Seckl, J R; Breen, K C

1999-11-26

92

Scintigraphic imaging and turnover studies with iodine-131 labelled serum amyloid P component in systemic amyloidosis  

International Nuclear Information System (INIS)

Radiolabelled serum amyloid P component (SAP) is a specific tracer for amyloid. Iodine-123 has ideal physical characteristics for scintigraphy but is expensive and not widely available. Here we report serial imaging and turnover studies in which we labelled SAP with iodine-131, a cheap alternative isotope which would be expected to yield poorer images but permit more prolonged turnover measurements. Imaging and plasma clearance and whole body retention (WBR) of tracer were studied for up to 7 days in ten patients with proven systemic AL amyloidosis and two patients in whom the diagnosis was suspected, after i.v. administration of about 37 MBq of 131I-SAP. Normal blood pool images were obtained in the latter two subjects and amyloidosis was subsequently refuted histologically. WBR at 48 h was 65% of the injected dose (i.d.). Among the other ten patients, amyloid deposits were identified in the spleen in eight cases, liver in five and kidneys in four; other sites that gave positive results included bone, joints and soft tissues, and the myocardium in one case. Up to 95% of the tracer localised into amyloid within 6-h, and the values for WBR became progressively more discriminating during the study period, exceeding the normal reference value (131I-SAP produced diagnostic scans in every patient in this series and, coupled with the detailed turnover information, is adequate for monitoring disease progress. (orig.)

93

Cellular prion protein regulates ?-secretase cleavage of the Alzheimer's amyloid precursor protein  

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Proteolytic processing of the amyloid precursor protein (APP) by ?-secretase, ?-site APP cleaving enzyme (BACE1), is the initial step in the production of the amyloid ? (A?) peptide, which is involved in the pathogenesis of Alzheimer's disease. The normal cellular function of the prion protein (PrPC), the causative agent of the transmissible spongiform encephalopathies such as Creutzfeldt–Jakob disease in humans, remains enigmatic. Because both APP and PrPC are subject to proteolytic pr...

Parkin, Edward T.; Watt, Nicole T.; Hussain, Ishrut; Eckman, Elizabeth A.; Eckman, Christopher B.; Manson, Jean C.; Baybutt, Herbert N.; Turner, Anthony J.; Hooper, Nigel M.

2007-01-01

94

Importance of the caspase cleavage site in amyloid-? protein precursor.  

Science.gov (United States)

Reports from multiple laboratories have now been published analyzing the critical nature of the caspase cleavage site of amyloid-? protein precursor (A?PP) for cell death induction, synaptic loss, hippocampal atrophy, long-term potentiation, memory loss, neophobia, and other aspects of the Alzheimer's phenotype. Here we review the results and implications of these studies for the understanding of Alzheimer's disease pathophysiology and the potential development of therapeutics that target this site in A?PP. PMID:20847422

Bredesen, Dale E; John, Varghese; Galvan, Veronica

2010-01-01

95

Protein Polymers and Amyloids : Focus on ?1-Antitrypsin  

DEFF Research Database (Denmark)

Several human disorders are caused by a common general disease mechanism arising from abnormal folding and aggregation of the underlying protein. These include the prevalent dementias like Alzheimer’s and Parkinson’s, where accumulation of protein fibrillar structures, known as amyloid fibrils, is a general hallmark. They also include the ?1-antitrypsin deficiency, where disease-causing mutations in the serine protease inhibitor, ?1-antitrypsin (?1AT), leads to accumulation of the aberrant protein in the liver of these patients. The native metastable structure of ?1AT constitutes a molecular trap that inhibits its target protease through a large conformational change but mutations compromise this function and cause premature structural collapse into hyperstable polymers. Understanding the conformational disorders at a molecular level is not only important for our general knowledge on protein folding and misfolding but also for rationalizing efficient therapeutic strategies to ameliorate the associated disease. In this work, we focussed on the C-terminal part of ?1AT to understand its role in the disease-causing polymerization events and to investigate the amyloid fibril formation of a proteolytically generated fragment from here. To enable a detailed structural analysis by Nuclear Magnetic Resonance Spectroscopy an in vitro ligation procedure was established that reconstituted ?1AT from two separate fragments. In this way, it would be possible to incorporate NMR-active isotopes in the C-terminal part selectively. Extensive biochemical work established successful expressed protein ligation for two separate ligation joints in ?1AT and provided proof-of-concept for the strategy. The polymerization of ?1AT can happen trough the insertion of the C-terminal tail into the succeeding molecule and features of this mechanism were investigated through a series of interaction experiments. Despite a very buried location in the native structure, evidence here suggest that the C-terminal tail is labile under slightly destabilizing conditions, providing new detail to this matter. A small infectious polymer unit was also constructed and used to show how polymerogenic seeding and polymer propagation might happen inside the body. The locking of central structural elements during ?1AT folding or in the native state represents a therapeutic strategy to prevent polymerization. Using Molecular Dynamics simulations, we identified a new druggable pocket on the surface of ?1AT that could be targeted to serve this purpose. The proteolytically generated C-terminal tail from ?1AT is 36 residues long (C- 36) and is present in various bodily fluids. The peptide is able to form amyloid fibrils and we provide the first characterization of the fibrillation mechanism and of the amyloid structures that arise. The fibrillation is greatly enhanced by the presence of the anionic heparin sugar chain and we establish a model to describe these effects. Such negatively charged sugar molecules are ubiquitously associated with amyloid deposits in vivo, underlining the importance of understanding this relationship. The monomeric C-36 peptide was investigated by liquid-state NMR spectroscopy and found to be intrinsically disordered with minor propensities towards ?-sheet structure. The plasticity of such a peptide makes it suitable for a whole range of interactions, including its conversion to the tightly packed repetitive ?-sheet arrangement of an amyloid fibre. The ultra-structural details of these fibres were established by electron microscopy and solid-state NMR was employed to resolve the underlying molecular structure. This last task has not been completed yet but solving such structures to atomic resolution is of high value for understanding and targeting the culprits of the amyloid-related conformational disorders. Lastly, this work also includes a study on the protease HtrA1 that localizes to certain amyloid plaques. We explore the mechanism behind its automaturation process and find it to depend on the integrity of a disulphide bond network

RisØr, Michael Wulff

2014-01-01

96

A carbon nanotube metal semiconductor field effect transistor-based biosensor for detection of amyloid-beta in human serum.  

Science.gov (United States)

We have developed a carbon nanotube (CNT) film-based biosensor with a metal semiconductor field effect transistor structure (MESFET). A gold top gate was deposited on the middle of the CNT channel and probe antibodies were immobilized on the gold top gate with an antibody-binding protein, protein G or Escherichia coli outer membrane (OM) with autodisplayed Z-domains of protein A. These CNT-MESFET biosensors exhibited a higher sensitivity than the CNT-FET biosensor with probe antibodies immobilized using a chemical linker, since the orientation of immobilized antibodies was controlled by the antibody-binding proteins. In addition, nonspecific binding was effectively inhibited by E. coli OM. Using the CNT-MESFET biosensors with E. coli OM containing Z domain, we detected amyloid-? (A?) in human serum, one of the biomarkers for early diagnosis of Alzheimer's disease. A? at the level of 1 pg/mL in human serum could be measured in real-time and without labeling, which was lower than a limit of detection for plasma A? using an enzyme-linked immune sorbent assay. These results suggested that our CNT-MESFET biosensors might be applicable for an early diagnosis of Alzheimer's disease. PMID:23891796

Oh, Jeseung; Yoo, Gu; Chang, Young Wook; Kim, Hyung Joon; Jose, Joachim; Kim, Eosu; Pyun, Jae-Chul; Yoo, Kyung-Hwa

2013-12-15

97

Administration of perioperative penicillin reduces postoperative serum amyloid A response in horses being castrated standing  

DEFF Research Database (Denmark)

Objectives: To compare postoperative in?ammatory responses in horses administered perioperative procaine penicillin and those not administered penicillin using acute phase protein serum amyloid A (SAA) as a marker of in?ammation. Study Design: Randomized clinical trial. Animals: Stallions (n = 50) castrated under ?eld conditions. Methods: SAA concentrations were determined on days 0, 3, and 8. Six horses were subsequently excluded because of elevated SAA concentrations on day 0. Of the remaining 50 horses, 26 were administered nonsteroidal anti-in?ammatory drug (NSAID) therapy and 24 were administered NSAID and 25,000 U/kg procaine penicillin on day 0, 1, and 2. Results: SAA concentrations increased signi?cantly from preoperative levels in both groups, and on day 8 concentrations were signi?cantly (P o .02) higher in horses administered only NSAID than in those administered procaine penicillin and NSAID. Infectious complications occurred more frequently (P o .01) in horses with preoperatively elevated SAA concentrations (the excluded horses) than in horses with normal preoperative SAA concentrations (the included horses). Conclusions: Perioperative antimicrobial therapy reduced the postoperative SAA response, suggesting that bacteria were present in the surgical wound and contributed to in?ammation after castration. Horses with elevated preoperative SAA concentrations developed infectious complications more often than horses with normal preoperative SAA concentrations. Clinical Relevance: Administration of antimicrobials may be important in horses being castrated standing under ?eld conditions. Increased SAA concentrations seem to be an indicator of increased surgical risk in horses and may be useful before elective surgery for planning.

Busk, Peter; Jacobsen, Stine

2010-01-01

98

Serum Amyloid A in the Placenta and Its Role in Trophoblast Invasion  

Science.gov (United States)

The serum amyloid A (SAA) protein is known to function in the acute phase response and immunoregulation. Recently, SAA has been shown to be involved in cell proliferation, differentiation and migratory behavior in different cell types. Here, we evaluated whether exogenous SAA could influence trophoblast invasion and differentiation using both the trophoblast-like BeWo cell line and fully differentiated human extravillous trophoblast cells (EVT) isolated from term placentae. SAA stimulated BeWo cell invasion, as measured in Matrigel invasion assays, and induced metalloprotease mRNA expression and activity. Given that BeWo cells express Toll-like receptor 4 (TLR4), a known receptor for SAA, we examined the role of TLR4 in SAA-induced invasion using a TLR4 neutralizing antibody. We also tested whether SAA could affect markers of trophoblast syncytialization in BeWo cells. We observed that SAA decreased ?hCG secretion and did not influence trophoblast syncytialization. Using EVT cells isolated from human term basal plates, we confirmed that SAA at 1 and 10 µg/mL doubled EVT invasion in a TLR4-dependent manner, but at 20 µg/mL inhibited EVT cells invasiveness. In addition, we observed that SAA was expressed in both BeWo cells and human term placentae, specifically in the syncytiotrophoblast, decidual cells and EVT. In conclusion, SAA was identified as a molecule that functions in the placental microenvironment to regulate metalloprotease activity and trophoblast invasion, which are key processes in placentation and placental homeostasis. PMID:24614130

Sandri, Silvana; Urban Borbely, Alexandre; Fernandes, Isabella; Mendes de Oliveira, Edson; Knebel, Franciele Hinterholz; Ruano, Rodrigo; Zugaib, Marcelo; Filippin-Monteiro, Fabiola; Bevilacqua, Estela; Campa, Ana

2014-01-01

99

Protein misfolding, congophilia, oligomerization, and defective amyloid processing in preeclampsia.  

Science.gov (United States)

Preeclampsia is a pregnancy-specific disorder of unknown etiology and a leading contributor to maternal and perinatal morbidity and mortality worldwide. Because there is no cure other than delivery, preeclampsia is the leading cause of iatrogenic preterm birth. We show that preeclampsia shares pathophysiologic features with recognized protein misfolding disorders. These features include urine congophilia (affinity for the amyloidophilic dye Congo red), affinity for conformational state-dependent antibodies, and dysregulation of prototype proteolytic enzymes involved in amyloid precursor protein (APP) processing. Assessment of global protein misfolding load in pregnancy based on urine congophilia (Congo red dot test) carries diagnostic and prognostic potential for preeclampsia. We used conformational state-dependent antibodies to demonstrate the presence of generic supramolecular assemblies (prefibrillar oligomers and annular protofibrils), which vary in quantitative and qualitative representation with preeclampsia severity. In the first attempt to characterize the preeclampsia misfoldome, we report that the urine congophilic material includes proteoforms of ceruloplasmin, immunoglobulin free light chains, SERPINA1, albumin, interferon-inducible protein 6-16, and Alzheimer's ?-amyloid. The human placenta abundantly expresses APP along with prototype APP-processing enzymes, of which the ?-secretase ADAM10, the ?-secretases BACE1 and BACE2, and the ?-secretase presenilin-1 were all up-regulated in preeclampsia. The presence of ?-amyloid aggregates in placentas of women with preeclampsia and fetal growth restriction further supports the notion that this condition should join the growing list of protein conformational disorders. If these aggregates play a pathophysiologic role, our findings may lead to treatment for preeclampsia. PMID:25031267

Buhimschi, Irina A; Nayeri, Unzila A; Zhao, Guomao; Shook, Lydia L; Pensalfini, Anna; Funai, Edmund F; Bernstein, Ira M; Glabe, Charles G; Buhimschi, Catalin S

2014-07-16

100

Lysine 624 of the Amyloid Precursor Protein (APP) Is a Critical Determinant of Amyloid ? Peptide Length  

Science.gov (United States)

?-Secretase is a multiprotein intramembrane cleaving aspartyl protease (I-CLiP) that catalyzes the final cleavage of the amyloid ? precursor protein (APP) to release the amyloid ? peptide (A?). A? is the primary component of senile plaques in Alzheimer's disease (AD), and its mechanism of production has been studied intensely. ?-Secretase executes multiple cleavages within the transmembrane domain of APP, with cleavages producing A? and the APP intracellular domain (AICD), referred to as ? and ?, respectively. The heterogeneous nature of the ? cleavage that produces various A? peptides is highly relevant to AD, as increased production of A? 1–42 is genetically and biochemically linked to the development of AD. We have identified an amino acid in the juxtamembrane region of APP, lysine 624, on the basis of APP695 numbering (position 28 relative to A?) that plays a critical role in determining the final length of A? peptides released by ?-secretase. Mutation of this lysine to alanine (K28A) shifts the primary site of ?-secretase cleavage from 1–40 to 1–33 without significant changes to ? cleavage. These results further support a model where ? cleavage occurs first, followed by sequential proteolysis of the remaining transmembrane fragment, but extend these observations by demonstrating that charged residues at the luminal boundary of the APP transmembrane domain limit processivity of ?-secretase. PMID:21868378

Kukar, Thomas L.; Ladd, Thomas B.; Robertson, Paul; Pintchovski, Sean A.; Moore, Brenda; Bann, Maralyssa A.; Ren, Zhao; Jansen-West, Karen; Malphrus, Kim; Eggert, Simone; Maruyama, Hiroko; Cottrell, Barbara A.; Das, Pritam; Basi, Guriqbal S.; Koo, Edward H.; Golde, Todd E.

2011-01-01

 
 
 
 
101

Structural and Functional Alterations in Amyloid-? Precursor Protein Induced by Amyloid-? Peptides  

Science.gov (United States)

Alzheimer's disease-associated amyloid-? (A?) peptide is neurotoxic as an oligomer, but not as a monomer, by an unknown mechanism. We showed previously that A? interacts with the amyloid-? precursor protein (A?PP), leading to caspase cleavage and cell death induction. To characterize this structure and interaction further, we purified the extracellular domain of A?PP695 (eA?PP) and its complex with A? oligomers (A?Os) of varying sizes, and then performed small angle X-ray scattering (SAXS). In the absence of any A?, eA?PP was a compact homodimer with a tight association between the E1 and E2 domains. Dimeric A? oligomers induced monomerization of eA?PP while larger oligomers also bound eA?PP but preserved the homodimer. Efficient binding of the larger oligomers correlated with the presence of prefibrillar oligomers, suggesting that the eA?PP binding is limited to a conformational subset of A? oligomers. Both forms of A? bound to eA?PP at the A?-cognate region and induced dissociation of the E1 and E2 domains. Our data provide the first structural evidence for A?-A?PP binding and suggest a mechanism for differential modulation of A?PP processing and cell death signaling by A? dimers versus conformationally-specific larger oligomers. PMID:21471643

Libeu, Clare Peters; Poksay, Karen S.; John, Varghese; Bredesen, Dale E.

2014-01-01

102

Structural and functional alterations in amyloid-? precursor protein induced by amyloid-? peptides.  

Science.gov (United States)

Alzheimer's disease-associated amyloid-? (A?) peptide is neurotoxic as an oligomer, but not as a monomer, by an unknown mechanism. We showed previously that A? interacts with the amyloid-? precursor protein (A?PP), leading to caspase cleavage and cell death induction. To characterize this structure and interaction further, we purified the extracellular domain of A?PP695 (eA?PP) and its complex with A? oligomers (A?Os) of varying sizes, and then performed small angle X-ray scattering (SAXS). In the absence of any A?, eA?PP was a compact homodimer with a tight association between the E1 and E2 domains. Dimeric A? oligomers induced monomerization of eA?PP while larger oligomers also bound eA?PP but preserved the homodimer. Efficient binding of the larger oligomers correlated with the presence of prefibrillar oligomers, suggesting that the eA?PP binding is limited to a conformational subset of A? oligomers. Both forms of A? bound to eA?PP at the A?-cognate region and induced dissociation of the E1 and E2 domains. Our data provide the first structural evidence for A?-A?PP binding and suggest a mechanism for differential modulation of A?PP processing and cell death signaling by A? dimers versus conformationally-specific larger oligomers. PMID:21471643

Libeu, Clare Peters; Poksay, Karen S; John, Varghese; Bredesen, Dale E

2011-01-01

103

The role of multivesicular bodies in the trafficking of the amyloid precursor protein  

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In Alzheimer’s disease, the beta-amyloid (A?) protein accumulates within neurons prior to amyloid plaque development. Recent evidence suggests that intracellular A? accumulation, rather than extracellular plaques, correlates best with disease progression. A? is generated by proteolytic processing of the amyloid precursor protein (APP) but the intracellular site of A? production is unclear. A? has been localised to multivesicular endosomes/bodies (MVBs), which have many cellular functio...

Edgar, J.

2013-01-01

104

Nerve growth factor and ras regulate ?-amyloid precursor protein gene expression in PC12 cells  

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The ?-amyloid protein, the major component of the vascular and plaque amyloid deposits that characterize Alzheimer's disease, derives from a larger ?-amyloid precursor protein (APP) that is expressed in both neural and nonneural cells. An increased expression of APP might actively contribute to the development of the pathology; however, the mechanisms involved in the regulation of APP gene expression are not yet well understood. In PC12 cells, a rat pheochromocytoma cell line, we have demon...

Cosgaya, Jose? Miguel; Latasa, Mari?a Jesu?s; Pascual, A?ngel

1996-01-01

105

Validación analítica de técnicas comerciales para la determinación de haptoglobina, proteína C reactiva y amiloide A sérico en caninos Analytical / validation of commercial assays for the determination of haptoglobin, C-reactive protein and serum amyloid A in dogs  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: Spanish Abstract in spanish Los métodos analíticos deben ser validados antes de emplearlos de forma rutinaria en un laboratorio. El objetivo de este trabajo fue validar tres métodos analíticos comerciales usados en nuestro laboratorio para la determinación de haptoglobina (Hp), proteína C reactiva (CRP) y amiloide A sérico (SA [...] A) en muestras de la especie canina con concentraciones bajas y altas de proteínas de fase aguda (PFAs). Los parámetros que se evaluaron para la validación fueron: (1) Precisión, determinada mediante el cálculo de los coeficientes de variación (CVs) intra e interdeterminación. (2) Exactitud, evaluada indirectamente comprobando la linealidad bajo dilución. (3) Límite de detección, determinado como la mínima concentración que puede distinguirse de una muestra de valor 0. Todos los CVs intradeterminación fueron Abstract in english All laboratory tests must be validated before being introduced for patient testing. The objective of this work was to perform the analytical validation of three commercial assays that are being used at our laboratory for the determination of haptoglobin (Hp), C reactive protein (CRP) and serum amylo [...] id A (SAA) in canine samples with low and high concentrations of these acute phase proteins (APPs). The parameters evaluated for the validation of the methods were: (1) Precision, assessed by determination of the within and between-run coefficients of variation (CVs). (2) Inaccuracy, evaluated indirectly by investigating linearity under dilution. (3) Limit of detection, determined as the lowest concentration of the APPs which could be distinguished from a zero sample. All within-run CVs were lower than 10%, however between-run CVs were lower than 10% only for Hp. Dilution of a serum sample with high concentrations of the different APPs resulted in a linear regression equation with correlation coefficient R²0.98 in all cases; so all methods showed a good accuracy. The detection limit of each assay was 0.02 g/L for Hp, 0.15 mg/L for CRP and 0.79 mg/L for SAA. Additionally they differentiate animals with inflammatory or infectious diseases from healthy subjects. Overall results of validation showed that the assays tested can be suitable for the routine measurement of APPs in canine samples, although it would be desirable to reduce the between run imprecision found for CRP and SAA assays.

S., Martínez-Subiela; J. J., Cerón.

106

Investigation of the solubility and the potentials for purification of serum amyloid A (SAA) from equine acute phase serum : a pilot study  

DEFF Research Database (Denmark)

BACKGROUND: Serum amyloid A (SAA) is useful as a diagnostic marker of systemic inflammation in horses, but only heterologous assays based on non-equine calibration and standardization are available for measurements of equine SAA. More accurate measurements could be obtained using purified species-specific SAA in native conformation for assay calibration and standardization. Further knowledge about the biochemical properties of SAA would facilitate a future production of native species-specific calibration material Therefore, the aim of the study was an investigation of the solubility and potentials for purification of equine SAA based on biochemical properties.Freeze dried equine acute phase serum was dissolved in 70% 2-propanol, 8 M urea, and milli-Q water, respectively. Supercritical fluid extraction (SFE), size-exclusive chromatography (FPLC-SEC), and preparative isoelectric focusing (IEF) were performed in the attempt to purify. Immunostaining of IEF blots were used for isoform-specific detection of SAA in the preparations and purity was assessed by silverstained SDS-PAGE. FINDINGS: SAA was soluble in 70% 2-propanol, 8 M urea and Milli-Q water. SAA was not separated in the lipophilic or ampipathic fractions following SFE. SAA was included in a FPLC-SEC-fraction of 237 kDa, despite the molecular weight known to be much smaller, suggesting binding to other serum constituents. SAA precipitated following separation of other serum proteins by preparative IEF. DISCUSSION: No effective purification of SAA was achieved in the present study, but findings important for future investigations were made. The study suggested that SAA is not exclusively hydrophobic, but appears less hydrophobic when interacting with other serum components. These results suggest more complex aspects of solubility than previously believed, and indicate potentials for purification of native SAA.

Christensen, Michelle BrØnniche; SØrensen, Jens Christian

2013-01-01

107

Ichthyophthirius multifiliis infection induces massive up-regulation of serum amyloid A in carp (Cyprinus carpio)  

DEFF Research Database (Denmark)

A real time quantitative PCR (RQ-PCR) assay was developed for measurement of differential expression of the genes encoding the acute phase reactant serum amyloid A (SAA), transferrin (TF) and a C-type lectin molecule (CL) in skin, blood and liver from Cyprinus carpio following infection with the ectoparasite Ichthyophthirius multifiliis. Serum amyloid A and CL were constitutively expressed in all organs evaluated while TF transcripts were only detected in the liver. A dramatic up-regulation (1600 times) in the expression levels of SAA was observed in skin 36 h after the parasite infection. A similar increase in the number of RNA molecules encoding for SAA was observed in the liver. The CL expression was significantly down regulated in all the organs and no significant change was observed in the expression levels of the TF in the liver. These results indicate that SAA plays a major role in the acute phase response in fish infected with I. multififiis and emphasize the importance of the fish skin as an active organ in response to an ectoparasite infection. (c) 2006 Elsevier B.V. All rights reserved.

Gonzales, S.F.; Buchmann, Kurt

2007-01-01

108

Amyloid precursor protein processing is stimulated by metabotropic glutamate receptors.  

Science.gov (United States)

Stimulation of muscarinic m1 or m3 receptors can, by generating diacylglycerol and activating protein kinase C, accelerate the breakdown of the amyloid precursor protein (APP) to form soluble, nonamyloidogenic derivatives (APPs), as previously shown. This relationship has been demonstrated in human glioma and neuroblastoma cells, as well as in transfected human embryonic kidney 293 cells and PC-12 cells. We now provide evidence that stimulation of metabotropic glutamate receptors (mGluRs), which also are coupled to phosphatidylinositol 4,5-bisphosphate hydrolysis, similarly accelerates processing of APP into nonamyloidogenic APPs. This process is demonstrated both in hippocampal neurons derived from fetal rats and in human embryonic kidney 293 cells transfected with cDNA expression constructs encoding the mGluR 1 alpha subtype. In hippocampal neurons, both an mGluR antagonist, L-(+)-2-amino-3-phosphonopropionic acid, and an inhibitor of protein kinase C, GF 109203X, blocked the APPs release evoked by glutamate receptor stimulation. Ionotropic glutamate agonists, N-methyl-D-aspartate or S(-)-5-fluorowillardiine, failed to affect APPs release. These data show that selective mGluR agonists that initiate signal-transduction events can regulate APP processing in bona fide primary neurons and transfected cells. As glutamatergic neurons in the cortex and hippocampus are damaged in Alzheimer disease, amyloid production in these regions may be enhanced by deficits in glutamatergic neurotransmission. PMID:7644542

Lee, R K; Wurtman, R J; Cox, A J; Nitsch, R M

1995-08-15

109

Regulation of Major Histocompatibility Complex Class I Molecule Expression on Cancer Cells by Amyloid Precursor-Like Protein 2  

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The three members of the amyloid precursor protein family in mammals (amyloid precursor protein, amyloid precursor-like protein-1, and amyloid precursor-like-protein-2 (APLP2)) have been implicated in a large array of intracellular processes, which include development, transcription, apoptosis, metabolism, and the cell cycle. A series of studies by our laboratories has demonstrated that APLP2 is highly expressed by many cancer cell lines (with the highest expression in pancreatic cancer cell ...

Peters, Haley L.; Tuli, Amit; Sharma, Mahak; Naslavsky, Naava; Caplan, Steve; Macdonald, Richard G.; Solheim, Joyce C.

2011-01-01

110

Novel neuritic clusters with accumulations of amyloid precursor protein and amyloid precursor-like protein 2 immunoreactivity in brain regions damaged by thiamine deficiency.  

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Experimental thiamine deficiency (TD) is a classical model of a nutritional deficit associated with a generalized impairment of oxidative metabolism and selective cell loss in the brain. In rats, TD-induced cell degeneration is accompanied by an accumulation of amyloid precursor protein (APP)/amyloid precursor-like protein 2 (APLP2) immunoreactivity in abnormal neurites and perikarya along the periphery of, or scattered within, the lesion. Prompted by these data and our previous findings of a...

Calingasan, N. Y.; Gandy, S. E.; Baker, H.; Sheu, K. F.; Smith, J. D.; Lamb, B. T.; Gearhart, J. D.; Buxbaum, J. D.; Harper, C.; Selkoe, D. J.; Price, D. L.; Sisodia, S. S.; Gibson, G. E.

1996-01-01

111

Herpes simplex virus interferes with amyloid precursor protein processing  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The early events underlying Alzheimer's disease (AD remain uncertain, although environmental factors may be involved. Work in this laboratory has shown that the combination of herpes simplex virus type 1 (HSV1 in brain and carriage of the APOE-?4 allele of the APOE gene strongly increases the risk of developing AD. The development of AD is thought to involve abnormal aggregation or deposition of a 39–43 amino acid protein – ? amyloid (A? – within the brain. This is cleaved from the much larger transmembranal protein 'amyloid precursor protein' (APP. Any agent able to interfere directly with A? or APP metabolism may therefore have the capacity to contribute towards AD. One recent report showed that certain HSV1 glycoprotein peptides may aggregate like A?; a second study described a role for APP in transport of virus in squid axons. However to date the effects of acute herpesvirus infection on metabolism of APP in human neuronal-type cells have not been investigated. In order to find if HSV1 directly affects APP and its degradation, we have examined this protein from human neuroblastoma cells (normal and transfected with APP 695 infected with the virus, using Western blotting. Results We have found that acute HSV1 (and also HSV2 infection rapidly reduces full length APP levels – as might be expected – yet surprisingly markedly increases levels of a novel C-terminal fragment of APP of about 55 kDa. This band was not increased in cells treated with the protein synthesis inhibitor cycloheximide Conclusion Herpes virus infection leads to rapid loss of full length APP from cells, yet also causes increased levels of a novel 55 kDa C-terminal APP fragment. These data suggest that infection can directly alter the processing of a transmembranal protein intimately linked to the aetiology of AD.

Itzhaki Ruth F

2005-08-01

112

Scintigraphic imaging and turnover studies with iodine-131 labelled serum amyloid P component in systemic amyloidosis  

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Radiolabelled serum amyloid P component (SAP) is a specific tracer for amyloid. Iodine-123 has ideal physical characteristics for scintigraphy but is expensive and not widely available. Here we report serial imaging and turnover studies in which we labelled SAP with iodine-131, a cheap alternative isotope which would be expected to yield poorer images but permit more prolonged turnover measurements. Imaging and plasma clearance and whole body retention (WBR) of tracer were studied for up to 7 days in ten patients with proven systemic AL amyloidosis and two patients in whom the diagnosis was suspected, after i.v. administration of about 37 MBq of {sup 131}I-SAP. Normal blood pool images were obtained in the latter two subjects and amyloidosis was subsequently refuted histologically. WBR at 48 h was <60% and 6-h plasma activity was >65% of the injected dose (i.d.). Among the other ten patients, amyloid deposits were identified in the spleen in eight cases, liver in five and kidneys in four; other sites that gave positive results included bone, joints and soft tissues, and the myocardium in one case. Up to 95% of the tracer localised into amyloid within 6-h, and the values for WBR became progressively more discriminating during the study period, exceeding the normal reference value (<25%) in all cases by day 7. The optimal imaging time was found to be between 24 and 48 h. The duration of the study enabled us to measure the tracer elimination half-life which was increased in all cases by up to tenfold. Follow-up studies performed after 2-24 months in four patients who were treated with iododoxorubicin showed regression of amyloid in one patient and a small increase in one case; in the other two patients the imaging and turnover studies were identical to baseline. Despite its unfavourable imaging characteristics, {sup 131}I-SAP produced diagnostic scans in every patient in this series and, coupled with the detailed turnover information, is adequate for monitoring disease progress. (orig.) With 5 figs., 2 tabs., 19 refs.

Hawkins, P.N.; Pepys, M.B. [Immunological Medicine Unit, Department of Medicine, Royal Postgraduate Medical School, London (United Kingdom); Aprile, C. [Nuclear Medicine Service, Scientific Institute Fondazione Maugeri, Pavia (Italy); Capri, G.; Vigano, L.; Munzone, E.; Gianni, L. [Division of Medical Oncology, Istituto Nazionale Tumori, Milan (Italy); Merlini, G. [Biotechnology Research Laboratories, University Hospital S. Matteo, Pavia (Italy)

1998-07-01

113

A Drosophila gene encoding a protein resembling the human ?-amyloid protein precursor  

International Nuclear Information System (INIS)

The authors have isolated genomic and cDNA clones for a Drosophila gene resembling the human ?-amyloid precursor protein (APP). This gene produces a nervous system-enriched 6.5-kilobase transcript. Sequencing of cDNAs derived from the 6.5-kilobase transcript predicts an 886-amino acid polypeptide. This polypeptide contains a putative transmembrane domain and exhibits strong sequence similarity to cytoplasmic and extracellular regions of the human ?-amyloid precursor protein. There is a high probability that this Drosophila gene corresponds to the essential Drosophila locus vnd, a gene required for embryonic nervous system development

114

Constitutive and regulated ?-secretase cleavage of Alzheimer’s amyloid precursor protein by a disintegrin metalloprotease  

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Amyloid ? peptide (A?), the principal proteinaceous component of amyloid plaques in brains of Alzheimer’s disease patients, is derived by proteolytic cleavage of the amyloid precursor protein (APP). Proteolytic cleavage of APP by a putative ?-secretase within the A? sequence precludes the formation of the amyloidogenic peptides and leads to the release of soluble APPs? into the medium. By overexpression of a disintegrin and metalloprotease (ADAM), classified as ADAM 10, in HEK 293 cell...

Lammich, Sven; Kojro, Elzbieta; Postina, Rolf; Gilbert, Sandra; Pfeiffer, Roland; Jasionowski, Marek; Haass, Christian; Fahrenholz, Falk

1999-01-01

115

Deletion of the amyloid precursor-like protein 2 (APLP2) does not affect hippocampal neuron morphology or function  

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Amyloid precursor protein (APP), the parent molecule to amyloid ? peptide, is part of larger gene family with two mammalian homologues, amyloid precursor-like protein 1 (APLP1) and amyloid precursor-like protein 2 (APLP2). Initial knock-out studies demonstrated that while single APP family gene deletions produced relatively mild phenotypes, deficiency of APLP2 and one other member of the gene family resulted in perinatal lethality, suggesting vital roles masked by functional redundancy of th...

Midthune, Brea; Tyan, Sheue-houy; Walsh, Jessica J.; Sarsoza, Floyd; Eggert, Simone; Hof, Patrick R.; Dickstein, Dara L.; Koo, Edward H.

2012-01-01

116

Studies of the cell biological function of the Amyloid Precursor Protein (APP) family in Drosophila melanogaster and mammals  

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Alzheimer?s disease (AD) is the most common neurodegenerative disorder affecting cognitive functions of the brain, and is pathologically characterized by extracellular Amyloid plaques and intracellular neurofibrillary tangles are the main pathological features of AD. Amyloid-beta, the main protein constituent of Amyloid plaques, is derived from the Amyloid Precursor Protein (APP) by proteolytic processing. APP is a type I transmembrane protein, which resembles a cell surface receptor, and c...

Soba, Peter

2004-01-01

117

Serum Protein Profile Alterations in Hemodialysis Patients  

Energy Technology Data Exchange (ETDEWEB)

Background: Serum protein profiling patterns can reflect the pathological state of a patient and therefore may be useful for clinical diagnostics. Here, we present results from a pilot study of proteomic expression patterns in hemodialysis patients designed to evaluate the range of serum proteomic alterations in this population. Methods: Surface-Enhanced Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (SELDI-TOFMS) was used to analyze serum obtained from patients on periodic hemodialysis treatment and healthy controls. Serum samples from patients and controls were first fractionated into six eluants on a strong anion exchange column, followed by application to four array chemistries representing cation exchange, anion exchange, metal affinity and hydrophobic surfaces. A total of 144 SELDI-TOF-MS spectra were obtained from each serum sample. Results: The overall profiles of the patient and control samples were consistent and reproducible. However, 30 well-defined protein differences were observed; 15 proteins were elevated and 15 were decreased in patients compared to controls. Serum from one patient exhibited novel protein peaks suggesting possible additional changes due to a secondary disease process. Conclusion: SELDI-TOF-MS demonstrated dramatic serum protein profile differences between patients and controls. Similarity in protein profiles among dialysis patients suggests that patient physiological responses to end-stage renal disease and/or dialysis therapy have a major effect on serum protein profiles.

Murphy, G A; Davies, R W; Choi, M W; Perkins, J; Turteltaub, K W; McCutchen-Maloney, S L; Langlois, R G; Curzi, M P; Trebes, J E; Fitch, J P; Dalmasso, E A; Colston, B W; Ying, Y; Chromy, B A

2003-11-18

118

Amyloid precursor protein modulates ERK-1 and -2 signaling.  

Science.gov (United States)

The amyloid precursor protein (APP) is a transmembrane protein with a short cytoplasmic tail whose physiological function is unclear, although it is well documented that the proteolytic processing of APP could influence the development of Alzheimer's disease (AD) through the formation of membrane-bound C-terminal fragments (CTFs) and of beta-amyloid peptides (Abeta). We have recently shown that tyrosine-phosphorylated APP and CTFs may interact with Grb2 and ShcA adaptor proteins and that this coupling occurs at a higher extent in AD subjects only. To study the interaction between APP or CTFs and ShcA/Grb2 and to investigate their molecular target we have used as experimental model two different cell lines: H4 human neuroglioma cells and APP/APLP null mouse embryonic fibroblast cells (MEFs). Here we show that in H4 cells APP interacts with Grb2; conversely in APP/APLP-null MEF cells this interaction is possible only after the reintroduction of human APP by transfection. We have also shown that in MEF cells the transfection of a plasmid encoding for human APP wild-type enhances the phosphorylation of ERK-1 and -2 as revealed by Western blotting and immunofluorescence experiments. Finally, also in H4 cells the overexpression of APP upregulates the levels of phospho-ERK-1 and -2. In summary our data suggest that APP may influence phospho-ERK-1 and -2 signaling through its binding with Grb2 and ShcA adaptors. The meaning of this event is not clear, but APP interaction with these adaptors could be relevant to regulate mitogenic pathway. PMID:17384289

Venezia, Valentina; Nizzari, Mario; Repetto, Emanuela; Violani, Elisabetta; Corsaro, Alessandro; Thellung, Stefano; Villa, Valentina; Carlo, Pia; Schettini, Gennaro; Florio, Tullio; Russo, Claudio

2006-12-01

119

Inhibition of dynamin-dependent endocytosis increases shedding of the amyloid precursor protein ectodomain and reduces generation of amyloid ? protein  

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Full Text Available Abstract Background The amyloid precursor protein (APP is transported via the secretory pathway to the cell surface, where it may be cleaved within its ectodomain by ?-secretase, or internalized within clathrin-coated vesicles. An alternative proteolytic pathway occurs within the endocytic compartment, where the sequential action of ?- and ?-secretases generates the amyloid ? protein (A?. In this study, we investigated the effects of modulators of endocytosis on APP processing. Results Human embryonic kidney cells were transfected with a dominant negative mutant of dynamin I, an important mediator of clathrin-dependent endocytosis, and APP proteolysis was analyzed. Overexpression of the mutant dynamin (dyn I K44A resulted in increased shedding of the APP ectodomain (sAPP?, accumulation of the C-terminal ?-secretase product C83, and a reduction in the release of A?. Levels of mature APP on the cell surface were increased in cells expressing dyn I K44A, and internalization of surface-immunolabeled APP, assessed by fluorescence microscopy, was inhibited. Dynamin is a substrate for protein kinase C (PKC, and it was hypothesized that activators of PKC, which are known to stimulate ?-secretase-mediated cleavage of APP, might exert their effects by inhibiting dynamin-dependent endocytosis. However, the internalization of surface-biotinylated APP was unaffected by treatment of cells with phorbol 12-myristate 13-acetate in the presence of the ?-secretase inhibitor TAPI-1. Conclusion The results indicate that APP is internalized by a dynamin-dependent process, and suggest that alterations in the activity of proteins that mediate endocytosis might lead to significant changes in A? production.

Lopez-Coviella Ignacio

2005-08-01

120

?-Secretase processing of the Alzheimer amyloid-? precursor protein and its homolog APLP2  

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The amyloid-? precursor protein (APP) has been widely studied due to its role in Alzheimer´s disease (AD). When APP is sequentially cleaved by ?- and ?-secretase, amyloid-? (A?) is formed. A? is prone to aggregate and is toxic to neurons. However, the main processing pathway for APP involves initial cleavage at the ?-site, within the A? region, instead generating a neuroprotective soluble fragment, sAPP?. APP is a member of a protein family, also including the proteins APLP1 and APL...

Jacobsen, Kristin

2013-01-01

 
 
 
 
121

Amyloid fibrils from readily available sources: milk casein and lens crystallin proteins.  

Science.gov (United States)

Amyloid fibrils are a highly ordered and robust aggregated form of protein structure in which the protein components are arranged in long fibrillar arrays comprised of ?-sheet. Because of these properties, along with their biocompatibility, amyloid fibrils have attracted much research attention as bionanomaterials, for example as template structures (in some cases following modification) that can be used as biosensors, encapsulators, and biomimetic materials. To use amyloid fibrils for such a range of applications will require them to be obtained relatively easily in large quantities. In this chapter, we describe methods for isolating crystallin and casein proteins from readily available sources that contain abundant protein, i.e., the eye lens and milk, respectively, and the subsequent conversion of these proteins into amyloid fibrils. PMID:23504420

Ecroyd, Heath; Garvey, Megan; Thorn, David C; Gerrard, Juliet A; Carver, John A

2013-01-01

122

Islet amyloid polypeptide forms rigid lipid-protein amyloid fibrils on supported phospholipid bilayers.  

Science.gov (United States)

Islet amyloid polypeptide (IAPP) forms fibrillar amyloid deposits in the pancreatic islets of Langerhans of patients with type 2 diabetes mellitus, and its misfolding and aggregation are thought to contribute to beta-cell death. Increasing evidence suggests that IAPP fibrillization is strongly influenced by lipid membranes and, vice versa, that the membrane architecture and integrity are severely affected by amyloid growth. Here, we report direct fluorescence microscopic observations of the morphological transformations accompanying IAPP fibrillization on the surface of supported lipid membranes. Within minutes of application in submicromolar concentrations, IAPP caused extensive remodeling of the membrane including formation of defects, vesiculation, and tubulation. The effects of IAPP concentration, ionic strength, and the presence of amyloid seeds on the bilayer perturbation and peptide aggregation were examined. Growth of amyloid fibrils was visualized using fluorescently labeled IAPP or thioflavin T staining. Two-color imaging of the peptide and membranes revealed that the fibrils were initially composed of the peptide only, and vesiculation occurred in the points where growing fibers touched the lipid membrane. Interestingly, after 2-5 h of incubation, IAPP fibers became "wrapped" by lipid membranes derived from the supported membrane. Progressive increase in molecular-level association between amyloid and membranes in the maturing fibers was confirmed by Förster resonance energy transfer spectroscopy. PMID:18155730

Domanov, Yegor A; Kinnunen, Paavo K J

2008-02-01

123

Cleavage of amyloid-beta precursor protein and amyloid-beta precursor-like protein by BACE 1.  

Science.gov (United States)

Site-specific proteolysis of the amyloid-beta precursor protein (APP) by BACE 1 and gamma-secretase, a central event in Alzheimer disease, releases a large secreted extracellular fragment (called APP(S)), peptides of 40-43 residues derived from extracellular and transmembrane sequences (Abeta), and a short intracellular fragment (APP intracellular domain) that may function as a transcriptional activator in a complex with the adaptor protein Fe65 and the nuclear protein Tip60. APP is closely related to APP-like protein (APLP) 1 and APLP2, but only APP is known to be cleaved by BACE 1 and to be involved in Alzheimer disease. We now demonstrate that similar to APP, APLP1 and APLP2 are also cleaved by BACE 1 but not by ADAM 9, another APP protease, and also transactivate nuclear Tip60 in a complex with Fe65. Paradoxically, although BACE 1 cleavage appears to be specific for APP and APLPs, their cleavage sequences exhibit no homology, and a short sequence (7 amino acids) from APP that when placed close to the membrane converts a membrane protein that is normally not cleaved by BACE 1 into a BACE 1 substrate. Our data demonstrate that APLPs and APP are processed similarly to act via the same nuclear target, suggesting that BACE 1 cleavage regulates a common function of APP and APLPs in neurons. PMID:14699153

Li, Qiming; Südhof, Thomas C

2004-03-12

124

Mint proteins are required for synaptic activity-dependent amyloid precursor protein (APP) trafficking and amyloid ? generation.  

Science.gov (United States)

Aberrant amyloid ? (A?) production plays a causal role in Alzheimer disease pathogenesis. A major cellular pathway for A? generation is the activity-dependent endocytosis and proteolytic cleavage of the amyloid precursor protein (APP). However, the molecules controlling activity-dependent APP trafficking in neurons are less defined. Mints are adaptor proteins that directly interact with the endocytic sorting motif of APP and are functionally important in regulating APP endocytosis and A? production. We analyzed neuronal cultures from control and Mint knockout neurons that were treated with either glutamate or tetrodotoxin to stimulate an increase or decrease in neuronal activity, respectively. We found that neuronal activation by glutamate increased APP endocytosis, followed by elevated APP insertion into the cell surface, stabilizing APP at the plasma membrane. Conversely, suppression of neuronal activity by tetrodotoxin decreased APP endocytosis and insertion. Interestingly, we found that activity-dependent APP trafficking and A? generation were blocked in Mint knockout neurons. We showed that wild-type Mint1 can rescue APP internalization and insertion in Mint knockout neurons. In addition, we found that Mint overexpression increased excitatory synaptic activity and that APP was internalized predominantly to endosomes associated with APP processing. We demonstrated that presenilin 1 (PS1) endocytosis requires interaction with the PDZ domains of Mint1 and that this interaction facilitates activity-dependent colocalization of APP and PS1. These findings demonstrate that Mints are necessary for activity-induced APP and PS1 trafficking and provide insight into the cellular fate of APP in endocytic pathways essential for A? production. PMID:24742670

Sullivan, Sarah E; Dillon, Gregory M; Sullivan, Josefa M; Ho, Angela

2014-05-30

125

The Role of Phospholipase D in Amyloid Precursor Protein Processing  

Directory of Open Access Journals (Sweden)

Full Text Available The generation of a secreted N-terminal fragment of the amyloid precursor protein (A PPs can be stimulated by a variety of signaling pathways many of which are also known to modulate the activity of the phospholipase D (PLD enzyme. This study used primary rat neuronal cerebellar granule (CG cultures and SH-SY5Y human neuroblastoma cell lines to determine the potential role of PLD in the protein kinase C (PKC-associated generation of A PPs. Protein release was markedly enhanced by direct PKC stimulation following treatment of both cell type with either phorbol ester or indirectly by the muscarinic agonist carbachol and these effects were greatly attenuated by co-incubation with the PKC inhibitor GF109203X. A partial inhibition of PKC- and carbachol-stimulated A PPs secretion was also achieved by pre-treatment of the cells with toxin B, a PLD inhibitor. This suggested that PLD may play a role downstream of PKC in the control of A PPs secretion.

Mark McLaughlin

2005-01-01

126

Amyloid Precursor Protein Expression Modulates Intestine Immune Phenotype  

Science.gov (United States)

Amyloid precursor protein (APP) is widely expressed across many tissue and cell types. Proteolytic processing of the protein gives rise to a plethora of protein fragments with varied biological activities. Although a large amount of data has been generated describing the metabolism of the protein in neurons, its role in regulating the phenotype of other cells remains unclear. Based upon prior work demonstrating that APP regulates the activation phenotype of monocytic lineage cells, we hypothesized that APP can regulate macrophage activation phenotype in tissues other than brain. Ileums of the small intestines from C57BL6/J wild type and APP?/? mice were compared as a representative tissue normally associated with abundant macrophage infiltration. APP?/? intestines demonstrated diminished CD68 immunoreactivity compared to wild type mice. This correlated with significantly less cycloxygenase-2 (cox-2), CD68, CD40, CD11c, and ?III-tubulin protein levels. Peritoneal macrophage from APP?/? mice demonstrated decreased in vitro migratory ability compared to wild type cells and diminished basal KC cytokine secretion. Whereas, APP?/? intestinal macrophage had an increase in basal KC cytokine secretion compared to wild type cells. Conversely, there was a significant decrease in multiple cytokine levels in APP?/? compared to wild type ileums. Finally, APP?/? mice demonstrated impaired absorption and increased motility compared to wild type mice. These data demonstrate the APP expression regulates immune cell secretions and phenotype and intestinal function. This data set describes a novel function for this protein or its metabolites that may be relevant not only for Alzheimer’s disease but a range of immune-related disorders. PMID:22124967

Puig, Kendra L.; Swigost, Adam J.; Zhou, Xudong; Sens, MaryAnn; Combs, Colin K.

2014-01-01

127

Comparative transcriptome profiling of amyloid precursor protein family members in the adult cortex  

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Abstract Background The ?-amyloid precursor protein (APP) and the related ?-amyloid precursor-like proteins (APLPs) undergo complex proteolytic processing giving rise to several fragments. Whereas it is well established that A? accumulation is a central trigger for Alzheimer's disease, the physiological role of APP family members and their diverse proteolytic products is still largely unknown. The secreted APPs? ectodomain has been shown to be involved in neuroprotection ...

Eils Roland; Prinz Marco; Gretz Norbert; Tschäpe Jakob-Andreas; Filippov Mikhail A; Aydin Dorothee; Brors Benedikt; Müller Ulrike C

2011-01-01

128

Cellular localization and mechanism of amyloid precursor protein (APP) homodimer formation in an oxidizing environment  

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Das Amyloid Vorläufer Protein („amyloid precursor protein“, APP) ist ein Typ I integrales Membranprotein, welches strukturelle Ähnlichkeiten mit Zelloberflächenrezeptoren aufweist. Es umfasst einen großen N-terminalen extrazellulären Abschnitt, einen Transmembranbereich, sowie einen kurzen cytoplasmatischen C-Terminus im Innern der Zelle. APP ist Teil einer konservierten Genfamilie mit insgesamt 17 Mitgliedern, zu denen auch die beiden Säuger-APP-homologen APLP1 und APLP2 („amylo...

Isbert, Simone

2012-01-01

129

Untersuchungen zur Induktion von Filopodien durch das Amyloid Precursor Like Protein 1 (APLP1), ein Mitglied der APP-Genfamilie  

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Die Alzheimer Krankheit ist eine fortschreitendende Demenzerkrankung von der in Deutschland ca. 1,6 Millionen Menschen betroffen sind. Im Gehirn der Patienten finden sich sogenannte amyloide Plaques, deren Hauptbestandteil das A?-Protein ist. Dieses Peptid ist ein Spaltprodukt des APP-Proteins (engl. amyloid precursor protein). APP ist das namensgebende Mitglied der APP-Proteinfamilie zu der neben APP die beiden APP-Homologen APLP1 und APLP2 (engl. amyloid precursor like protein) gehören. O...

Langer, Andreas

2006-01-01

130

Pathogenetic mechanisms of amyloid A amyloidosis.  

Science.gov (United States)

Systemic amyloid A (AA) amyloidosis is a serious complication of chronic inflammation. Serum AA protein (SAA), an acute phase plasma protein, is deposited extracellularly as insoluble amyloid fibrils that damage tissue structure and function. Clinical AA amyloidosis is typically preceded by many years of active inflammation before presenting, most commonly with renal involvement. Using dose-dependent, doxycycline-inducible transgenic expression of SAA in mice, we show that AA amyloid deposition can occur independently of inflammation and that the time before amyloid deposition is determined by the circulating SAA concentration. High level SAA expression induced amyloidosis in all mice after a short, slightly variable delay. SAA was rapidly incorporated into amyloid, acutely reducing circulating SAA concentrations by up to 90%. Prolonged modest SAA overexpression occasionally produced amyloidosis after long delays and primed most mice for explosive amyloidosis when SAA production subsequently increased. Endogenous priming and bulk amyloid deposition are thus separable events, each sensitive to plasma SAA concentration. Amyloid deposits slowly regressed with restoration of normal SAA production after doxycycline withdrawal. Reinduction of SAA overproduction revealed that, following amyloid regression, all mice were primed, especially for rapid glomerular amyloid deposition leading to renal failure, closely resembling the rapid onset of renal failure in clinical AA amyloidosis following acute exacerbation of inflammation. Clinical AA amyloidosis rarely involves the heart, but amyloidotic SAA transgenic mice consistently had minor cardiac amyloid deposits, enabling us to extend to the heart the demonstrable efficacy of our unique antibody therapy for elimination of visceral amyloid. PMID:23959890

Simons, J Paul; Al-Shawi, Raya; Ellmerich, Stephan; Speck, Ivana; Aslam, Samrina; Hutchinson, Winston L; Mangione, Palma P; Disterer, Petra; Gilbertson, Janet A; Hunt, Toby; Millar, David J; Minogue, Shane; Bodin, Karl; Pepys, Mark B; Hawkins, Philip N

2013-10-01

131

Ribonuclease A suggests how proteins self-chaperone against amyloid fiber formation  

Energy Technology Data Exchange (ETDEWEB)

Genomic analyses have identified segments with high fiber-forming propensity in many proteins not known to form amyloid. Proteins are often protected from entering the amyloid state by molecular chaperones that permit them to fold in isolation from identical molecules; but, how do proteins self-chaperone their folding in the absence of chaperones? Here, we explore this question with the stable protein ribonuclease A (RNase A). We previously identified fiber-forming segments of amyloid-related proteins and demonstrated that insertion of these segments into the C-terminal hinge loop of nonfiber-forming RNase A can convert RNase A into the amyloid state through three-dimensional domain-swapping, where the inserted fiber-forming segments interact to create a steric zipper spine. In this study, we convert RNase A into amyloid-like fibers by increasing the loop length and hence conformational freedom of an endogenous fiber-forming segment, SSTSAASS, in the N-terminal hinge loop. This is accomplished by sandwiching SSTSAASS between inserted Gly residues. With these inserts, SSTSAASS is now able to form the steric zipper spine, allowing RNase A to form amyloid-like fibers. We show that these fibers contain RNase A molecules retaining their enzymatic activity and therefore native-like structure. Thus, RNase A appears to prevent fiber formation by limiting the conformational freedom of this fiber-forming segment from entering a steric zipper. Our observations suggest that proteins have evolved to self-chaperone by using similar protective mechanisms.

Teng, Poh K.; Anderson, Natalie J.; Goldschmidt, Lukasz; Sawaya, Michael R.; Sambashivan, Shilpa; Eisenberg, David (UCB)

2012-05-29

132

Exploring new biological functions of amyloids: bacteria cell agglutination mediated by host protein aggregation.  

Science.gov (United States)

Antimicrobial proteins and peptides (AMPs) are important effectors of the innate immune system that play a vital role in the prevention of infections. Recent advances have highlighted the similarity between AMPs and amyloid proteins. Using the Eosinophil Cationic Protein as a model, we have rationalized the structure-activity relationships between amyloid aggregation and antimicrobial activity. Our results show how protein aggregation can induce bacteria agglutination and cell death. Using confocal and total internal reflection fluorescence microscopy we have tracked the formation in situ of protein amyloid-like aggregates at the bacteria surface and on membrane models. In both cases, fibrillar aggregates able to bind to amyloid diagnostic dyes were detected. Additionally, a single point mutation (Ile13 to Ala) can suppress the protein amyloid behavior, abolishing the agglutinating activity and impairing the antimicrobial action. The mutant is also defective in triggering both leakage and lipid vesicle aggregation. We conclude that ECP aggregation at the bacterial surface is essential for its cytotoxicity. Hence, we propose here a new prospective biological function for amyloid-like aggregates with potential biological relevance. PMID:23133388

Torrent, Marc; Pulido, David; Nogués, M Victòria; Boix, Ester

2012-01-01

133

Amyloid precursor protein and tau transgenic models of Alzheimer's disease: insights from the past and directions for the future  

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During the last 20 years, our understanding of the mechanisms underlying Alzheimer's disease (AD) has considerably improved, in part owing to both in vitro and in vivo model systems. Studies in mice expressing both human amyloid precursor protein and human tau have provided clear evidence that amyloid-? and tau interact in the pathogenesis of AD. Moreover, amyloid-? toxicity has been shown to be tau-dependent since reducing tau levels prevents behavioral deficits and sudden death in amyloid...

Sahara, Naruhiko; Lewis, Jada

2010-01-01

134

Neurogenin 2 Mediates Amyloid-? Precursor Protein-stimulated Neurogenesis.  

Science.gov (United States)

Amyloid-? precursor protein (APP) is well studied for its role in Alzheimer disease, although its normal function remains uncertain. It has been reported that APP stimulates the proliferation and neuronal differentiation of neural stem/progenitor cells (NSPCs). In this study we examined the role of APP in NSPC differentiation. To identify proteins that may mediate the effect of APP on NSPC differentiation, we used a gene array approach to find genes whose expression correlated with APP-induced neurogenesis. We found that the expression of neurogenin 2 (Ngn2), a basic helix-loop-helix transcription factor, was significantly down-regulated in NSPCs from APP knock-out mice (APPKO) and increased in APP transgenic (Tg2576) mice. Ngn2 overexpression in APPKO NSPCs promoted neuronal differentiation, whereas siRNA knockdown of Ngn2 expression in wild-type NSPCs decreased neuronal differentiation. The results demonstrate that APP-stimulated neuronal differentiation of NSPCs is mediated by Ngn2. PMID:25217641

Bolós, Marta; Hu, Yanling; Young, Kaylene M; Foa, Lisa; Small, David H

2014-11-01

135

Alzheimer beta A4-amyloid protein precursor in immunocompetent cells.  

Science.gov (United States)

The mechanism of proteolytic breakdown of the beta A4-amyloid protein precursor (APP) has attracted much attention because of its relevance for Alzheimer's disease. Apart from the pathological role of APP in the amyloidogenesis, many efforts have been made to identify the functional significance of this widely expressed protein in various biological processes. Employing biochemical techniques, we demonstrate that APP is involved in the initiation of the immune response. Upon stimulation, it is expressed by the major functional types of T-lymphocytes, i.e. CD4+ and CD8+ cells. As was demonstrated for the CD4+ lymphoid cell line H9, APP is predominantly secreted. The remaining COOH-terminal fragments generated upon secretion were highly unstable. Of the APP produced by immunocompetent cells, considerable amounts were shown to be leukocyte-derived APP (L-APP). In addition, we were able to identify the KPI-containing L-APP isoform, L-APP733, as the major expressed L-APP isoform in immunocompetent cells, including rat microglial cells and astrocytes. The L-APP expression pattern of these cells showed high similarity. These findings seem to be indicative of an important function of APP within the immune system. Therefore, APP may be involved in various immunopathogenic conditions of the periphery and in the central nervous system. PMID:1429732

Mönning, U; König, G; Banati, R B; Mechler, H; Czech, C; Gehrmann, J; Schreiter-Gasser, U; Masters, C L; Beyreuther, K

1992-11-25

136

The physical chemistry of the amyloid phenomenon: thermodynamics and kinetics of filamentous protein aggregation.  

Science.gov (United States)

In this chapter, we present an overview of the kinetics and thermodynamics of protein aggregation into amyloid fibrils. The perspective we adopt is largely experimental, but we also discuss recent developments in data analysis and we show that only a combination of well-designed experiments with appropriate theoretical modelling is able to provide detailed mechanistic insight into the complex pathways of amyloid formation. In the first part of the chapter, we describe measurements of the thermodynamic stability of the amyloid state with respect to the soluble state of proteins, as well as the magnitude and origin of this stability. In the second part, we discuss in detail the kinetics of the individual molecular steps in the overall mechanism of the conversion of soluble protein into amyloid fibrils. Finally, we highlight the effects of external factors, such as salt type and concentration, chemical denaturants and molecular chaperones on the kinetics of aggregation. PMID:25131584

Buell, Alexander K; Dobson, Christopher M; Knowles, Tuomas P J

2014-01-01

137

Serum acute phase proteins in dogs with symptomatic esophageal spirocercosis.  

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Canine spirocercosis (CS) is a helminthic infection caused by the nematode Spirocerca lupi. The clinical hallmark of the disease is esophageal dysphagia due to parasite-induced esophageal nodules. Currently, there is limited information on the involvement of serum acute phase proteins (APPs) in the symptomatic CS. The objective of this study was to investigate whether C-reactive protein (CRP), serum amyloid A (SAA), haptoglobin (Hp) and albumin are involved in CS, and if their concentrations measured on admission reflect the severity of benign esophageal lesions. Nineteen dogs with spontaneous symptomatic esophageal spirocercosis and 7 clinically healthy dogs were studied retrospectively. The most consistently increased APP in the symptomatic dogs was Hp (95% of the dogs), followed by CRP (68%). The SAA concentrations were infrequently increased (5% of the dogs), while albumin concentrations were decreased in 58% of the affected dogs. The dogs with spirocercosis had significantly higher median concentrations of Hp (p=0.0001) and CRP (p=0.02) compared to healthy dogs. Median albumin concentrations did not differ between the two groups of dogs. The median concentrations of Hp, CRP and albumin did not differ significantly between the dogs having a single or multiple esophageal nodules. The results of this study indicate that in symptomatic CS, Hp and CRP are significantly and consistently increased, while SAA and albumin may be of limited value as diagnostic markers. No association was established between the concentrations of Hp, CRP and albumin measured on admission and the number of esophageal nodules. PMID:22683300

Mylonakis, Mathios E; Ceron, Jose J; Leontides, Leonidas; Rallis, Tim S; Koutinas, Alexander F

2012-11-23

138

Human serum amyloid P functions as a negative regulator of the innate and adaptive immune responses to DNA vaccines.  

Science.gov (United States)

The utility of DNA vaccines has been limited by their failure to elicit sufficiently potent immune responses in many human applications, whereas DNA vaccinations in mice have been very successful. However, the underlying mechanisms remain unknown. We hypothesize that serum amyloid P component (SAP), which has a species-specific, DNA-binding ability, contributes to the differences between human and mice and then limits DNA vaccine's efficacy in vivo. In our study, DNA vaccine-induced adaptive immune responses were also significantly decreased in the human SAP (hSAP) transgenic mice. Using human promonocytic cell line THP-1-derived macrophages as a cell model, we found that cells incubated with a hSAP-DNA complex showed significant defects in innate immune activations, whereas mouse SAP had similar, albeit very weak, activities. hSAP also significantly inhibited the functions of two identified DNA sentinels, high-mobility group B protein 1 and antimicrobial peptide LL37, and redirected DNA update to FcRs leading to endocytosis and endosomal degradation. We also found that a chemical SAP inhibitor strongly recovered the suppressed innate immune responses to DNA in the presence of human serum and enhanced the immunogenicity of DNA vaccines in vivo. Our data indicated that SAP is a key negative regulator for innate immune responses to DNA and may be partly responsible for the insufficient immune responses after DNA vaccinations in humans. SAP suppression may be a novel strategy for improving efficacy of human DNA vaccines and requires further clinical investigations. PMID:21278351

Wang, Yue; Guo, Yingjun; Wang, Xiaohui; Huang, Jinfeng; Shang, Jingli; Sun, Shuhan

2011-03-01

139

Characterization of reconstituted high-density lipoprotein particles formed by lipid interactions with human serum amyloid A.  

Science.gov (United States)

The acute-phase human protein serum amyloid A (SAA) is enriched in high-density lipoprotein (HDL) in patients with inflammatory diseases. Compared with normal HDL containing apolipoprotein A-I, which is the principal protein component, characteristics of acute-phase HDL containing SAA remain largely undefined. In the present study, we examined the physicochemical properties of reconstituted HDL (rHDL) particles formed by lipid interactions with SAA. Fluorescence and circular dichroism measurements revealed that although SAA was unstructured at physiological temperature, ?-helix formation was induced upon binding to phospholipid vesicles. SAA also formed rHDL particles by solubilizing phospholipid vesicles through mechanisms that are common to other exchangeable apolipoproteins. Dynamic light scattering and nondenaturing gradient gel electrophoresis analyses of rHDL after gel filtration revealed particle sizes of approximately 10nm, and a discoidal shape was verified by transmission electron microscopy. Thermal denaturation experiments indicated that SAA molecules in rHDL retained ?-helical conformations at 37°C, but were almost completely denatured around 60°C. Furthermore, trypsin digestion experiments showed that lipid binding rendered SAA molecules resistant to protein degradation. In humans, three major SAA1 isoforms (SAA1.1, 1.3, and 1.5) are known. Although these isoforms have different amino acids at residues 52 and 57, no major differences in physicochemical properties between rHDL particles resulting from lipid interactions with SAA isoforms have been found. The present data provide useful insights into the effects of SAA enrichment on the physicochemical properties of HDL. PMID:25063355

Takase, Hiroka; Furuchi, Hiroki; Tanaka, Masafumi; Yamada, Toshiyuki; Matoba, Kyoko; Iwasaki, Kenji; Kawakami, Toru; Mukai, Takahiro

2014-10-01

140

Organization and Biology of the Porcine Serum Amyloid A (SAA) Gene Cluster: Isoform Specific Responses to Bacterial Infection.  

DEFF Research Database (Denmark)

Serum amyloid A (SAA) is a prominent acute phase protein. Although its biological functions are debated, the wide species distribution of highly homologous SAA proteins and their uniform behavior in response to injury or inflammation in itself suggests a significant role for this protein. The pig is increasingly being used as a model for the study of inflammatory reactions, yet only little is known about how specific SAA genes are regulated in the pig during acute phase responses and other responses induced by pro-inflammatory host mediators. We designed SAA gene specific primers and quantified the gene expression of porcine SAA1, SAA2, SAA3, and SAA4 by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) in liver, spleen, and lung tissue from pigs experimentally infected with the Gram-negative swine specific bacterium Actinobacillus pleuropneumoniae, as well as from pigs experimentally infected with the Gram-positive bacterium Staphylococcus aureus. Our results show that: 1) SAA1 may be a pseudogene in pigs; 2) we were able to detect two previously uncharacterized SAA transcripts, namely SAA2 and SAA4, of which the SAA2 transcript is primarily induced in the liver during acute infection and presumably contributes to circulating SAA in pigs; 3) Porcine SAA3 transcription is induced both hepatically and extrahepatically during acute infection, and may be correlated to local organ affection; 4) Hepatic transcription of SAA4 is markedly induced in pigs infected with A. pleuropneumoniae, but only weakly in pigs infected with S. aureus. These results for the first time establish the infection response patterns of the four porcine SAA genes which will be of importance for the use of the pig as a model for human inflammatory responses, e.g. within sepsis, cancer, and obesity research.

Skovgaard, Kerstin; Heegaard, Peter M. H.

2013-01-01

 
 
 
 
141

Clinical correlation of serum protein electrophoresis  

International Nuclear Information System (INIS)

Serum protein electrophoresis is a simple, reliable and specific method of separating different protein fractious. A study was carried out in 1556 symptomatic cases reporting to Dept of Chemical pathology, Army Medical College of serum protein electrophoresis. Aims and objectives were to see the diagnostic significance of protein electrophoresis especially in comparison to other related investigations like serum protein estimation and albumin. globulin (A; G) ratio etc. Deluxe electrophoresis chamber and sample applicator was used. Gelman DCD-16 digital computing densitometer was used for quantification. Results revealed that protein electrophoresis was essential for diseases like paraproteinaemia. Immune deficiencies and aantitrypsin deficiency. It is helpful along with other investigations in liver disease, nephritic syndrome, collagen disease and malignancy. Three hundred and sixty five cases had subacute/chronic infection, 340 had normal electrophoretic pattern, 250 cases were of congestive cardiac failure, 152 cases of nephritic syndrome. 90 cases of hepatic cirrhosis, 53 cases of chronic renal failure and 25 cases of paraproteinaemia were identified. Its sensitivity and specificity was more than serum protein estimation by dye methods. It is recommended that, full use of this diagnostic technique should be made for better diagnostic sensitivity and specificity. (author)

142

Neutron activation analysis of human serum proteins  

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Neutron activation analysis may be used to determine about 12 elements in serum samples. The blood has to be taken with a teflon cannula and processed in a dust-poor room. The pretreatment consists of lyophilisation or wet mineralisation. If necessary, post-irradiation separation is performed, based on adsorption on active carbon or hydride formation. The scope of the analysis is extended by gel permeation chromatography of the serum. The obtained fractions are then analyzed by instrumental neutron activation analysis. Alternatively, the combined proteins are separated from the ''salt'' fraction and analyzed. Results for normal human serum are given.

Woittiez, J.R.W.; Das, H.A. (Stichting Energieonderzoek Centrum Nederland, Petten); van Kamp, G.J. (Amsterdam Univ. (Netherlands))

1982-01-01

143

Neutron activation analysis of human serum proteins  

International Nuclear Information System (INIS)

Neutron activation analysis may be used to determine about 12 elements in serum samples. The blood has to be taken with a teflon cannula and processed in a dust-poor room. The pretreatment consists of lyophilisation or wet mineralisation. If necessary, post-irradiation separation is performed, based on adsorption on active carbon or hydride formation. The scope of the analysis is extended by gel permeation chromatography of the serum. The obtained fractions are then analyzed by instrumental neutron activation analysis. Alternatively, the combined proteins are separated from the ''salt'' fraction and analyzed. Results for normal human serum are given. (author)

144

MicroRNA regulation of Alzheimer's Amyloid precursor protein expression.  

Science.gov (United States)

Gene dosage effects of Amyloid precursor protein (APP) can cause familial AD. Recent evidence suggest that microRNA (miRNA) pathways, implicated in gene transcriptional control, could be involved in the development of sporadic Alzheimer's disease (AD). We therefore investigated whether miRNAs could participate in the regulation of APP gene expression. We show that miRNAs belonging to the miR-20a family (that is, miR-20a, miR-17-5p and miR-106b) could regulate APP expression in vitro and at the endogenous level in neuronal cell lines. A tight correlation between these miRNAs and APP was found during brain development and in differentiating neurons. We thus identify miRNAs as novel endogenous regulators of APP expression, suggesting that variations in miRNA expression could contribute to changes in APP expression in the brain during development and disease. This possibility is further corroborated by the observation that a statistically significant decrease in miR-106b expression was found in sporadic AD patients. PMID:19110058

Hébert, Sébastien S; Horré, Katrien; Nicolaï, Laura; Bergmans, Bruno; Papadopoulou, Aikaterini S; Delacourte, André; De Strooper, Bart

2009-03-01

145

Pregnancy amelioration of arthritis in SKG mice corresponds with alterations in serum amyloid A3 levels.  

Science.gov (United States)

OBJECTIVES: Pregnancy leads to rheumatoid arthritis remission in humans. The objective of this study was to determine if the SKG mouse could serve as a model for pregnancy-associated inflammatory arthritis amelioration. In addition, the maternal peripheral blood mononuclear cell (PBMC) transcriptome was assessed to define a biomarker associated with remission. METHODS: Cohorts of zymosan-treated pregnant SKG mice and controls were monitored for arthritis progression. Microarray analysis evaluated alterations in gene expression in maternal PBMCs at embryonic day 14.5 (E14.5) between arthritic and pregnancy-remitted mice. A selected target, serum amyloid A3 (SAA3), was further investigated using quantitative reverse transcriptase PCR (qRT-PCR) and an enzyme-linked immunosorbent assay (ELISA). RESULTS: Pregnancy resulted in complete or partial remission in the majority of the zymosan-treated SKG mice. Twenty-seven transcripts were differentially expressed in the PBMCs between arthritic and pregnancy-remitted mice. Expression and plasma SAA3 levels decreased with pregnancy-induced arthritis amelioration and plasma SAA3 levels correlated with arthritis severity. CONCLUSIONS: These results establish the SKG mouse as a model system to study pregnancy-induced amelioration of arthritis. These studies also establish SAA3 as a biomarker of arthritis amelioration in SKG mice. This model can be used to elucidate the molecular and cellular mechanisms underlying the impact of pregnancy on the maternal immune system that results in arthritis amelioration. PMID:23097751

Shaw, Laura A; Stefanski, Adrianne L; Peterson, Lisa K; Rumer, Kristen K; Vondracek, Andrea; Phang, Tzu L; Sakaguchi, Shimon; Winn, Virginia D; Dragone, Leonard L

2012-06-30

146

Elevated levels of serum amyloid A indicate poor prognosis in patients with esophageal squamous cell carcinoma  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Increase of Serum amyloid A (SAA level has been observed in patients with a variety of cancers. The objective of this study was to determined whether SAA level could be used as a prognostic parameter in patients with esophageal squamous cell carcinoma (ESCC. Methods SAA levels were measured by rate nephelometry immunoassay in 167 healthy controls and 167 ESCC patients prior to surgical resection. Statistical associations between clinicopathological observations and SAA levels were determined using the Mann–Whitney U test. The clinical value of SAA level as a prognostic parameter was evaluated using the Cox’s proportional hazards model. Results SAA levels were significantly higher in patients with ESCC compared to levels in healthy controls (13.88?±?15.19 mg/L vs. 2.26?±?1.66 mg/L, P?P?P?=?0.015, T classification (P?P?P?P?P? Conclusions An elevated level of preoperative SAA was found to associate with tumor progression and poor survival in patients with ESCC.

Wang Jun-Ye

2012-08-01

147

Serum amyloid P component inhibits influenza A virus infections: in vitro and in vivo studies.  

DEFF Research Database (Denmark)

Serum amyloid P component (SAP) binds in vitro Ca(2+)-dependently to several ligands including oligosaccharides with terminal mannose and galactose. We have earlier reported that SAP binds to human influenza A virus strains, inhibiting hemagglutinin (HA) activity and virus infectivity in vitro. These studies were extended to comprise five mouse-adapted influenza A strains, two swine influenza A strains, a mink influenza A virus, a ferret influenza A reassortant virus, a influenza B virus and a parainfluenza 3 virus. The HA activity of all these viruses was inhibited by SAP. Western blotting showed that SAP bound to HA trimers, monomers and HA1 and HA2 subunits of influenza A virus. Binding studies indicated that galactose, mannose and fucose moieties contributed to the SAP reacting site(s). Intranasal administration of human SAP to mice induced no demonstrable toxic reactions, and circulating antibodies against SAP were not detected. Preincubation of virus (A/Japan/57) with SAP prevented primary infection of mice and development of antiviral antibodies. After a single intranasal administration of SAP (40 microg) 1 h before primary infection with virus (2LD(50)), nine out of 10 mice survived on day 10 and these mice approached normal body weight, whereas control mice (one out of five surviving on day 10) died. The data provide evidence of the potential of intranasally administered SAP for prophylactic treatment of influenza A virus infections in humans.

Horvath, A; Andersen, I

2001-01-01

148

Ectodomain shedding of the amyloid precursor protein: Cellular control mechanisms and novel modifiers  

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Proteolytic cleavage in the ectodomain of the amyloid precursor protein (APP) is a key regulatory step in the generation of the Alzheimer's disease amyloid-beta (A beta) pepticle and occurs through two different protease activities termed alpha- and beta-secretase. Both proteases compete for APP cleavage, but have opposite effects on A beta generation. At present, little is known about the cellular pathways that control APP alpha- or beta-secretase cleavage and thus A beta generation. To expl...

Lichtenthaler, Stefan F.

2006-01-01

149

?-Secretase Inhibitor Potency Is Decreased by Aberrant ?-Cleavage Location of the “Swedish Mutant” Amyloid Precursor Protein  

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The amyloid-? (A?) peptide, widely known as the causative molecule of Alzheimer disease (AD), is generated by the sequential cleavage of amyloid precursor protein (APP) by the aspartyl proteases BACE1/?-secretase and presenilin/?-secretase. Inhibition of BACE1, therefore, is a promising strategy for preventing the progression of AD. However, ?-secretase inhibitors (BSIs) exhibit unexpectedly low potency in cells expressing “Swedish mutant” APP (APPswe) and in the transgenic mouse Tg2...

Yagishita, Sosuke; Futai, Eugene; Ishiura, Shoichi

2010-01-01

150

Brain amyloid β protein and memory disruption in Alzheimer’s disease  

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Weiming XiaCenter for Neurologic Diseases, Department of Neurology, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA, USAAbstract: The development of amyloid-containing neuritic plaques is an invariable characteristic of Alzheimer’s diseases (AD). The conversion from monomeric amyloid ? protein (A?) to oligomeric A? and finally neuritic plaques is highly dynamic. The specific Aß species that is correlated with disease...

Weiming Xia

2010-01-01

151

Membrane-protein interactions hold the key to understanding amyloid formation  

CERN Document Server

In this perspective we describe the critical role membranes play in modulating the structures of the Amyloid Precursor Proteins to produce the peptides involved in the Alzheimer's disease. Some of the key concepts related to protein aggregation including the potential role of the excited states of monomers in initiating protein aggregation are described.

Straub, John E

2014-01-01

152

A positive correlation between serum amyloid ? levels and depressive symptoms among community-dwelling elderly individuals in Japan  

Science.gov (United States)

Background Amyloid beta (A?) levels have been associated with an increased risk of Alzheimer’s disease (AD). As depression is common before the onset of AD, serum A? levels could be associated with depressive symptoms. The aim of this study was to investigate whether serum A? levels are associated with depressive symptoms and/or cognitive function in community-dwelling elderly individuals. Methods We examined the association between serum A? levels and depression among 419 Japanese community-dwelling elderly individuals aged 60 years and over. Subjects were divided into two subgroups: younger elderly between 60 and 69 years old and older elderly over 69 years old. The Mini-Mental State Examination (MMSE) was used to assess cognitive function, and symptoms of depression were assessed using the Center for Epidemiologic Studies Depression Scale (CES-D). The ability to perform activities of daily living was evaluated using the Tokyo Metropolitan Institute of Gerontology Index of Competence. Serum A? levels were measured with a human amyloid beta enzyme-linked immunosorbent assay kit. Results After controlling for potential confounding variables, a multiple linear regression analysis showed that increased levels of serum A?40 and A?42 were associated with higher CES-D scores in the older elderly subgroup. Under the same condition, multiple regression showed that serum A? levels were not associated with MMSE scores among the total subjects, younger elderly, or older elderly. Conclusion Serum A? levels were associated with depressive symptoms in community-dwelling elderly individuals. The present study indicates the possibility that serum A? may be involved in the development of late-onset depression.

Tsuruga, Koji; Sugawara, Norio; Yasui-Furukori, Norio; Takahashi, Ippei; Tsuchimine, Shoko; Kaneda, Ayako; Nakaji, Shigeyuki; Nakamura, Kazuhiko

2014-01-01

153

A positive correlation between serum amyloid ? levels and depressive symptoms among community-dwelling elderly individuals in Japan  

Directory of Open Access Journals (Sweden)

Full Text Available Koji Tsuruga,1 Norio Sugawara,1 Norio Yasui-Furukori,1 Ippei Takahashi,2 Shoko Tsuchimine,1 Ayako Kaneda,1 Shigeyuki Nakaji,2 Kazuhiko Nakamura1 1Department of Neuropsychiatry, 2Department of Social Medicine, Hirosaki University School of Medicine, Hirosaki, Japan Background: Amyloid beta (A? levels have been associated with an increased risk of Alzheimer’s disease (AD. As depression is common before the onset of AD, serum Aß levels could be associated with depressive symptoms. The aim of this study was to investigate whether serum A? levels are associated with depressive symptoms and/or cognitive function in community-dwelling elderly individuals. Methods: We examined the association between serum A? levels and depression among 419 Japanese community-dwelling elderly individuals aged 60 years and over. Subjects were divided into two subgroups: younger elderly between 60 and 69 years old and older elderly over 69 years old. The Mini-Mental State Examination (MMSE was used to assess cognitive function, and symptoms of depression were assessed using the Center for Epidemiologic Studies Depression Scale (CES-D. The ability to perform activities of daily living was evaluated using the Tokyo Metropolitan Institute of Gerontology Index of Competence. Serum A? levels were measured with a human amyloid beta enzyme-linked immunosorbent assay kit. Results: After controlling for potential confounding variables, a multiple linear regression analysis showed that increased levels of serum A?40 and A?42 were associated with higher CES-D scores in the older elderly subgroup. Under the same condition, multiple regression showed that serum A? levels were not associated with MMSE scores among the total subjects, younger elderly, or older elderly. Conclusion: Serum A? levels were associated with depressive symptoms in community-dwelling elderly individuals. The present study indicates the possibility that serum A? may be involved in the development of late-onset depression. Keywords: Alzheimer’s disease, depression, dementia, Japanese

Tsuruga K

2014-08-01

154

Purification and aggregation of the amyloid precursor protein intracellular domain.  

Science.gov (United States)

Amyloid precursor protein (APP) is a type I transmembrane protein associated with the pathogenesis of Alzheimer's disease (AD). APP is characterized by a large extracellular domain and a short cytosolic domain termed the APP intracellular domain (AICD). During maturation through the secretory pathway, APP can be cleaved by proteases termed ?, ?, and ?-secretases. Sequential proteolytic cleavage of APP with ? and ?-secretases leads to the production of a small proteolytic peptide, termed A?, which is amyloidogenic and the core constituent of senile plaques. The AICD is also liberated from the membrane after secretase processing, and through interactions with Fe65 and Tip60, can translocate to the nucleus to participate in transcription regulation of multiple target genes. Protein-protein interactions involving the AICD may affect trafficking, processing, and cellular functions of holo-APP and its C-terminal fragments. We have recently shown that AICD can aggregate in vitro, and this process is inhibited by the AD-implicated molecular chaperone ubiquilin-1. Consistent with these findings, the AICD has exposed hydrophobic domains and is intrinsically disordered in vitro, however it obtains stable secondary structure when bound to Fe65. We have proposed that ubiquilin-1 prevents inappropriate inter- and intramolecular interactions of AICD, preventing aggregation in vitro and in intact cells. While most studies focus on the role of APP in the pathogenesis of AD, the role of AICD in this process is not clear. Expression of AICD has been shown to induce apoptosis, to modulate signaling pathways, and to regulate calcium signaling. Over-expression of AICD and Fe65 in a transgenic mouse model induces Alzheimer's like pathology, and recently AICD has been detected in brain lysates by western blotting when using appropriate antigen retrieval techniques. To facilitate structural, biochemical, and biophysical studies of the AICD, we have developed a procedure to produce recombinantly large amounts of highly pure AICD protein. We further describe a method for inducing the in vitro thermal aggregation of AICD and analysis by atomic force microscopy. The methods described are useful for biochemical, biophysical, and structural characterization of the AICD and the effects of molecular chaperones on AICD aggregation. PMID:22952038

El Ayadi, Amina; Stieren, Emily S; Barral, José M; Oberhauser, Andres F; Boehning, Darren

2012-01-01

155

Analysis of the acute phase responses of Serum Amyloid A, Haptoglobin and Type 1 Interferon in cattle experimentally infected with foot-and-mouth disease virus serotype O  

DEFF Research Database (Denmark)

A series of challenge experiments were performed in order to investigate the acute phase responses to foot-and-mouth disease virus (FMDV) infection in cattle and possible implications for the development of persistently infected "carriers". The host response to infection was investigated through measurements of the concentrations of the acute phase proteins (APPs) serum amyloid A (SAA) and haptoglobin (HP), as well as the bioactivity of type 1 interferon (IFN) in serum of infected animals. Results were based on measurements from a total of 36 infected animals of which 24 were kept for observational periods exceeding 28 days in order to determine the carrier-status of individual animals. The systemic host response to FMDV in infected animals was evaluated in comparison to similar measurements in sera from 6 mock-inoculated control animals.There was a significant increase in serum concentrations of both APPs and type 1 IFN in infected animals coinciding with the onset of viremia and clinical disease. The measured parameters declined to baseline levels within 21 days after inoculation, indicating that there was no systemically measurable inflammatory reaction related to the carrier state of FMD. There was a statistically significant difference in the HP response between carriers and non-carriers with a lower response in the animals that subsequently developed into FMDV carriers. It was concluded that the induction of SAA, HP and type 1 IFN in serum can be used as markers of acute infection by FMDV in cattle.

Stenfeldt, Carolina; Heegaard, Peter M. H.

2011-01-01

156

Amyloid-related biomarkers and axonal damage proteins in parkinsonian syndromes  

DEFF Research Database (Denmark)

Clinical differentiation between parkinsonian syndromes (PS) remains a challenge despite well-established clinical diagnostic criteria. Specific diagnostic biomarkers have yet to be identified, though in recent years, studies have been published on the aid of certain brain related proteins (BRP) in the diagnosing of PS. We investigated the levels of the light subunit of neurofilament triplet protein (NF-L), total tau and phosphorylated tau, amyloid-ß(1-42), and the soluble a- and ß-cleaved fragments of amyloid precursor proteins in a cohort of patients with various PS.

Bech, Sara; Hjermind, Lena E

2012-01-01

157

Self-assembling of amyloid-like proteins  

International Nuclear Information System (INIS)

Full text: Septins are proteins from the GTP-binding family and participate in cell division cycle performing functions such as secretion and cytoskeletal division. They can also be found in neurodegenerative conditions as Alzheimers and Parkinson's diseases, forming highly organized fiber-like aggregates known as amyloids. In this work, we used small angle x-ray scattering (SAXS) to investigate the formation and time evolution of septins aggregates under the influence of temperature and concentration. The SAXS measurements were performed with the GTPase domain of human Septin 2 (SEPT2G) at 0.5 and 1 mg/mL and temperatures between 4 and 45 deg C. At 0.5 mg/mL and 4 deg C, the protein self-aggregates as a dimer, being stable over one hour of observation. When the temperature was increased to 15 deg C, the results demonstrate that cylinder-like aggregates are formed and coexist with some dimer population and a small amount of larger aggregates. However, the number of very large aggregates increases with time concomitantly with the decrease of cylinder amount in the solution. At 37 deg C cylinder-like aggregates are not longer present in solution, whereas a significant amount of dimers decreases from 50% to 20% in less than 1 hour. At 45 deg C such an effect is even more accentuated: the percentage of dimers is only 6% in solution into a favor of 94% of very larger aggregates. When we analyze the protein at 1 mg/mL, at 4 deg C cylinder-like aggregates (36 nm-long and 12 nm-cross section) are already formed, coexisting with dimers and, as occurred for lower concentration, the two populations remained unchanged over one hour of observation. Out results also indicate that the dimensions of these cylinders increase with the concentration and the percentage of cylinders and larger aggregates are higher than those found for 0.5 mg/mL. In conclusion, our results showed the coexistence of dimers of SEPT2G with small fibers and larger aggregates in solution that evolve not only with concentration and temperature but also with time. (author)

158

Self-assembling of amyloid-like proteins  

Energy Technology Data Exchange (ETDEWEB)

Full text: Septins are proteins from the GTP-binding family and participate in cell division cycle performing functions such as secretion and cytoskeletal division. They can also be found in neurodegenerative conditions as Alzheimers and Parkinson's diseases, forming highly organized fiber-like aggregates known as amyloids. In this work, we used small angle x-ray scattering (SAXS) to investigate the formation and time evolution of septins aggregates under the influence of temperature and concentration. The SAXS measurements were performed with the GTPase domain of human Septin 2 (SEPT2G) at 0.5 and 1 mg/mL and temperatures between 4 and 45 deg C. At 0.5 mg/mL and 4 deg C, the protein self-aggregates as a dimer, being stable over one hour of observation. When the temperature was increased to 15 deg C, the results demonstrate that cylinder-like aggregates are formed and coexist with some dimer population and a small amount of larger aggregates. However, the number of very large aggregates increases with time concomitantly with the decrease of cylinder amount in the solution. At 37 deg C cylinder-like aggregates are not longer present in solution, whereas a significant amount of dimers decreases from 50% to 20% in less than 1 hour. At 45 deg C such an effect is even more accentuated: the percentage of dimers is only 6% in solution into a favor of 94% of very larger aggregates. When we analyze the protein at 1 mg/mL, at 4 deg C cylinder-like aggregates (36 nm-long and 12 nm-cross section) are already formed, coexisting with dimers and, as occurred for lower concentration, the two populations remained unchanged over one hour of observation. Out results also indicate that the dimensions of these cylinders increase with the concentration and the percentage of cylinders and larger aggregates are higher than those found for 0.5 mg/mL. In conclusion, our results showed the coexistence of dimers of SEPT2G with small fibers and larger aggregates in solution that evolve not only with concentration and temperature but also with time. (author)

Sales, E.M.; Barbosa, L.R.S.; Itri, R. [Universidade de Sao Paulo (USP), SP (Brazil); Damalio, J.C.P.; Araujo, A.P.U. [Universidade de Sao Paulo (USP-SC), Sao Carlos, SP (Brazil); Spinozzi, F.; Mariani, P. [Universita Politecnica delle Marche, Ancona (Italy)

2012-07-01

159

Cannabidiol promotes amyloid precursor protein ubiquitination and reduction of beta amyloid expression in SHSY5YAPP+ cells through PPAR? involvement.  

Science.gov (United States)

The amyloidogenic cascade is regarded as a key factor at the basis of Alzheimer's disease (AD) pathogenesis. The aberrant cleavage of amyloid precursor protein (APP) induces an increased production and a subsequent aggregation of beta amyloid (A?) peptide in limbic and association cortices. As a result, altered neuronal homeostasis and oxidative injury provoke tangle formation with consequent neuronal loss. Cannabidiol (CBD), a Cannabis derivative devoid of psychotropic effects, has attracted much attention because it may beneficially interfere with several A?-triggered neurodegenerative pathways, even though the mechanism responsible for such actions remains unknown. In the present research, the role of CBD was investigated as a possible modulating compound of APP processing in SHSY5Y(APP+) neurons. In addition, the putative involvement of peroxisome proliferator-activated receptor-? (PPAR?) was explored as a candidate molecular site responsible for CBD actions. Results indicated the CBD capability to induce the ubiquitination of APP protein which led to a substantial decrease in APP full length protein levels in SHSY5Y(APP+) with the consequent decrease in A? production. Moreover, CBD promoted an increased survival of SHSY5Y(APP+) neurons, by reducing their long-term apoptotic rate. Obtained results also showed that all, here observed, CBD effects were dependent on the selective activation of PPAR?. PMID:24288245

Scuderi, Caterina; Steardo, Luca; Esposito, Giuseppe

2014-07-01

160

The acute phase response of haptoglobin and serum amyloid A (SAA) in cattle undergoing experimental infection with bovine respiratory syncytial virus  

DEFF Research Database (Denmark)

The ability of a pure virus infection to induce an acute phase protein response is of interest as viral infections are normally considered to be less efficient in inducing an acute phase protein response than bacterial infections. This was studied in a bovine model for infection with bovine respiratory syncytial virus (BRSV), analysing the induction of the two most dominant bovine acute phase proteins haptoglobin and serum amyloid A (SAA). Strong and reproducible acute phase responses were detected for both proteins, peaking at around 7-8 days after inoculation of BRSV, while no response was seen in mock-inoculated control animals. The serum concentrations reached for SAA and haptoglobin during the BRSV-induced acute phase response were generally the same or higher than previously reported for bacterial infections in calves. The magnitude and the duration of the haptoglobin response was found to correlate well with the severity of clinical signs (fever) and with the extent of lung consolidation while SAA responded most rapidly to infection.

Heegaard, Peter M. H.; TjØrnehØj, Kirsten

2000-01-01

 
 
 
 
161

Investigation of amyloid deposition in uterine leiomyoma patients  

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Objects: To investigate the pathogenesis of amyloid presented in uterine leiomyoma. Methods: 36 uterine leiomyoma patients were recruited and divided into two groups according to Congo red staining results. 6 cases are Congo red staining-positive, and 30 cases Congo red staining-negative which represented amyloid positive and amyloid negative respectively. All patients’ serum total protein (TP), albumin (Alb) and prealbumin (PA) levels were measured as well as blood hemoglobin (Hb), cell co...

Jinping Liu; Fei Zhai; Peng Ge; Jinhai Lu; Yi Qin; Xuguo Sun

2012-01-01

162

Macrophage differentiation and polarization via phosphatidylinositol 3-kinase/Akt-ERK signaling pathway conferred by serum amyloid P component.  

Science.gov (United States)

Macrophage differentiation and polarization is influenced by, and act on, many processes associated with autoimmunity. However, the molecular mechanisms underlying macrophage polarization in systemic lupus erythematosus (SLE) remain largely debated. We previously demonstrated that macrophage M2b polarization conferred by activated lymphocyte-derived (ALD)-DNA immunization could initiate and propagate murine lupus nephritis. Serum amyloid P component (SAP), a conserved acute-phase protein in mice, has been reported to bind to DNA and modulate immune responses. In this study, murine SAP was shown to promote macrophage-mediated ALD-DNA uptake through binding to ALD-DNA (SAP/ALD-DNA). Moreover, macrophage phenotypic switch from a proinflammatory M2b phenotype induced by ALD-DNA alone to an anti-inflammatory M2a phenotype stimulated with SAP/ALD-DNA were found because of PI3K/Akt-ERK signaling activation. Both in vivo SAP supplements and adoptive transfer of ex vivo programmed M2a macrophages induced by SAP/ALD-DNA into SLE mice could efficiently alleviate lupus nephritis. Importantly, increased IL-10 secretion, accompanied by anti-inflammatory effect exerted by M2a macrophages, was found to predominantly impede macrophage M2b polarization. Furthermore, neutralization of IL-10 notably reduced the suppressive effect of M2a macrophages. Our results demonstrate that binding of SAP to ALD-DNA could switch macrophage phenotypic polarization from proinflammatory M2b to anti-inflammatory M2a via PI3K/Akt-ERK signaling activation, thus exerting protective and therapeutic interventions on murine lupus nephritis. These data provide a possible molecular mechanism responsible for modulation of macrophage polarization in the context of lupus nephritis and open a new potential therapeutic avenue for SLE. PMID:21753147

Zhang, Weijuan; Xu, Wei; Xiong, Sidong

2011-08-15

163

Curcumin Reduces Amyloid Fibrillation of Prion Protein and Decreases Reactive Oxidative Stress  

Directory of Open Access Journals (Sweden)

Full Text Available Misfolding and aggregation into amyloids of the prion protein (PrP is responsible for the development of fatal transmissible neurodegenerative diseases. Various studies on curcumin demonstrate promise for the prevention of Alzheimer’s disease and inhibition of PrPres accumulation. To evaluate the effect of curcumin on amyloid fibrillation of prion protein, we first investigated the effect of curcumin on mouse prion protein (mPrP in a cell-free system. Curcumin reduced the prion fibril formation significantly. Furthermore, we monitored the change in apoptosis and reactive oxygen species (ROS level upon curcumin treatment in mouse neuroblastoma cells (N2a. Curcumin effectively rescues the cells from apoptosis and decreases the ROS level caused by subsequent co-incubation with prion amyloid fibrils. The assays in cell-free mPrP and in N2a cells of this work verified the promising effect of curcumin on the prevention of transmissible neurodegenerative diseases.

Raymond Chung

2013-07-01

164

Physiological functions of the amyloid precursor protein secretases ADAM10, BACE1, and presenilin.  

Science.gov (United States)

Alzheimer's disease causing mutations in the amyloid precursor protein (APP) or in the Presenilin 1 (PS1) or Presenilin 2 (PS2) genes increase the production of amyloid peptides (A?) that precipitate in amyloid plaques. Since amyloid plaques are also a prominent feature of sporadic Alzheimer's disease (AD), abnormal proteolysis of APP and the generation of amyloid beta (A?) are key events in the pathogenesis of AD. The proteases (secretases) that cleave APP are therefore important therapeutic targets, both for the rare familial forms but likely also for the sporadic forms of AD. The identification and understanding of the (neuro)biological functions of the ?-, ?-, and presenilin/?-secretase (complexes) is important for the development of drugs and the delineation of their associated side effects. The potential impact of this type of research exceeds the AD field since the function of these secretases are also linked to cellular pathways like ectodomain shedding of growth factors and regulated intramembrane proteolysis of receptors in developmental biology, tissue homeostasis, and tumorigenesis. The generation of mice deficient in presenilin 1, presenilin 2, the ?-secretase ADAM10, and the ?-secretases BACE1 and BACE2 were instrumental for the elucidation of the physiological functions of these proteases. Using these mouse models understanding how these secretases regulate amyloid peptide formation and how they exert their diverse biological functions could be significantly increased. This review attempts to summarize selected aspects of the current view of the multiple roles such proteases play in health and disease. PMID:22120156

Prox, Johannes; Rittger, Andrea; Saftig, Paul

2012-04-01

165

Dual amyloid domains promote differential functioning of the chaplin proteins during Streptomyces aerial morphogenesis  

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The chaplin proteins are functional amyloids found in the filamentous Streptomyces bacteria. These secreted proteins are required for the aerial development of Streptomyces coelicolor, and contribute to an intricate rodlet ultrastructure that decorates the surfaces of aerial hyphae and spores. S. coelicolor encodes eight chaplin proteins. Previous studies have revealed that only three of these proteins (ChpC, ChpE, and ChpH) are necessary for promoting aerial development, and of these three, ...

Capstick, David S.; Jomaa, Ahmad; Hanke, Chistopher; Ortega, Joaquin; Elliot, Marie A.

2011-01-01

166

Relevance of amyloid precursor-like protein 2 C-terminal fragments in pancreatic cancer cells  

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In some cellular systems, particularly neurons, amyloid precursor-like protein 2 (APLP2), and its highly homologous family member amyloid precursor protein (APP), have been linked to cellular growth. APLP2 and APP undergo regulated intramembrane proteolysis to produce C-terminal fragments. In this study, we found comprehensive expression of APLP2 C-terminal fragments in a panel of pancreatic cancer cell lines; however, APP C-terminal fragments were notably limited to the BxPC3 cell line. Exte...

Peters, Haley L.; Tuli, Amit; Wang, Xiaojian; Liu, Cuiling; Pan, Zenggang; Ouellette, Michel M.; Hollingsworth, Michael A.; Macdonald, Richard G.; Solheim, Joyce C.

2012-01-01

167

[Expression of beta-amyloid and tau-protein in mastocytes in Alzheimer disease].  

Science.gov (United States)

With immunochemical assay, the expression of B-amyloid and tau-protein in the mast cells of the stomach and skin was first detected in patients with Alzheimer's disease (AD), which supports the hypothesis that AD is a systemic disease involving different organs and tissues. Verification of expression of two key peptides that participate in the pathogenesis of AD in the samples of extra-brain tissues allows B-amyloid and tau-protein to be regarded as promising markers and mast cells to be considered to the most suitable object for lifetime diagnosis of neurodegenerative diseases. PMID:12669611

Kvetno?, I M; Kvetnaia, T V; Riadnova, I Iu; Fursov, B B; Ernandes-Jago, H; Blesa, J R

2003-01-01

168

Trafficking of cell surface beta-amyloid precursor protein: retrograde and transcytotic transport in cultured neurons  

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Amyloid beta-protein (A beta), the principal constituent of senile plaques seen in Alzheimer's disease (AD), is derived by proteolysis from the beta-amyloid precursor protein (beta PP). The mechanism of A beta production in neurons, which are hypothesized to be a rich source of A beta in brain, remains to be defined. In this study, we describe a detailed localization of cell surface beta PP and its subsequent trafficking in primary cultured neurons. Full-length cell surface beta PP was presen...

1995-01-01

169

Amyloid-clearing proteins and their epigenetic regulation as a therapeutic target in Alzheimer's disease  

Science.gov (United States)

Abnormal elevation of amyloid ?-peptide (A?) levels in the brain is the primary trigger for neuronal cell death specific to Alzheimer’s disease (AD). It is now evident that A? levels in the brain are manipulable due to a dynamic equilibrium between its production from the amyloid precursor protein (APP) and removal by amyloid clearance proteins. Clearance can be either enzymic or non-enzymic (binding/transport proteins). Intriguingly several of the main amyloid-degrading enzymes (ADEs) are members of the M13 peptidase family (neprilysin (NEP), NEP2 and the endothelin converting enzymes (ECE-1 and -2)). A distinct metallopeptidase, insulin-degrading enzyme (IDE), also contributes to A? degradation in the brain. The ADE family currently embraces more than 20 members, both membrane-bound and soluble, and of differing cellular locations. NEP plays an important role in brain function terminating neuropeptide signals. Its decrease in specific brain areas with age or after hypoxia, ischaemia or stroke contribute significantly to the development of AD pathology. The recently discovered mechanism of epigenetic regulation of NEP (and other genes) by the APP intracellular domain (AICD) and its dependence on the cell type and APP isoform expression suggest possibilities for selective manipulation of NEP gene expression in neuronal cells. We have also observed that another amyloid-clearing protein, namely transthyretin (TTR), is also regulated in the neuronal cell by a mechanism similar to NEP. Dependence of amyloid clearance proteins on histone deacetylases and the ability of HDAC inhibitors to up-regulate their expression in the brain opens new avenues for developing preventive strategies in AD.

Nalivaeva, Natalia N.; Belyaev, Nikolai D.; Kerridge, Caroline; Turner, Anthony J.

2014-01-01

170

Self-assembly of protein amyloid: a competition between amorphous and ordered aggregation  

CERN Document Server

Protein aggregation in the form of amyloid fibrils has important biological and technological implications. Although the self-assembly process is highly efficient, aggregates not in the fibrillar form would also occur and it is important to include these disordered species when discussing the thermodynamic equilibrium behavior of the system. Here, we initiate such a task by considering a mixture of monomeric proteins and the corresponding aggregates in the disordered form (micelles) and in the fibrillar form (amyloid fibrils). Starting with a model on the respective binding free energies for these species, we calculate their concentrations at thermal equilibrium. We then discuss how the incorporation of the disordered structure furthers our understanding on the various amyloid promoting factors observed empirically, and on the kinetics of fibrilization.

Lee, Chiu Fan

2010-01-01

171

Nox2-derived radicals contribute to neurovascular and behavioral dysfunction in mice overexpressing the amyloid precursor protein  

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Alterations in cerebrovascular regulation related to vascular oxidative stress have been implicated in the mechanisms of Alzheimer's disease (AD), but their role in the amyloid deposition and cognitive impairment associated with AD remains unclear. We used mice overexpressing the Swedish mutation of the amyloid precursor protein (Tg2576) as a model of AD to examine the role of reactive oxygen species produced by NADPH oxidase in the cerebrovascular alterations, amyloid deposition, and behavio...

Park, Laibaik; Zhou, Ping; Pitstick, Rose; Capone, Carmen; Anrather, Josef; Norris, Erin H.; Younkin, Linda; Younkin, Steven; Carlson, George; Mcewen, Bruce S.; Iadecola, Costantino

2008-01-01

172

Selective localization of amyloid precursor-like protein 1 in the cerebral cortex postsynaptic density.  

Science.gov (United States)

Senile plaques, a hallmark of Alzheimer's disease (AD), contain amyloid beta-peptide (A beta), which is generated from the larger amyloid beta protein precursor (APP). In addition to APP, several APP-related proteins have been recently identified in different organisms, including Drosophila amyloid precursor protein-like protein (APPL). Deficiency of APPL causes behavioral deficits in Drosophila, implicating a role in brain function. Moreover, mouse and human cDNA clones encoding amyloid precursor-like proteins (APLP1 and APLP2) have been identified and exhibit extensive sequence similarity to the APPL and APP genes. To define the potential role of APLP in the mammalian brain, we sought to directly localize APLP1 within the complex cortical synaptic structure. We focused on the postsynaptic density (PSD), which appears to be central to synaptic function. We now report that the 90 kDa APLP1, the first known APLP, is localized to the PSD from rat and human cerebral cortex. APLP1 increased during cortical synaptic development, suggesting a role in synaptogenesis or synaptic maturation. In contrast, APP was predominantly expressed in the synaptic membrane fraction, but was barely detectable in the PSD, including different subcellular distributions of APP and APLP1. Our observations raise the possibility that APLP1, a homologue of APPL, which appears to be necessary for normal behavior in Drosophila, participates in brain synaptic function in mammals. PMID:7494461

Kim, T W; Wu, K; Xu, J L; McAuliffe, G; Tanzi, R E; Wasco, W; Black, I B

1995-08-01

173

A novel mechanism for the regulation of amyloid precursor protein metabolism  

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Modifier of cell adhesion protein (MOCA; previously called presenilin [PS] binding protein) is a DOCK180-related molecule, which interacts with PS1 and PS2, is localized to brain areas involved in Alzheimer's disease (AD) pathology, and is lost from the soluble fraction of sporadic Alzheimer's disease (AD) brains. Because PS1 has been associated with ?-secretase activity, MOCA may be involved in the regulation of ?-amyloid precursor protein (APP) processing. Here we show that the expression...

Chen, Qi; Kimura, Hideo; Schubert, David

2002-01-01

174

Serum amyloid A and pairing formyl peptide receptor 2 are expressed in corneas and involved in inflammation-mediated neovascularization  

Directory of Open Access Journals (Sweden)

Full Text Available AIM:To solidify the involvement of Saa-related pathway in corneal neovascularization (CorNV. The pathogenesis of inflammatory CorNV is not fully understood yet, and our previous study implicated that serum amyloid A (Saa 1 (Saa1 and Saa3 were among the genes up-regulated upon CorNV induction in mice.METHODS:Microarray data obtained during our profiling project on CorNV were analyzed for the genes encoding the four SAA family members (Saa1-4, six reported SAA receptors (formyl peptide receptor 2, Tlr2, Tlr4, Cd36, Scarb1, P2rx7 and seven matrix metallopeptidases (Mmp 1a, 1b, 2, 3, 9, 10, 13 reportedly to be expressed upon SAA pathway activation. The baseline expression or changes of interested genes were further confirmed in animals with CorNV using molecular or histological methods. CorNV was induced in Balb/c and C57BL/6 mice by placing either three interrupted 10-0 sutures or a 2 mm filter paper soaked with sodium hydroxide in the central area of the cornea. At desired time points, the corneas were harvested for histology examination or for extraction of mRNA and protein. The mRNA levels of Saa1, Saa3, Fpr2, Mmp2 and Mmp3 in corneas were detected using quantitative reverse transcription-PCR, and SAA3 protein in tissues detected using immunohistochemistry or western blotting.RESULTS:Microarray data analysis revealed that Saa1, Saa3, Fpr2, Mmp2, Mmp3 messengers were readily detected in normal corneas and significantly up-regulated upon CorNV induction. The changes of these five genes were confirmed with real-time PCR assay. On the contrary, other SAA members (Saa2, Saa4, other SAA receptors (Tlr2, Tlr4, Cd36, P2rx7, etc, or other Mmps (Mmp1a, Mmp1b, Mmp9, Mmp10, Mmp13 did not show consistent changes. Immunohistochemistry study and western blotting further confirmed the expression of SAA3 products in normal corneas as well as their up-regulation in corneas with CorNV.CONCLUSION:SAA-FPR2 pathway composing genes were expressed in normal murine corneas and, upon inflammatory stimuli challenge to the corneas, their expressions were up-regulated, suggesting their roles in pathogenesis of CorNV. The potential usefulness of SAA-FPR2 targets in future management of CorNV-related diseases deserves investigation.

Sheng-Wei Ren

2014-04-01

175

Association of a novel human FE65-like protein with the cytoplasmic domain of the beta-amyloid precursor protein.  

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We identified a novel human homologue of the rat FE65 gene, hFE65L, by screening the cytoplasmic domain of beta-amyloid precursor protein (beta PP) with the "interaction trap." The cytoplasmic domains of the beta PP homologues, APLP1 and APLP2 (amyloid precursor-like proteins), were also tested for interaction with hFE65L. APLP2, but not APLP1, was found to interact with hFE65L. We confirmed these interactions in vivo by successfully coimmunoprecipatating endogenous beta PP and APLP2 from mam...

Gue?nette, S. Y.; Chen, J.; Jondro, P. D.; Tanzi, R. E.

1996-01-01

176

Mechanisms of protein oligomerization: inhibitor of functional amyloids templates ?-synuclein fibrillation.  

Science.gov (United States)

Small organic molecules that inhibit functional bacterial amyloid fibers, curli, are promising new antibiotics. Here we investigated the mechanism by which the ring-fused 2-pyridone FN075 inhibits fibrillation of the curli protein CsgA. Using a variety of biophysical techniques, we found that FN075 promotes CsgA to form off-pathway, non-amyloidogenic oligomeric species. In light of the generic properties of amyloids, we tested whether FN075 would also affect the fibrillation reaction of human ?-synuclein, an amyloid-forming protein involved in Parkinson's disease. Surprisingly, FN075 stimulates ?-synuclein amyloid fiber formation as measured by thioflavin T emission, electron microscopy (EM), and atomic force microscopy (AFM). NMR data on (15)N-labeled ?-synuclein show that upon FN075 addition, ?-synuclein oligomers with 7 nm radius form in which the C-terminal 40 residues remain disordered and solvent exposed. The polypeptides in these oligomers contain ?-like secondary structure, and the oligomers are detectable by AFM, EM, and size-exclusion chromatography (SEC). Taken together, FN075 triggers oligomer formation of both proteins: in the case of CsgA, the oligomers do not proceed to fibers, whereas for ?-synuclein, the oligomers are poised to rapidly form fibers. We conclude that there is a fine balance between small-molecule inhibition and templation that depends on protein chemistry. PMID:22260746

Horvath, Istvan; Weise, Christoph F; Andersson, Emma K; Chorell, Erik; Sellstedt, Magnus; Bengtsson, Christoffer; Olofsson, Anders; Hultgren, Scott J; Chapman, Matthew; Wolf-Watz, Magnus; Almqvist, Fredrik; Wittung-Stafshede, Pernilla

2012-02-22

177

Mapping the structure of amyloid nucleation precursors by protein engineering kinetic analysis.  

Science.gov (United States)

Understanding the early molecular mechanisms governing amyloid aggregation is crucial to learn how to prevent it. Here, we used a site-directed mutagenesis approach to explore the molecular mechanism of nucleation of amyloid structure in the N47A Spc-SH3 domain. The changes in the native state stability produced by a series of mutations on each structural element of the domain were uncorrelated with the changes in the aggregation rates, although the overall aggregation mechanism was not altered. Analysis of the thioflavin T initial rates based on a simple kinetic model allowed us to extract thermodynamic magnitudes of the precursor states of nucleation and map the regions of the protein participating in the structure of the amyloidogenic precursors. This structure differs from that of the folding transition state of the SH3 domains, strongly suggesting that the regions of the conformational landscape leading to amyloid formation are divergent from those leading to the native fold. PMID:24394436

Ruzafa, David; Varela, Lorena; Azuaga, Ana I; Conejero-Lara, Francisco; Morel, Bertrand

2014-02-21

178

Stabilization of neurotoxic Alzheimer amyloid-? oligomers by protein engineering  

Science.gov (United States)

Soluble oligomeric aggregates of the amyloid-? peptide (A?) have been implicated in the pathogenesis of Alzheimer’s disease (AD). Although the conformation adopted by A? within these aggregates is not known, a ?-hairpin conformation is known to be accessible to monomeric A?. Here we show that this ?-hairpin is a building block of toxic A? oligomers by engineering a double-cysteine mutant (called A?cc) in which the ?-hairpin is stabilized by an intramolecular disulfide bond. A?40cc and A?42cc both spontaneously form stable oligomeric species with distinct molecular weights and secondary-structure content, but both are unable to convert into amyloid fibrils. Biochemical and biophysical experiments and assays with conformation-specific antibodies used to detect A? aggregates in vivo indicate that the wild-type oligomer structure is preserved and stabilized in A?cc oligomers. Stable oligomers are expected to become highly toxic and, accordingly, we find that ?-sheet-containing A?42cc oligomers or protofibrillar species formed by these oligomers are 50 times more potent inducers of neuronal apoptosis than amyloid fibrils or samples of monomeric wild-type A?42, in which toxic aggregates are only transiently formed. The possibility of obtaining completely stable and physiologically relevant neurotoxic A? oligomer preparations will facilitate studies of their structure and role in the pathogenesis of AD. For example, here we show how kinetic partitioning into different aggregation pathways can explain why A?42 is more toxic than the shorter A?40, and why certain inherited mutations are linked to protofibril formation and early-onset AD. PMID:20713699

Sandberg, Anders; Luheshi, Leila M.; Sollvander, Sofia; Pereira de Barros, Teresa; Macao, Bertil; Knowles, Tuomas P. J.; Biverstal, Henrik; Lendel, Christofer; Ekholm-Petterson, Frida; Dubnovitsky, Anatoly; Lannfelt, Lars; Dobson, Christopher M.; Hard, Torleif

2010-01-01

179

Amyloid fibril formation by a normally folded protein in the absence of denaturants and agitation.  

Science.gov (United States)

The conversion of normally folded proteins into amyloid-like fibrils is an important process in protein chemistry, biology, pathology and biotechnology. This process generally requires harsh conditions, such as pH extremes, organic cosolvents, high temperatures, high pressures or shear forces. Such conditions promote aggregation because they partially unfold structured proteins or allow the sampling of locally unfolded native-like states, both of which possibly represent amyloidogenic states. Here we report the formation of amyloid-like fibrils by the lipase from Pseudomonas sp. under conditions that are close to physiological, that is, in the absence of denaturants and agitation. The resulting aggregates bind thioflavin T and Congo red, causing their characteristic spectral changes observed in the presence of amyloid fibrils. They possess a significant quantity of ?-sheet structure, as detected with Fourier transform infrared and far-UV circular dichroism spectroscopies, and appear fibrillar using transmission electron microscopy. These results indicate that the lipase from Pseudomonas sp. can be a useful model system for the characterization of a key process, such as amyloid fibril formation under physiological conditions. PMID:24053331

Shokri, Maryam Monsef; Ahmadian, Shahin; Bemporad, Francesco; Khajeh, Khosro; Chiti, Fabrizio

2013-12-01

180

Determination of the Proteolytic Cleavage Sites of the Amyloid Precursor-Like Protein 2 by the Proteases ADAM10, BACE1 and ?-Secretase  

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Regulated intramembrane proteolysis of the amyloid precursor protein (APP) by the protease activities ?-, ?- and ?-secretase controls the generation of the neurotoxic amyloid ? peptide. APLP2, the amyloid precursor-like protein 2, is a homolog of APP, which shows functional overlap with APP, but lacks an amyloid ? domain. Compared to APP, less is known about the proteolytic processing of APLP2, in particular in neurons, and the cleavage sites have not yet been determined. APLP2 is cleave...

2011-01-01

 
 
 
 
181

Determination of the proteolytic cleavage sites of the amyloid precursor-like protein 2 by the proteases ADAM10, BACE1 and ?-secretase.  

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Regulated intramembrane proteolysis of the amyloid precursor protein (APP) by the protease activities ?-, ?- and ?-secretase controls the generation of the neurotoxic amyloid ? peptide. APLP2, the amyloid precursor-like protein 2, is a homolog of APP, which shows functional overlap with APP, but lacks an amyloid ? domain. Compared to APP, less is known about the proteolytic processing of APLP2, in particular in neurons, and the cleavage sites have not yet been determined. APLP2 is cleave...

2011-01-01

182

Mice with combined gene knock-outs reveal essential and partially redundant functions of amyloid precursor protein family members.  

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The amyloid precursor protein (APP) involved in Alzheimer's disease is a member of a larger gene family including amyloid precursor-like proteins APLP1 and APLP2. We generated and examined the phenotypes of mice lacking individual or all possible combinations of APP family members to assess potential functional redundancies within the gene family. Mice deficient for the nervous system-specific APLP1 protein showed a postnatal growth deficit as the only obvious abnormality. In contrast to this...

Heber, S.; Herms, J.; Gajic, V.; Hainfellner, J. A.; Aguzzi, A.; Ru?licke, T.; Von Kretzschmar, H.; Von Koch, C.; Sisodia, S.; Tremml, P.; Lipp, H. P.; Wolfer, D. P.; Mu?ller, U.

2000-01-01

183

99mTc-MAMA-chrysamine G, a probe for beta-amyloid protein of Alzheimer's disease  

International Nuclear Information System (INIS)

Chrysamine G (CG), an analogue of Congo red, is known to bind in vitro to the ?-amyloid protein (A? 10-43) and to homogenates of several regions of the brain of Alzheimer's disease (AD) patients. We synthesised a conjugate of 2-(acetamido)-CG with a bis-S-trityl protected monoamide-monoaminedithiol (MAMA-Tr2) tetraligand, which was efficiently deprotected and labelled with a 75% yield with technetium-99m, to obtain 99mTc-MAMA-CG. In mice, 99mTc-MAMA-CG was cleared mainly by the hepatobiliary system, resulting in a fast blood clearance. Brain uptake of 99mTc-MAMA-CG was low. Co-injection with the blood pool tracer iodine-125 human serum albumin (125I-HSA) demonstrated a brain/blood activity ratio for 99mTc-MAMA-CG that was significantly higher than that for 125I-HSA (t test for dependent samples, P99mTc-MAMA-CG to cross the blood-brain barrier. In vitro autoradiography demonstrated pronounced binding of 99mTc-MAMA-CG to ?-amyloid deposits in autopsy sections of the parietal and occipital cortex of an AD patient as compared with controls. Adding 10 ?M Congo red during incubation displaced the binding of 99mTc-MAMA-CG. Congo red staining and autoradiography identified the same lesions. 99mTc-MAMA-CG seems to bind selectively to ?-amyloid deposition in human brain parenchyma and blood vessels in vitro and thus might be a lead compound for further development of a useful tracer agent for the in vivo diagnosis of Alzheimer's disease. (orig.)

184

Neuronal sorting protein-related receptor sorLA/LR11 regulates processing of the amyloid precursor protein  

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sorLA (sorting protein-related receptor) is a type-1 membrane protein of unknown function that is expressed in neurons. Its homology to sorting receptors that shuttle between the plasma membrane, endosomes, and the Golgi suggests a related function in neuronal trafficking processes. Because expression of sorLA is reduced in the brain of patients with Alzheimer's disease (AD), we tested involvement of this receptor in intracellular transport and processing of the amyloid precursor protein (APP...

Andersen, Olav M.; Reiche, Juliane; Schmidt, Vanessa; Gotthardt, Michael; Spoelgen, Robert; Behlke, Joachim; Von Arnim, Christine A. F.; Breiderhoff, Tilman; Jansen, Pernille; Wu, Xin; Bales, Kelly R.; Cappai, Roberto; Masters, Colin L.; Gliemann, Jørgen; Mufson, Elliott J.

2005-01-01

185

Crystal Structure of the E2 Domain of Amyloid Precursor Protein-like Protein 1 in Complex with Sucrose Octasulfate*  

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Missense mutations in the amyloid precursor protein (APP) gene can cause familial Alzheimer disease. It is thought that APP and APP-like proteins (APLPs) may play a role in adhesion and signal transduction because their ectodomains interact with components of the extracellular matrix. Heparin binding induces dimerization of APP and APLPs. To help explain how these proteins interact with heparin, we have determined the crystal structure of the E2 domain of APLP1 in complex with sucrose octasul...

Xue, Yi; Lee, Sangwon; Wang, Yongcheng; Ha, Ya

2011-01-01

186

Total serum protein, serum protein fractions and serum immunoglobulins in colostrum-fed and colostrum-deprived calves.  

Science.gov (United States)

Total serum protein levels, serum protein fraction levels, and specific serum immunoglobulin class or subclass levels were measured in colostrum-fed (CF) and colostrum-deprived (CD) calves during the first 144 hours after birth. Total serum protein values increased at 24 hours in the CF group and then decreased slightly at 144 hours. The increase in total serum protei5) in beta1-, beta2-, and gamma-globulins. The beta2- and gamma-globulin levels decreased by 144 hours, while the serum level of beta1-globulin continued to increase. The CD calves exhibited a significant decrease (P less than 0.05) in total serum protein at 24 hours, folhours, the level of beta1-globulin inlowed by a significant increase (P less than 0.05) at 144 hours. At 24 hours, the level of beta1-globulin decreased slightly, and the level of beta2- and gamma-globulins increased slightly. At 144 creased, and the level of gamma-globulin decreased. The beta2-globulin level did not change. At birth, immunoglobulin (Ig) M was detected in 5 of the 10 calves, IgG1 in 6 of the 10 calves, and IgG2 in 3 of the 10 calves. By 24 hours after birth, all CF calves had detectable levels of IgM, IgG1, and IgG2, and there were significant increases (P less than 0.01) in the mean serum levels of all 3 immunoglobulins. By 144 hours after birth, the serum levels of IgM, IgG1, and IgG2 decreased to various degrees. At 24 hours, the IgM level had not increased in CD calves; however, the level of IgG2 appeared to increase slightly, and the mean IgG1 level increased by approximately 50%. By 144 hours after birth, there was a significant increase (P less than 0.01) in the mean level of serum IgM. The level of IgG, also appeared to increase substantially, while the level of IgG2 appeared to increase slightly. PMID:65932

LaMotte, G B

1977-02-01

187

Self-assembly of protein amyloid: a competition between amorphous and ordered aggregation  

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Protein aggregation in the form of amyloid fibrils has important biological and technological implications. Although the self-assembly process is highly efficient, aggregates not in the fibrillar form would also occur and it is important to include these disordered species when discussing the thermodynamic equilibrium behavior of the system. Here, we initiate such a task by considering a mixture of monomeric proteins and the corresponding aggregates in the disordered form (m...

Lee, Chiu Fan

2008-01-01

188

Amyloid Precursor-like Protein 2 Association with HLA Class I Molecules  

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Amyloid precursor-like protein 2 (APLP2) is a ubiquitously expressed protein. The previously demonstrated functions for APLP2 include binding to the mouse major histocompatibility complex (MHC) class I molecule H-2Kd and down regulating its cell surface expression. In this study, we have investigated the interaction of APLP2 with the human leukocyte antigen (HLA) class I molecule in human tumor cell lines. APLP2 was readily detected in pancreatic, breast, and prostate tumor lines, although it...

Tuli, Amit; Sharma, Mahak; Wang, Xiaojian; Simone, Laura C.; Capek, Haley L.; Cate, Steven; Hildebrand, William H.; Naslavsky, Naava; Caplan, Steve; Solheim, Joyce C.

2009-01-01

189

Knockdown of Amyloid Precursor Protein in Zebrafish Causes Defects in Motor Axon Outgrowth  

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Amyloid precursor protein (APP) plays a pivotal role in Alzheimer’s disease (AD) pathogenesis, but its normal physiological functions are less clear. Combined deletion of the APP and APP-like protein 2 (APLP2) genes in mice results in post-natal lethality, suggesting that APP performs an essential, if redundant, function during embryogenesis. We previously showed that injection of antisense morpholino to reduce APP levels in zebrafish embryos caused convergent-extension defects. Here we rep...

Song, Ping; Pimplikar, Sanjay W.

2012-01-01

190

Turnover of amyloid precursor protein family members determines their nuclear signaling capability  

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The amyloid precursor protein (APP) as well as its homologues, APP-like protein 1 and 2 (APLP1 and APLP2), are cleaved by ?-, ?-, and ?-secretases, resulting in the release of their intracellular domains (ICDs). We have shown that the APP intracellular domain (AICD) is transported to the nucleus by Fe65 where they jointly bind the histone acetyltransferase Tip60 and localize to spherical nuclear complexes (AFT complexes), which are thought to be sites of transcription. We have now analyzed...

Gersbacher, Manuel T.; Goodger, Zoe? V.; Trutzel, Annette; Bundschuh, Diana; Nitsch, Roger M.; Konietzko, Uwe

2013-01-01

191

Proteolyse des Amyloid-Vorläufer-Protein-verwandten APLP2 durch alpha-Sekretasen  

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Die im Laufe der Evolution konservierte Genfamilie des Amyloid-Vorläufer-Proteins APP beinhaltet sowohl bei der Maus als auch beim Menschen die beiden APP-ähnlichen ProteineAPLP1 und APLP2. Ziel dieser Arbeit war es, die proteolytische Prozessierung des APLP2 zu charakterisieren und die beteiligten Proteasen aufzuzeigen. Ausgehend von Stimulations- und Inhibitionsversuchen wurde die Metzincin-Familie der Metalloproteinasen als APLP2-Proteasen identifiziert. Durch Überexpression von ADAM10 ...

Endres, Kristina

2005-01-01

192

Correlations between serum levels of beta amyloid, cerebrospinal levels of tau and phospho tau, and delayed response tasks in young and aged cynomolgus monkeys (Macaca fascicularis)  

DEFF Research Database (Denmark)

In an attempt to explore cynomolgus monkeys as an animal model for Alzheimer's disease, the present study focused on the Alzheimer's biomarkers beta amyloid 1-42 (A?42 ) in serum, and total tau (t-tau) and phosphorylated tau (p-tau) levels in cerebrospinal fluid.

Darusman, Huda Shalahudin; Sajuthi, D

2013-01-01

193

Adipose Tissue-Derived Human Serum Amyloid A Does Not Affect Atherosclerotic Lesion Area in hSAA1+/?/ApoE?/? Mice  

Science.gov (United States)

Chronically elevated serum levels of serum amyloid A (SAA) are linked to increased risk of cardiovascular disease. However, whether SAA is directly involved in atherosclerosis development is still not known. The aim of this study was to investigate the effects of adipose tissue-derived human SAA on atherosclerosis in mice. hSAA1+/? transgenic mice (hSAA1 mice) with a specific expression of human SAA1 in adipose tissue were bred with ApoE-deficient mice. The hSAA1 mice and their wild type (wt) littermates were fed normal chow for 35 weeks. At the end of the experiment, the mice were euthanized and blood, gonadal adipose tissue and aortas were collected. Plasma levels of SAA, cholesterol and triglycerides were measured. Atherosclerotic lesion areas were analyzed in the aortic arch, the thoracic aorta and the abdominal aorta in en face preparations of aorta stained with Sudan IV. The human SAA protein was present in plasma from hSAA1 mice but undetectable in wt mice. Similar plasma levels of cholesterol and triglycerides were observed in hSAA1 mice and their wt controls. There were no differences in atherosclerotic lesion areas in any sections of the aorta in hSAA1 mice compared to wt mice. In conclusion, our data suggest that adipose tissue-derived human SAA does not influence atherosclerosis development in mice. PMID:24751653

Ahlin, Sofie; Olsson, Maja; Wilhelmson, Anna S.; Skalen, Kristina; Boren, Jan; Carlsson, Lena M. S.; Svensson, Per-Arne; Sjoholm, Kajsa

2014-01-01

194

Adipose tissue-derived human serum amyloid a does not affect atherosclerotic lesion area in hSAA1+/-/ApoE-/- mice.  

Science.gov (United States)

Chronically elevated serum levels of serum amyloid A (SAA) are linked to increased risk of cardiovascular disease. However, whether SAA is directly involved in atherosclerosis development is still not known. The aim of this study was to investigate the effects of adipose tissue-derived human SAA on atherosclerosis in mice. hSAA1+/- transgenic mice (hSAA1 mice) with a specific expression of human SAA1 in adipose tissue were bred with ApoE-deficient mice. The hSAA1 mice and their wild type (wt) littermates were fed normal chow for 35 weeks. At the end of the experiment, the mice were euthanized and blood, gonadal adipose tissue and aortas were collected. Plasma levels of SAA, cholesterol and triglycerides were measured. Atherosclerotic lesion areas were analyzed in the aortic arch, the thoracic aorta and the abdominal aorta in en face preparations of aorta stained with Sudan IV. The human SAA protein was present in plasma from hSAA1 mice but undetectable in wt mice. Similar plasma levels of cholesterol and triglycerides were observed in hSAA1 mice and their wt controls. There were no differences in atherosclerotic lesion areas in any sections of the aorta in hSAA1 mice compared to wt mice. In conclusion, our data suggest that adipose tissue-derived human SAA does not influence atherosclerosis development in mice. PMID:24751653

Ahlin, Sofie; Olsson, Maja; Wilhelmson, Anna S; Skålén, Kristina; Borén, Jan; Carlsson, Lena M S; Svensson, Per-Arne; Sjöholm, Kajsa

2014-01-01

195

Oxidative stress activates a positive feedback between the ?- and ?-secretase cleavages of the ?-amyloid precursor protein  

Science.gov (United States)

Sequential cleavages of the ?-amyloid precursor protein cleaving enzyme 1 (BACE1) by ?-secretase and ?-secretase generate the amyloid ?-peptides, believed to be responsible of synaptic dysfunction and neuronal cell death in Alzheimer's disease (AD). Levels of BACE1 are increased in vulnerable regions of the AD brain, but the underlying mechanism is unknown. Here we show that oxidative stress (OS) stimulates BACE1 expression by a mechanism requiring ?-secretase activity involving the c-jun N-terminal kinase (JNK)/c-jun pathway. BACE1 levels are increased in response to OS in normal cells, but not in cells lacking presenilins or amyloid precursor protein. Moreover, BACE1 is induced in association with OS in the brains of mice subjected to cerebral ischaemia/reperfusion. The OS-induced BACE1 expression correlates with an activation of JNK and c-jun, but is absent in cultured cells or mice lacking JNK. Our findings suggest a mechanism by which OS induces BACE1 transcription, thereby promoting production of pathological levels of amyloid ? in AD. PMID:18005001

Tamagno, Elena; Guglielmotto, Michela; Aragno, Manuela; Borghi, Roberta; Autelli, Riccardo; Giliberto, Luca; Muraca, Giuseppe; Danni, Oliviero; Zhu, Xiongwei; Smith, Mark A.; Perry, George; Jo, Dong-Gyu; Mattson, Mark P.; Tabaton, Massimo

2008-01-01

196

Cerebrospinal fluid soluble amyloid-? protein precursor as a potential novel biomarkers of Alzheimer's disease.  

Science.gov (United States)

In this report, we confirm our previous findings of increased concentrations of soluble amyloid-? protein precursor (sA?PP) in cerebrospinal fluid (CSF) of patients with Alzheimer's disease (AD) and mild cognitive impairment (MCI) in a large cohort of patients (n = 314), not overlapping with those of our previous study, and we extend our observations by including a control group of participants with normal cognition. In addition, we investigate the effects of age, the APOE?4 genotype, and the blood-CSF barrier function on the concentrations of sA?PP? and sA?PP?. The study participants were categorized according to clinical-neuropsychological criteria, supported by CSF neurochemical dementia diagnostics (NDD) analyses. sA?PP? concentrations in the AD group (132.0 ± 44.8) were significantly higher than in the control group (105.3 ± 37.3, p significantly from the MCI-O (97.3 ± 34.3, p significantly higher than in the control group (129.9 ± 44.6, p significantly from the MCI-O (127.8 ± 46.2, p 0.99). We observed highly significant correlation of the two sA?PP forms. Age and the CSF-serum albumin ratio were significant albeit weak predictors of the sA?PP? and sA?PP? concentrations, while carrying the APOE?4 allele did not influenced the levels of the sA?PP forms. Taken together, the results strongly suggest that CSF sA?PP concentrations may be considered as an extension of already available NDD tools. PMID:21971403

Lewczuk, Piotr; Popp, Julius; Lelental, Natalia; Kölsch, Heike; Maier, Wolfgang; Kornhuber, Johannes; Jessen, Frank

2012-01-01

197

Bilobalide regulates soluble amyloid precursor protein release via phosphatidyl inositol 3 kinase-dependent pathway.  

Science.gov (United States)

Bilobalide (BB) is a sesquiterpenoid extracted from Ginkgo biloba leaves. An increasing number of studies have demonstrated its neuroprotective effects. The neuroprotective mechanisms may be associated with modulation of intracellular signaling cascades such as the phosphatidyl inositol 3-kinase (PI3K) pathway. Using differentiated SH-SY5Y cells, this study investigated whether BB modulation of intracellular signaling pathways, such as the protein kinase C (PKC) and PI3K pathways, contributes to amyloid precursor protein (APP) metabolism, a key event in the pathogenesis of Alzheimer's disease (AD). We demonstrated in this study that BB enhanced the secretion of ?-secretase-cleaved soluble amyloid precursor protein (sAPP?, a by-product of non-amyloidogenic processing of APP) and decreased the ? amyloid protein (A?, a by-product of amyloidogenic processing of APP) via PI3K-dependent pathway. The PI3K pathway mediated the rapid effect of BB on APP processing possibly via regulation of intracellular APP trafficking. After longer time BB incubation (12h), this effect was reinforced by PI3K pathway-mediated up-regulation of disintegrin and metalloproteinase domain-containing protein 10 (ADAM10, an ?-secretase candidate). Given the strong association between APP metabolism and AD pathogenesis, the ability of BB to regulate APP processing suggests its potential use in AD prevention. PMID:21679734

Shi, Chun; Wu, Fengming; Xu, Jie; Zou, Juntao

2011-08-01

198

A novel mechanism for the regulation of amyloid precursor protein metabolism.  

Science.gov (United States)

Modifier of cell adhesion protein (MOCA; previously called presenilin [PS] binding protein) is a DOCK180-related molecule, which interacts with PS1 and PS2, is localized to brain areas involved in Alzheimer's disease (AD) pathology, and is lost from the soluble fraction of sporadic Alzheimer's disease (AD) brains. Because PS1 has been associated with gamma-secretase activity, MOCA may be involved in the regulation of beta-amyloid precursor protein (APP) processing. Here we show that the expression of MOCA decreases both APP and amyloid beta-peptide secretion and lowers the rate of cell-substratum adhesion. In contrast, MOCA does not lower the secretion of amyloid precursor-like protein (APLP) or several additional type 1 membrane proteins. The phenotypic changes caused by MOCA are due to an acceleration in the rate of intracellular APP degradation. The effect of MOCA expression on the secretion of APP and cellular adhesion is reversed by proteasome inhibitors, suggesting that MOCA directs nascent APP to proteasomes for destruction. It is concluded that MOCA plays a major role in APP metabolism and that the effect of MOCA on APP secretion and cell adhesion is a downstream consequence of MOCA-directed APP catabolism. This is a new mechanism by which the expression of APP is regulated. PMID:12093789

Chen, Qi; Kimura, Hideo; Schubert, David

2002-07-01

199

Expression and distribution of amyloid precursor protein-like protein-2 in Alzheimer's disease and in normal brain.  

Science.gov (United States)

Amyloid precursor-like protein-2 (APLP-2) belongs to a family of homologous amyloid precursor-like proteins. In the present study we report on the expression and distribution of APLP-2 in fetal and adult human brain and in brains of patients with Alzheimer's disease. We demonstrate that APLP-2 mRNAs encoding isoforms predicted to undergo post-translational modification by chondroitin sulfate glycosaminoglycans are elevated in fetal and aging brains relative to the brains of young adults. Immunocytochemical labeling with APLP-2-specific antibodies demonstrates APLP-2 immunoreactivity in cytoplasmic compartments in neurons and astrocytes, in large part overlapping the distribution of the amyloid precursor protein. In Alzheimer's disease brain, APLP-2 antibodies also label a subset of neuritic plaques. APLP-2 immunoreactivity is particularly conspicuous in large dystrophic neurites that also label with antibodies specific for APP and chromogranin A. In view of the age-dependent increase in levels of chondroitin sulfate glycosaminoglycan-modified forms of APLP-2 in aging brain and the accumulation of APLP-2 in dystrophic presynaptic elements, we suggest that APLP-2 may play roles in neuronal sprouting or in the aggregation, deposition, and/or persistence of beta-amyloid deposits. PMID:8863657

Crain, B J; Hu, W; Sze, C I; Slunt, H H; Koo, E H; Price, D L; Thinakaran, G; Sisodia, S S

1996-10-01

200

NMDA receptor/amyloid precursor protein interactions: A comparison between wild-type and amyloid precursor protein mutations associated with familial Alzheimer's disease  

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Two recent reports showed that amyloid precursor protein (APP) may contribute to postsynaptic mechanisms via the regulation of the surface trafficking of excitatory N-methyl-d-aspartate (NMDA) receptors. Here we have investigated the interactions and surface trafficking of NR1-1a/NR2A and NR1-1a/NR2B NMDA receptor subtypes with three APP mutations linked to familial Alzheimer's disease, APP695Indiana, APP695London and APP695Swedish. Flag-tagged mutated APP695s were generated and shown to be e...

Innocent, N.; Cousins, S. L.; Stephenson, F. A.

2012-01-01

 
 
 
 
201

Mapping of the gene encoding the ?-amyloid precursor protein and its relationship to the Down syndrome region of chromosome 21  

International Nuclear Information System (INIS)

The gene encoding the ?-amyloid precursor protein has been assigned to human chromosome 21, as has a gene responsible for at least some cases of familial Alzheimer disease. Linkage studies strongly suggest that the ?-amyloid precursor protein and the product corresponding to familial Alzheimer disease are from two genes, or at least that several million base pairs of DNA separate the markers. The precise location of the ?-amyloid precursor protein gene on chromosome 21 has not yet been determined. Here the authors show, by using a somatic-cell/hybrid-cell mapping panel, in situ hybridization, and transverse-alternating-field electrophoresis, that the ?-amyloid precursor protein gene is located on chromosome 21 very near the 21q21/21q/22 border and probably within the region of chromosome 21 that, when trisomic, results in Down syndrome

202

Amyloid Precursor Proteins Anchor CPEB to Membranes and Promote Polyadenylation-Induced Translation  

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The cytoplasmic polyadenylation element (CPE) binding factor, CPEB, is a sequence-specific RNA binding protein that controls polyadenylation-induced translation in germ cells and at postsynaptic sites of neurons. A yeast two-hybrid screen with a mouse brain cDNA library identified the transmembrane amyloid precursor-like protein 1 (APLP1) as a CPEB-interacting factor. CPEB binds the small intracellular domain (ICD) of APLP1 and the related proteins APLP2 and APP. These proteins promote polyad...

Cao, Quiping; Huang, Yi-shuian; Kan, Ming-chung; Richter, Joel D.

2005-01-01

203

Mechanism for Amyloid Precursor-like Protein 2 Enhancement of Major Histocompatibility Complex Class I Molecule Degradation*  

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Earlier studies have demonstrated interaction of the murine major histocompatibility complex (MHC) class I molecule Kd with amyloid precursor-like protein 2 (APLP2), a ubiquitously expressed member of the amyloid precursor protein family. Our current findings indicate that APLP2 is internalized in a clathrin-dependent manner, as shown by utilization of inhibitors of the clathrin pathway. Furthermore, we demonstrated that APLP2 and Kd bind at the cell surface and are internalized together. The...

Tuli, Amit; Sharma, Mahak; Capek, Haley L.; Naslavsky, Naava; Caplan, Steve; Solheim, Joyce C.

2009-01-01

204

Molecular cloning, characterization of one key molecule of teleost innate immunity from orange-spotted grouper (Epinephelus coioides): serum amyloid A.  

Science.gov (United States)

The orange-spotted grouper (Epinephelus coioides), a favorite marine food fish, is widely cultured in China and Southeast Asian countries. However, little is known about its acute phase response (APR) caused by viral diseases. Serum amyloid A (SAA) is a major acute phase protein (APP). In this study, a new SAA homologous (EcSAA) gene was cloned from grouper, E. coioides, by rapid amplification of cDNA ends (RACE) PCR. The full-length cDNA sequence of SAA was 508 bp and contained a 363 bp open reading frame (ORF) coding for a protein of 121 aa. Similar to other fish known SAA genes, the EcSAA gene contained four exons and three introns. Quantitative real-time PCR analysis revealed that EcSAA mRNA is predominately expressed in liver and gill of grouper. Furthermore, the expression of EcSAA was differentially up-regulated in liver after infection with Staphyloccocus aureus, Vibrio vulnificus, Vibrio parahaemolyticus, Saccharomyces cerevisiae and Singapore grouper iridovirus (SGIV). Recombinant EcSAA (rEcSAA) was expressed in Escherichia BL21 (DE3) and purified for mouse anti-EcSAA serum preparation. The rEcSAA fusion protein was demonstrated to bind to all tested bacteria and yeast, and inhibit the replication of SGIV. Overexpression of EcSAA in grouper spleen (GS) cells could also inhibit the replication of SGIV. These results suggest that EcSAA may be an important molecule in the innate immunity of grouper. PMID:23178260

Wei, Jingguang; Guo, Minglan; Ji, Huasong; Qin, Qiwei

2013-01-01

205

Sertoli cells secrete both testis-specific and serum proteins.  

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The secretions of the Sertoli cell were examined with two polyvalent antisera--one prepared against proteins in rat serum and the other against testis-specific proteins in rete testis fluid. These antisera detected 12 serum and 9 testis-specific proteins in rete testis fluid. To determine the origin of these proteins, primary cultures enriched in Sertoli cells were incubated with [35S]methionine, and the radiolabeled proteins in the medium were immunoprecipitated. Gel electrophoresis of the t...

Wright, W. W.; Musto, N. A.; Mather, J. P.; Bardin, C. W.

1981-01-01

206

Bioluminescence imaging reveals inhibition of tumor cell proliferation by Alzheimer's amyloid ? protein  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Cancer and Alzheimer's disease (AD are two seemingly distinct diseases and rarely occur simultaneously in patients. To explore molecular determinants differentiating pathogenic routes towards AD or cancer, we investigate the role of amyloid ? protein (A? on multiple tumor cell lines that are stably expressing luciferase (human glioblastoma U87; human breast adenocarcinoma MDA-MB231; and mouse melanoma B16F. Results Quantification of the photons emitted from the MDA-MB231 or B16F cells revealed a significant inhibition of cell proliferation by the conditioning media (CM derived from amyloid precursor protein (APP over-expressing cells. The inhibition of U87 cells was observed only after the media was conditioned for longer than 2 days with APP over-expressing cells. Conclusion Our results suggest that A? plays an inhibitory role in tumor cell proliferation; this effect could depend on the type of tumor cells and amount of A?.

Zhao Hong

2009-06-01

207

Serum amyloid A induces interleukin-6 in dermal fibroblasts via Toll-like receptor 2, interleukin-1 receptor-associated kinase 4 and nuclear factor-?B.  

Science.gov (United States)

Systemic sclerosis is an autoimmune idiopathic connective tissue disease, characterized by vasculopathy, inflammation and fibrosis. There appears to be a link between inflammation and fibrosis, although the exact nature of the relationship is unknown. Serum amyloid A (SAA) is an acute-phase protein that is elevated up to 1000-fold in times of infection or inflammation. This acute-phase reactant, as well as being a marker of inflammation, may initiate signals in a cytokine-like manner, possibly through toll-like receptors (TLRs) promoting inflammation. This study addressed the role of SAA in initiating interleukin-6 (IL-6) production in dermal fibroblasts and the role of TLR2 in this system. We show that SAA induces IL-6 secretion in healthy dermal fibroblasts and that blockade of TLR2 with a neutralizing antibody to TLR2 or specific small interfering RNA attenuated the SAA-induced IL-6 secretion and that this was also mediated through the TLR adaptor protein IL-1 receptor-associated kinase 4. The effect is nuclear factor-?B-mediated because blockade of nuclear factor-?B reduced the induction. We also demonstrate that dermal fibroblasts express TLR2; this is functional and over-expressed in the fibroblasts of patients with systemic sclerosis. Taken together these data suggest that SAA is a danger signal that initiates IL-6 signalling in systemic sclerosis via enhanced TLR2 signalling. PMID:24476318

O'Reilly, Steven; Cant, Rachel; Ciechomska, Marzena; Finnigan, James; Oakley, Fiona; Hambleton, Sophie; van Laar, Jacob M

2014-11-01

208

Effect of x-radiation on serum proteins  

International Nuclear Information System (INIS)

The inbred albine mice were exposed to 315 roentgen whole-body X-radiation at a rate of 22.5 roentgen per minute. Maximum decrease in the serum albumin and maximum increase in the serum globulin fractions were seen two hours after irradiation. Both of the serum proteins return to approximately normal levels within six hours. (author)

209

The Alzheimer's Disease-Associated Amyloid \\(\\beta\\)-Protein Is an Antimicrobial Peptide  

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Background: The amyloid \\(\\beta\\)-protein (A\\(\\beta\\)) is believed to be the key mediator of Alzheimer's disease (AD) pathology. A\\(\\beta\\) is most often characterized as an incidental catabolic byproduct that lacks a normal physiological role. However, A\\(\\beta\\) has been shown to be a specific ligand for a number of different receptors and other molecules, transported by complex trafficking pathways, modulated in response to a variety of environmental stressors, and able to induce pro-infla...

Soscia, Stephanie; Kirby, James Edward; Washicosky, Kevin J.; Tucker, Stephanie; Ingelsson, Martin; Hyman, Bradley Theodore; Burton, Mark A.; Duong, Scott; Tanzi, Rudolph Emile; Moir, Robert D.; Goldstein, Lee E.

2010-01-01

210

The Alzheimer's Disease-Associated Amyloid ?-Protein Is an Antimicrobial Peptide  

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Background: The amyloid ?-protein (A?) is believed to be the key mediator of Alzheimer's disease (AD) pathology. A? is most often characterized as an incidental catabolic byproduct that lacks a normal physiological role. However, A? has been shown to be a specific ligand for a number of different receptors and other molecules, transported by complex trafficking pathways, modulated in response to a variety of environmental stressors, and able to induce pro-inflammatory activities. Methodol...

Washicosky, Kevin J.; Tucker, Stephanie M.; Ingelsson, Martin; Burton, Mark A.; Duong, Scott; Kirby, James Edward; Hyman, Bradley Theodore; Goldstein, Lee E.; Tanzi, Rudolph Emile; Moir, Robert D.

2010-01-01

211

Exploring the Folding Pathways of Proteins through Energy Landscape Sampling: Application to Alzheimer's ?-Amyloid Peptide  

Directory of Open Access Journals (Sweden)

Full Text Available The determination of the folding mechanisms of proteins is critical to understand the topological change that can propagate Alzheimer's and prion diseases. The associated folding time scale generally precludes the use of molecular dynamics simulations. Here we present the details of the activation-relaxation simulations using the generic OPEP energy model. We illustrate the strengths of our approach by studying the folding of a dimer of the Alzheimer's ?-amyloid peptide.

Sebastien Santini

2003-09-01

212

Higher susceptibility to amyloid fibril formation of the recombinant ovine prion protein modified by transglutaminase.  

Science.gov (United States)

Prion proteins are known as the main agents of transmissible spongiform encephalopathies affecting humans as well as animals. A recombinant ovine prion protein was found to be in vitro able to act as an effective substrate for a microbial isoform of transglutaminase, an enzyme catalyzing the formation of isopeptide bonds inside the proteins. We proved that transglutaminase modifies the structure of the prion protein by leading to the formation of three intra-molecular crosslinks and that the crosslinked protein form is more competent in amyloid formation compared to the unmodified one. In addition, the crosslinked prion protein was shown also to be more resistant to proteinase K digestion. Our findings suggest a possible use of transglutaminase in stabilizing the prion protein three-dimensional structure in order to investigate the molecular basis of the conversion of the protein into its pathological form. PMID:22705206

Sorrentino, Angela; Giosafatto, Concetta Valeria L; Sirangelo, Ivana; De Simone, Carmela; Di Pierro, Prospero; Porta, Raffaele; Mariniello, Loredana

2012-10-01

213

Sensing of Proteins in Human Serum using Nanoparticle-Green Fluorescent Protein Conjugates  

Science.gov (United States)

There is a direct correlation between protein levels and disease states in human serum making it an attractive target for sensors and diagnostics. However this is made challenging because serum features more than 20,000 proteins with an overall protein content of greater than 1 mM. Here we report a hybrid synthetic-biomolecule based sensor that uses green fluorescent protein-nanoparticle arrays to detect proteins at biorelevant concentrations in both buffer and human serum. Distinct and reproducible fluorescence response patterns were obtained from five serum proteins (human serum albumin, immunoglobulin G, transferrin, fibrinogen and ?-antitrypsin) in buffer and when spiked into human serum. Using linear discriminant analysis we identified these proteins with an identification accuracy of 100% in buffer and 97% in human serum. The arrays were also able to discriminate between different concentrations of the same protein as well as a mixture of different proteins in human serum. PMID:20161380

DE, MRINMOY; RANA, SUBINOY; AKPINAR, HANDAN; MIRANDA, OSCAR R.; ARVIZO, ROCHELLE R.; BUNZ, UWE H. F.; ROTELLO, VINCENT M.

2009-01-01

214

Sensing of proteins in human serum using conjugates of nanoparticles and green fluorescent protein.  

Science.gov (United States)

There is a direct correlation between protein levels and disease states in human serum, which makes it an attractive target for sensors and diagnostics. However, this is challenging because serum features more than 20,000 proteins, with an overall protein content greater than 1 mM. Here we report a sensor based on a hybrid synthetic-biomolecule that uses arrays of green fluorescent protein and nanoparticles to detect proteins at biorelevant concentrations in both buffer and human serum. Distinct and reproducible fluorescence-response patterns were obtained from five serum proteins (human serum albumin, immunoglobulin G, transferrin, fibrinogen and a-antitrypsin), both in buffer and when spiked into human serum. Using linear discriminant analysis we identified these proteins with an identification accuracy of 100% in buffer and 97% in human serum. The arrays were also able to discriminate between different concentrations of the same protein, as well as a mixture of different proteins in human serum. PMID:20161380

De, Mrinmoy; Rana, Subinoy; Akpinar, Handan; Miranda, Oscar R; Arvizo, Rochelle R; Bunz, Uwe H F; Rotello, Vincent M

2009-09-01

215

Hydrolysis of the amyloid prion protein and nonpathogenic meat and bone meal by anaerobic thermophilic prokaryotes and streptomyces subspecies.  

Science.gov (United States)

Transmissible spongiform encephalopathies are caused by accumulation of highly resistant misfolded amyloid prion protein PrPres and can be initiated by penetration of such pathogen molecules from infected tissue to intact organism. Decontamination of animal meal containing amyloid prion protein is proposed thanks to the use of proteolytic enzymes secreted by thermophilic bacteria Thermoanaerobacter, Thermosipho, and Thermococcus subsp. and mesophilic soil bacteria Streptomyces subsp. Keratins alpha and beta, which resemble amyloid structures, were used as the substrates for the screening for microorganisms able to grow on keratins and producing efficient proteases specific for hydrolysis of beta-sheeted proteic structures, hence amyloids. Secretion of keratin-degrading proteases was evidenced by a zymogram method. Enzymes from thermophilic strains VC13, VC15, and S290 and Streptomyces subsp. S6 were strongly active against amyloid recombinant ovine prion protein and animal meal proteins. The studied proteases displayed broad primary specificities hydrolyzing low molecular mass peptide model substrates. Strong amyloidolytic activity of detected proteases was confirmed by experiments of hydrolysis of PrPres in SAFs produced from brain homogenates of mice infected with the 6PB1 BSE strain. The proteases from Thermoanaerobacter subsp. S290 and Streptomyces subsp. S6 are the best candidates for neutralization/elimination of amyloids in meat and bone meal and other protein-containing substances and materials. PMID:15453713

Tsiroulnikov, Kirill; Rezai, Human; Bonch-Osmolovskaya, Elisaveta; Nedkov, Peter; Gousterova, Adriana; Cueff, Valérie; Godfroy, Anne; Barbier, Georges; Métro, François; Chobert, Jean-Marc; Clayette, Pascal; Dormont, Dominique; Grosclaude, Jeanne; Haertlé, Thomas

2004-10-01

216

Unraveling the Early Events of Amyloid-? Protein (A? Aggregation: Techniques for the Determination of A? Aggregate Size  

Directory of Open Access Journals (Sweden)

Full Text Available The aggregation of proteins into insoluble amyloid fibrils coincides with the onset of numerous diseases. An array of techniques is available to study the different stages of the amyloid aggregation process. Recently, emphasis has been placed upon the analysis of oligomeric amyloid species, which have been hypothesized to play a key role in disease progression. This paper reviews techniques utilized to study aggregation of the amyloid-? protein (A? associated with Alzheimer’s disease. In particular, the review focuses on techniques that provide information about the size or quantity of oligomeric A? species formed during the early stages of aggregation, including native-PAGE, SDS-PAGE, Western blotting, capillary electrophoresis, mass spectrometry, fluorescence correlation spectroscopy, light scattering, size exclusion chromatography, centrifugation, enzyme-linked immunosorbent assay, and dot blotting.

N. Elizabeth Pryor

2012-03-01

217

Serum low-molecular-weight protein fractionation for biomarker discovery.  

Science.gov (United States)

Protein biomarkers provide the key diagnostic information for the detection of disease, risk of disease progression, and a patient's likely response to drug therapy. Potential biomarkers exist in biofluids, such as serum, urine, and cerebrospinal fluid. Unfortunately, discovering and validating protein biomarkers are hindered by the presence of high-molecular-weight proteins, such as serum albumin and immunoglobulins, which comprise 90% of the proteins present in these samples. High-abundance, high-molecular-weight proteins mask the low-molecular-weight (LMW) proteins and peptides using conventional protein detection methods. Candidate biomarkers are believed to exist in very low concentrations and comprise less than 1% of serum proteins, and may be highly labile as well. Therefore, it is imperative to isolate and enrich LMW proteins from complex mixtures for biomarker discovery. This chapter describes a continuous -elution electrophoresis method, based on molecular weight sieving, to isolate specific molecular weight fractions for mass spectrometric, western blotting, or protein array analysis. PMID:22081349

VanMeter, Amy J; Camerini, Serena; Polci, Maria Letizia; Tessitore, Alessandra; Trivedi, Nishant; Heiby, Michael; Kamal, Yasmin; Hansen, Jonathan; Zhou, Weidong

2012-01-01

218

Trans fatty acids enhance amyloidogenic processing of the Alzheimer amyloid precursor protein (APP).  

Science.gov (United States)

Hydrogenation of oils and diary products of ruminant animals leads to an increasing amount of trans fatty acids in the human diet. Trans fatty acids are incorporated in several lipids and accumulate in the membrane of cells. Here we systematically investigate whether the regulated intramembrane proteolysis of the amyloid precursor protein (APP) is affected by trans fatty acids compared to the cis conformation. Our experiments clearly show that trans fatty acids compared to cis fatty acids increase amyloidogenic and decrease nonamyloidogenic processing of APP, resulting in an increased production of amyloid beta (A?) peptides, main components of senile plaques, which are a characteristic neuropathological hallmark for Alzheimer's disease (AD). Moreover, our results show that oligomerization and aggregation of A? are increased by trans fatty acids. The mechanisms identified by this in vitro study suggest that the intake of trans fatty acids potentially increases the AD risk or causes an earlier onset of the disease. PMID:22209004

Grimm, Marcus O W; Rothhaar, Tatjana L; Grösgen, Sven; Burg, Verena K; Hundsdörfer, Benjamin; Haupenthal, Viola J; Friess, Petra; Kins, Stefan; Grimm, Heike S; Hartmann, Tobias

2012-10-01

219

Amyloid precursor proteins inhibit heme oxygenase activity and augment neurotoxicity in Alzheimer's disease.  

Science.gov (United States)

Amyloid precursor protein (APP) generates the beta-amyloid peptide, postulated to participate in the neurotoxicity of Alzheimer's disease. We report that APP and APLP bind to heme oxygenase (HO), an enzyme whose product, bilirubin, is antioxidant and neuroprotective. The binding of APP inhibits HO activity, and APP with mutations linked to the familial Alzheimer's disease (FAD) provides substantially greater inhibition of HO activity than wild-type APP. Cortical cultures from transgenic mice expressing Swedish mutant APP have greatly reduced bilirubin levels, establishing that mutant APP inhibits HO activity in vivo. Oxidative neurotoxicity is markedly greater in cerebral cortical cultures from APP Swedish mutant transgenic mice than wild-type cultures. These findings indicate that augmented neurotoxicity caused by APP-HO interactions may contribute to neuronal cell death in Alzheimer's disease. PMID:11144356

Takahashi, M; Doré, S; Ferris, C D; Tomita, T; Sawa, A; Wolosker, H; Borchelt, D R; Iwatsubo, T; Kim, S H; Thinakaran, G; Sisodia, S S; Snyder, S H

2000-11-01

220

The molecular link between beta- and gamma-secretase activity on the amyloid beta precursor protein.  

Science.gov (United States)

Alzheimer's disease (AD) is characterized by an accumulation in the brain of amyloid beta peptides (Abeta). The production of Abeta requires two sequential cleavages induced by beta- and gamma-secretases on the beta-amyloid precursor protein (APP). Altered activity of these secretases is involved in the pathogenesis of AD. The expression and activity of beta-secretase (BACE1) is augmented in the brain in late-onset sporadic AD. Mutant presenilin 1 (PS1), the major genetic defect of early-onset familial AD (FAD), alters the activity of gamma-secretase, leading to increased production of Abeta42. Here we review the role of oxidative stress as a molecular link between the beta- and the gamma-secretase activities, and provide a mechanistic explanation of the pathogenesis of sporadic late-onset AD. We also discuss evidence for a role of the same mechanism in the pathogenesis of familial AD carrying PS1 mutations. PMID:17604999

Tabaton, M; Tamagno, E

2007-09-01

 
 
 
 
221

Focally Elevated Creatine Detected in Amyloid Precursor Protein (APP) Transgenic Mice and Alzheimer Disease Brain Tissue  

Energy Technology Data Exchange (ETDEWEB)

The creatine/phosphocreatine system, regulated by creatine kinase, plays an important role in maintaining energy balance in the brain. Energy metabolism and the function of creatine kinase are known to be affected in Alzheimer diseased brain and in cells exposed to the {beta}-amyloid peptide. We used infrared microspectroscopy to examine hippocampal, cortical, and caudal tissue from 21-89-week-old transgenic mice expressing doubly mutant (K670N/M671L and V717F) amyloid precursor protein and displaying robust pathology from an early age. Microcrystalline deposits of creatine, suggestive of perturbed energetic status, were detected by infrared microspectroscopy in all animals with advanced plaque pathology. Relatively large creatine deposits were also found in hippocampal sections from post-mortem Alzheimer diseased human brain, compared with hippocampus from non-demented brain. We therefore speculate that this molecule is a marker of the disease process.

Gallant,M.; Rak, M.; Szeghalmi, A.; Del Bigio, M.; Westaway, D.; Yang, J.; Julian, R.; Gough, K.

2006-01-01

222

Dimeric structure of transmembrane domain of amyloid precursor protein in micellar environment.  

Science.gov (United States)

Some pathogenic mutations associated with Alzheimer's disease are thought to affect structural-dynamic properties and the lateral dimerization of amyloid precursor protein (APP) in neuron membrane. Dimeric structure of APP transmembrane fragment Gln(686)-Lys(726) was determined in membrane-mimicking dodecylphosphocholine micelles using high-resolution NMR spectroscopy. The APP membrane-spanning ?-helix Lys(699)-Lys(724) self-associates in a left-handed parallel dimer through extended heptad repeat motif I(702)X(3)M(706)X(2)G(709)X(3)A(713)X(2)I(716)X(3)I(720)X(2)I(723), whereas the juxtamembrane region Gln(686)-Val(695) constitutes the nascent helix, also sensing the dimerization. The dimerization mechanism of APP transmembrane domain has been described at atomic resolution for the first time and is important for understanding molecular events of APP sequential proteolytical cleavage resulting in amyloid-? peptide. PMID:22584060

Nadezhdin, Kirill D; Bocharova, Olga V; Bocharov, Eduard V; Arseniev, Alexander S

2012-06-12

223

Investigation of amyloid deposition in uterine leiomyoma patients  

Directory of Open Access Journals (Sweden)

Full Text Available Objects: To investigate the pathogenesis of amyloid presented in uterine leiomyoma. Methods: 36 uterine leiomyoma patients were recruited and divided into two groups according to Congo red staining results. 6 cases are Congo red staining-positive, and 30 cases Congo red staining-negative which represented amyloid positive and amyloid negative respectively. All patients’ serum total protein (TP, albumin (Alb and prealbumin (PA levels were measured as well as blood hemoglobin (Hb, cell counts of white blood cell (WBC, neutrophils (NEU and lymphocyte (LYM. Glycogen in tissue was compared between amyloid accumulated and amyloid negative sections with periodic acid schiff staining (PAS in leiomyoma patients. Results: All of blood Hb concentration, WBC, NEU and LYM have not been found significant differences between two groups. Also no obvious infiltration of inflammatory cells was observed in tissue with amyloid deposition in uterine leiomyoma patients. And levels of TP, Alb and prealbumin have not been found significant differences between two groups. The amyloid was negative in leiomyoma entity cells range by Congo red staining, while small blood vessels in myoma tissues were positively detected with high rate. Amyloid was found in normal tissue around myoma as well as in blood vessel of pseudo-capsule. Increased PAS-positive material induced by leiomyoma was not correlated with amyloid deposition. Conclusions: Metabolic changes in the setting of functional alterations of cell in local microenvironment with uterine leiomyoma, may be related to the amyloid deposition.

Jinping Liu

2012-08-01

224

Soluble amyloid precursor protein (APP) regulates transthyretin and Klotho gene expression without rescuing the essential function of APP  

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Amyloidogenic processing of the amyloid precursor protein (APP) generates a large secreted ectodomain fragment (APPs?), ?-amyloid (A?) peptides, and an APP intracellular domain (AICD). Whereas A? is viewed as critical for Alzheimer's disease pathogenesis, the role of other APP processing products remains enigmatic. Of interest, the AICD has been implicated in transcriptional regulation, and N-terminal cleavage of APPs? has been suggested to produce an active fragment that may mediate axo...

Li, Hongmei; Wang, Baiping; Wang, Zilai; Guo, Qinxi; Tabuchi, Katsuhiko; Hammer, Robert E.; Su?dhof, Thomas C.; Zheng, Hui

2010-01-01

225

An iron-export ferroxidase activity of ?-amyloid protein precursor is inhibited by zinc in Alzheimer’s Disease  

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Alzheimer’s Disease (AD) is complicated by pro-oxidant intraneuronal Fe2+ elevation as well as extracellular Zn2+ accumulation within amyloid plaque. We found that the AD ?-amyloid protein precursor (APP) possesses ferroxidase activity mediated by a conserved H-ferritin-like active site, which is inhibited specifically by Zn2+. Like ceruloplasmin, APP catalytically oxidizes Fe2+, loads Fe3+ into transferrin, and has a major interaction with ferroportin in HEK293T cells (that lack cerulopla...

Duce, James A.; Tsatsanis, Andrew; Cater, Michael A.; James, Simon A.; Robb, Elysia; Wikhe, Krutika; Leong, Su Ling; Perez, Keyla; Johanssen, Timothy; Greenough, Mark A.; Cho, Hyun-hee; Galatis, Denise; Moir, Robert D.; Masters, Colin L.; Mclean, Catriona

2010-01-01

226

The intramembrane cleavage site of the amyloid precursor protein depends on the length of its transmembrane domain  

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Proteolytic processing of the amyloid precursor protein by ?-secretase generates C99, which subsequently is cleaved by ?-secretase, yielding the amyloid ? peptide (A?). This ?-cleavage occurs within the transmembrane domain (TMD) of C99 and is similar to the intramembrane cleavage of Notch. However, Notch and C99 differ in their site of intramembrane cleavage. The main ?-cleavage of C99 occurs in the middle of the TMD, whereas the cleavage of Notch occurs close to the C-terminal end of ...

Lichtenthaler, Stefan F.; Beher, Dirk; Grimm, Heike S.; Wang, Rong; Shearman, Mark S.; Masters, Colin L.; Beyreuther, Konrad

2002-01-01

227

Introduction of yeast artificial chromosomes containing mutant human amyloid precursor protein genes into transgenic mice  

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Several hypothetical mechanisms have been proposed for the generation and deposition of the amyloid beta (A{beta}) peptide in Alzheimer`s disease (AD). These include overexpression of the amyloid precursor protein (APP) gene, as suggested by Down Syndrome (DS, trisomy 21), and mutation of APP, as suggested by mutations associated with the presence of disease/amyloid deposition in some cases of familial AD (FAD). Although numerous in vitro studies have lead to certain insights into the molecular basis for amyloid deposition, the mechanisms(s) of amyloidogenesis in vivo remains poorly defined. To examine the effect of FAD mutations on amyloidogenesis in an animal model, we have focused on producing APP YAC transgenic mice containing the human APP gene with FAD mutations. These APP YAC transgenics are being produced by introduction of a 650 kb APP YAC through lipid-mediated transfection of ES cells. This strategy has two principal advantages: the APP genomic sequences contain transcriptional regulatory elements required for proper spatial and temporal expression and contain appropriate splice donor and acceptor sites needed to generate the entire spectrum of alternatively spliced APP transcripts. As a first step, we cloned the genomic regions surrounding APP exons 16 and 17 from an APP YAC sublibrary. Both the Swedish and the 717 mutations were then introduced into exons 16 and 17, respectively, by PCR mutagenesis, and subsequently transferred into the 650 kb APP YAC by a two step gene replacement in yeast. The mutant YACs have been introduced into ES cells, and we have determined that these cells are expressing human mutant APP mRNA and protein. These cells are being used to generate transgenic mice. This paradigm should provide the appropriate test of whether a mutant APP gene is capable of producing AD-like pathology in a mouse model.

Call, L.M.; Lamb, B.T.; Boese, K.F. [Johns Hopkins Univ. School of Medicine, Baltimore, MD (United States)] [and others

1994-09-01

228

Protein-induced Photophysical Changes to the Amyloid Indicator Dye Thioflavin T  

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The small molecule thioflavin T (ThT) is a defining probe for the identification and mechanistic study of amyloid fiber formation. As such, ThT is fundamental to investigations of serious diseases such as Alzheimer's disease, Parkinson disease, and type II diabetes. For each disease, a different protein undergoes conformational conversion to a {beta}-sheet rich fiber. The fluorescence of ThT exhibits an increase in quantum yield upon binding these fibers. Despite its widespread use, the structural basis for binding specificity and for the changes to the photophysical properties of ThT remain poorly understood. Here, we report the co-crystal structures of ThT with two alternative states of {beta}-2 microglobulin ({beta}2m); one monomeric, the other an amyloid-like oligomer. In the latter, the dye intercalates between {beta}-sheets orthogonal to the {beta}-strands. Importantly, the fluorophore is bound in such a manner that a photophysically relevant torsion is limited to a range of angles generally associated with low, not high, quantum yield. Quantum mechanical assessment of the fluorophore shows the electronic distribution to be strongly stabilized by aromatic interactions with the protein. Monomeric {beta}2m gives little increase in ThT fluorescence despite showing three fluorophores, at two binding sites, in configurations generally associated with high quantum yield. Our efforts fundamentally extend existing understanding about the origins of amyloid-induced photophysical changes. Specifically, the {beta}-sheet interface that characterizes amyloid acts both sterically and electronically to stabilize the fluorophore's ground state electronic distribution. By preventing the fluorophore from adopting its preferred excited state configuration, nonradiative relaxation pathways are minimized and quantum yield is increased.

L Wolfe; M Calabrese; A Nath; D Blaho; A Miranker; Y Xiong

2011-12-31

229

The predominant form of the amyloid beta-protein precursor in human brain is protease nexin 2.  

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The amyloid beta protein and the amyloid beta-protein precursor (APP) are major constituents of senile plaques and cerebrovascular deposits in patients with Alzheimer disease and Down syndrome. Most human tissues contain mRNA that encodes forms of APP that contain the Kunitz protease inhibitor (KPI+) domain. A major 120-kDa protein corresponding to this KPI+ mRNA is also found in these tissues. This protein is identical to the protease inhibitor protease nexin 2. Brain contains an additional ...

Nostrand, W. E.; Farrow, J. S.; Wagner, S. L.; Bhasin, R.; Goldgaber, D.; Cotman, C. W.; Cunningham, D. D.

1991-01-01

230

Impact of the Nature and Size of the Polymeric Backbone on the Ability of Heterobifunctional Ligands to Mediate Shiga Toxin and Serum Amyloid P Component Ternary Complex Formation  

Directory of Open Access Journals (Sweden)

Full Text Available Inhibition of AB5-type bacterial toxins can be achieved by heterobifunctional ligands (BAITs that mediate assembly of supramolecular complexes involving the toxin’s pentameric cell membrane-binding subunit and an endogenous protein, serum amyloid P component, of the innate immune system. Effective in vivo protection from Shiga toxin Type 1 (Stx1 is achieved by polymer-bound, heterobifunctional inhibitors-adaptors (PolyBAITs, which exhibit prolonged half-life in circulation and by mediating formation of face-to-face SAP-AB5 complexes, block receptor recognition sites and redirect toxins to the spleen and liver for degradation. Direct correlation between solid-phase activity and protective dose of PolyBAITs both in the cytotoxicity assay and in vivo indicate that the mechanism of protection from intoxication is inhibition of toxin binding to the host cell membrane. The polymeric scaffold influences the activity not only by clustering active binding fragments but also by sterically interfering with the supramolecular complex assembly. Thus, inhibitors based on N-(2-hydroxypropyl methacrylamide (HPMA show significantly lower activity than polyacrylamide-based analogs. The detrimental steric effect can partially be alleviated by extending the length of the spacer, which separates pendant ligand from the backbone, as well as extending the spacer, which spans the distance between binding moieties within each heterobifunctional ligand. Herein we report that polymer size and payload of the active ligand had moderate effects on the inhibitor’s activity.

Glen D. Armstrong

2011-08-01

231

AMYPdb: A database dedicated to amyloid precursor proteins  

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Abstract Background Misfolding and aggregation of proteins into ordered fibrillar structures is associated with a number of severe pathologies, including Alzheimer's disease, prion diseases, and type II diabetes. The rapid accumulation of knowledge about the sequences and structures of these proteins allows using of in silico methods to investigate the molecular mechanisms of their abnormal conformational changes and assembly. However, such an approach requires the c...

2008-01-01

232

Elemental analysis of human serum and serum protein fractions by thermal neutron activation  

International Nuclear Information System (INIS)

Some applications of thermal neutron activation for the determination of elemental contents in human serum and human serum protein fractions are presented. Firstly total serum is dealt with, secondly serum protein fractions obtained by gel filtration are described. A brief review on the role of (trace) elements in human health and disease and a compilation of literature data for elemental contents in human serum, as obtained by neutron activation techniques, are given. The most important sources of statistical and systematic errors are evaluated. Results for the contents of sodium, potassium, magnesium, bromine, iron, copper, zinc, selenium, rubidium, cesium and antimony in serum are given, with emphasis on control of accuracy and precision. The possible relation between selenium in blood and cancer occurrence in humans is discussed. The results of elemental analyses from cancer patients and from a patient receiving a cytostatic treatment are presented. A survey of literature results for the determination of protein-bound elemental contents in serum is presented. Subsequently, results from a study on the behaviour of elements during gel filtration are discussed. Gel-element and protein-element interactions are studied. Finally the protein-bound occurrence of trace elements in human serum is determined by gel filtration and neutron activation analysis. Results for both desalting and fractionation are given, for the elements bromine, copper, manganese, vanadium, selenium, zinc, rubidium, iron and iodine. (Auth.)

233

Distribution of precursor amyloid-?-protein messenger RNA in human cerebral cortex: relationship to neurofibrillary tangles and neuritic plaques  

International Nuclear Information System (INIS)

Neurofibrillary tangles (NFT) and neuritic plaques (NP), two neuropathological markers of Alzheimer disease, may both contain peptide fragments derived from the human amyloid ? protein. However, the nature of the relationship between NFT and NP and the source of the amyloid ? proteins found in each have remained unclear. The authors used in situ hybridization techniques to map the anatomical distribution of precursor amyloid-?-protein mRNA in the neocortex of brains from three subjects with no known neurologic disease and from five patients with Alzheimer disease. In brains from control subjects, positively hybridizing neurons were present in cortical regions and layers that contain a high density of neuropathological markers in Alzheimer disease, as well as in those loci that contain NP but few NFT. Quantitative analyses of in situ hybridization patterns within layers III and V of the superior frontal cortex revealed that the presence of high numbers of NFT in Alzheimer-diseased brains was associated with a decrease in the number of positively hybridizing neurons compared to controls and Alzheimer-diseased brains with few NFT. These findings suggest that the expression of precursor amyloid-?-protein mRNA may be a necessary but is clearly not a sufficient prerequisite for NFT formation. In addition, these results may indicate that the amyloid ? protein, present in NP in a given region or layer of cortex, is not derived from the resident neuronal cell bodies that express the mRNA for the precursor protein

234

Bacterial curli protein promotes the conversion of PAP248-286 into the amyloid SEVI: cross-seeding of dissimilar amyloid sequences  

Directory of Open Access Journals (Sweden)

Full Text Available Fragments of prostatic acid phosphatase (PAP248-286 in human semen dramatically increase HIV infection efficiency by increasing virus adhesion to target cells. PAP248-286 only enhances HIV infection in the form of amyloid aggregates termed SEVI (Semen Enhancer of Viral Infection, however monomeric PAP248-286 aggregates very slowly in isolation. It has therefore been suggested that SEVI fiber formation in vivo may be promoted by exogenous factors. We show here that a bacterially-produced extracellular amyloid (curli or Csg acts as a catalytic agent for SEVI formation from PAP248-286 at low concentrations in vitro, producing fibers that retain the ability to enhance HIV (Human Immunodeficiency Virus infection. Kinetic analysis of the cross-seeding effect shows an unusual pattern. Cross-seeding PAP248-286 with curli only moderately affects the nucleation rate while significantly enhancing the growth of fibers from existing nuclei. This pattern is in contrast to most previous observations of cross-seeding, which show cross-seeding partially bypasses the nucleation step but has little effect on fiber elongation. Seeding other amyloidogenic proteins (IAPP (islet amyloid polypeptide and A?1?40 with curli showed varied results. Curli cross-seeding decreased the lag-time of IAPP amyloid formation but strongly inhibited IAPP elongation. Curli cross-seeding exerted a complicated concentration dependent effect on A?1?40 fibrillogenesis kinetics. Combined, these results suggest that the interaction of amyloidogenic proteins with preformed fibers of a different type can take a variety of forms and is not limited to epitaxial nucleation between proteins of similar sequence. The ability of curli fibers to interact with proteins of dissimilar sequences suggests cross-seeding may be a more general phenomenon than previously supposed.

Kevin Hartman

2013-02-01

235

What the evolution of the amyloid protein precursor supergene family tells us about its function.  

Science.gov (United States)

The Alzheimer's disease amyloid protein precursor (APP) gene is part of a multi-gene super-family from which sixteen homologous amyloid precursor-like proteins (APLP) and APP species homologues have been isolated and characterised. Comparison of exon structure (including the uncharacterised APL-1 gene), construction of phylogenetic trees, and analysis of the protein sequence alignment of known homologues of the APP super-family were performed to reconstruct the evolution of the family and to assess the functional significance of conserved protein sequences between homologues. This analysis supports an adhesion function for all members of the APP super family, with specificity determined by those sequences which are not conserved between APLP lineages, and provides evidence for an increasingly complex APP superfamily during evolution. The analysis also suggests that Drosophila APPL and Caenorhabditis elegans APL-1 may be a fourth APLP lineage indicating that these proteins, while not functional homologues of human APP, are similarly likely to regulate cell adhesion. Furthermore, the betaA4 sequence is highly conserved only in APP orthologues, strongly suggesting this sequence is of significant functional importance in this lineage. PMID:10676850

Coulson, E J; Paliga, K; Beyreuther, K; Masters, C L

2000-03-01

236

Role of sequence and membrane composition in structure of transmembrane domain of Amyloid Precursor Protein  

Science.gov (United States)

Aggregation of proteins of known sequence is linked to a variety of neurodegenerative disorders. The amyloid ? (A?) protein associated with Alzheimer's Disease (AD) is derived from cleavage of the 99 amino acid C-terminal fragment of Amyloid Precursor Protein (APP-C99) by ?-secretase. Certain familial mutations of APP-C99 have been shown to lead to altered production of A? protein and the early onset of AD. We describe simulation studies exploring the structure of APP-C99 in micelle and membrane environments. Our studies explore how changes in sequence and membrane composition influence (1) the structure of monomeric APP-C99 and (2) APP-C99 homodimer structure and stability. Comparison of simulation results with recent NMR studies of APP-C99 monomers and dimers in micelle and bicelle environments provide insight into how critical aspects of APP-C99 structure and dimerization correlate with secretase processing, an essential component of the A? protein aggregation pathway and AD.

Straub, John

2013-03-01

237

Functional Analysis of the Accessory Protein TapA in Bacillus subtilis Amyloid Fiber Assembly  

Science.gov (United States)

Bacillus subtilis biofilm formation relies on the assembly of a fibrous scaffold formed by the protein TasA. TasA polymerizes into highly stable fibers with biochemical and morphological features of functional amyloids. Previously, we showed that assembly of TasA fibers requires the auxiliary protein TapA. In this study, we investigated the roles of TapA sequences from the C-terminal and N-terminal ends and TapA cysteine residues in its ability to promote the assembly of TasA amyloid-like fibers. We found that the cysteine residues are not essential for the formation of TasA fibers, as their replacement by alanine residues resulted in only minor defects in biofilm formation. Mutating sequences in the C-terminal half had no effect on biofilm formation. However, we identified a sequence of 8 amino acids in the N terminus that is key for TasA fiber formation. Strains expressing TapA lacking these 8 residues were completely defective in biofilm formation. In addition, this TapA mutant protein exhibited a dominant negative effect on TasA fiber formation. Even in the presence of wild-type TapA, the mutant protein inhibited fiber assembly in vitro and delayed biofilm formation in vivo. We propose that this 8-residue sequence is crucial for the formation of amyloid-like fibers on the cell surface, perhaps by mediating the interaction between TapA or TapA and TasA molecules. PMID:24488317

Romero, Diego; Vlamakis, Hera; Losick, Richard

2014-01-01

238

Conformational stability of mammalian prion protein amyloid fibrils is dictated by a packing polymorphism within the core region.  

Science.gov (United States)

Mammalian prion strains are believed to arise from the propagation of distinct conformations of the misfolded prion protein PrP(Sc). One key operational parameter used to define differences between strains has been conformational stability of PrP(Sc) as defined by resistance to thermal and/or chemical denaturation. However, the structural basis of these stability differences is unknown. To bridge this gap, we have generated two strains of recombinant human prion protein amyloid fibrils that show dramatic differences in conformational stability and have characterized them by a number of biophysical methods. Backbone amide hydrogen/deuterium exchange experiments revealed that, in sharp contrast to previously studied strains of infectious amyloid formed from the yeast prion protein Sup35, differences in ?-sheet core size do not underlie differences in conformational stability between strains of mammalian prion protein amyloid. Instead, these stability differences appear to be dictated by distinct packing arrangements (i.e. steric zipper interfaces) within the amyloid core, as indicated by distinct x-ray fiber diffraction patterns and large strain-dependent differences in hydrogen/deuterium exchange kinetics for histidine side chains within the core region. Although this study was limited to synthetic prion protein amyloid fibrils, a similar structural basis for strain-dependent conformational stability may apply to brain-derived PrP(Sc), especially because large strain-specific differences in PrP(Sc) stability are often observed despite a similar size of the PrP(Sc) core region. PMID:24338015

Cobb, Nathan J; Apostol, Marcin I; Chen, Shugui; Smirnovas, Vytautas; Surewicz, Witold K

2014-01-31

239

Signaling via amyloid precursor-like proteins APLP1 and APLP2.  

Science.gov (United States)

The amyloid-? protein precursor (A?PP) has been implicated in Alzheimer's disease (AD) not only as a precursor of the amyloid-? peptide but also as a mediator of signal transduction. We recently identified novel mediators of A?PP signaling via interactions with Mint/X11 family proteins Mint1 and Mint3. These mediators include transcriptional co-activators Taz and Yap. Here we show that Taz and Yap also mediate signaling via the A?PP paralogues APLP1 and APLP2 through interactions with Mint1 and Mint3. APLP1 and APLP2 formed transcriptionally active triple protein complexes with the adaptor protein Mint3 and each of the transcriptional regulators Taz and Yap, and complex formation was regulated by the ?-secretase cleavage of APLP1 and APLP2. The presence of Mint1 instead of Mint3 in the complex prevented its translocation to the nucleus. APLP1 displayed much lower transactivation levels compared to A?PP and APLP2. These results indicate that all three A?PP family members are capable of activating gene transcription via Mint3-Taz and Mint3-Yap. PMID:21178287

Orcholski, Mark E; Zhang, Qiang; Bredesen, Dale E

2011-01-01

240

Photochemistry of modified proteins benzophenone-containing bovine serum albumin  

International Nuclear Information System (INIS)

The results of exploratory and mechanistic studies of the photochemistry of poly-p-benzoyl-acetimido-bovine serum albumin, a modified protein containing photoreactive and photosensitizing groups, are reported. Specifically described are recent findings concerning (1) the synthesis and characterization of a modified bovine serum albumin that contains benzophenone-like moieties, (2) the photochemistry of this modified protein which appeared to involve photoreductive coupling of the benzophenone chromophores to the protein backbone, and (3) triplet energy transfer from modified bovine serum albumin to small molecule acceptors resulting in quenching of the photoreaction. (author)

 
 
 
 
241

Serum amyloid A stimulates macrophage foam cell formation via lectin-like oxidized low-density lipoprotein receptor 1 upregulation  

International Nuclear Information System (INIS)

Highlights: ? SAA induced macrophage foam cell formation. ? SAA stimulated upregulation of lectin-like oxidized low-density lipoprotein receptor 1 (LOX1). ? SAA-induced LOX1 expression and foam cell formation is mediated by JNK/NF-?B signaling. ? HDL-conjugated SAA also stimulates foam cell formation via LOX1 upregulation. ? The finding reveals a novel mechanism of action of SAA in the pathogenesis of atherosclerosis. -- Abstract: Elevated levels of serum amyloid A (SAA) is a risk factor for cardiovascular diseases, however, the role of SAA in the pathophysiology of atherosclerosis remains unclear. Here we show that SAA induced macrophage foam cell formation. SAA-stimulated foam cell formation was mediated by c-jun N-terminal kinase (JNK) signaling. Moreover, both SAA and SAA-conjugated high density lipoprotein stimulated the expression of the important scavenger receptor lectin-like oxidized low-density lipoprotein receptor 1 (LOX1) via nuclear factor-?B (NF-?B). A LOX1 antagonist carrageenan significantly blocked SAA-induced foam cell formation, indicating that SAA promotes foam cell formation via LOX1 expression. Our findings therefore suggest that SAA stimulates foam cell formation via LOX1 induction, and thus likely contributes to atherogenesis

242

Serum Amyloid A, Procalcitonin, Tumor Necrosis Factor-?, and Interleukin-1? Levels in Neonatal Late-Onset Sepsis  

Science.gov (United States)

Background. Sepsis is an important cause of mortality in newborns. However, a single reliable marker is not available for the diagnosis of neonatal late-onset sepsis (NLS). The aim of this study is to evaluate the value of serum amyloid A (SAA) and procalcitonin (PCT) in the diagnosis and follow-up of NLS. Methods. 36 septic and healthy newborns were included in the study. However, SAA, PCT, TNF-?, IL-1?, and CRP were serially measured on days 0, 4, and 8 in the patients and once in the controls. Töllner's sepsis score (TSS) was calculated for each patient. Results. CRP, PCT, and TNF-? levels in septic neonates at each study day were significantly higher than in the controls (P = .001). SAA and IL-1? levels did not differ from healthy neonates. The sensitivity and specificity were 86.8% and 97.2% for PCT, 83.3% and 80.6% for TNF-?, 75% and 44.4% for SAA on day 0. Conclusion. Present study suggests that CRP seems to be the most helpful indicator and PCT and TNF-? may be useful markers for the early diagnosis of NLS. However, SAA, IL-1?, and TSS are not reliable markers for the diagnosis and follow-up of NLS. PMID:19043563

Ucar, Birsen; Yildiz, Bilal; Aksit, M. Arif; Yarar, Coskun; Colak, Omer; Akbay, Yildiz; Colak, Ertugrul

2008-01-01

243

Identification of a mouse brain cDNA that encodes a protein related to the Alzheimer disease-associated amyloid beta protein precursor.  

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We have isolated a cDNA from a mouse brain library that encodes a protein whose predicted amino acid sequence is 42% identical and 64% similar to that of the amyloid beta protein precursor (APP). This 653-amino acid protein, which we have termed the amyloid precursor-like protein (APLP), appears to be similar to APP in overall structure as well as amino acid sequence. The amino acid homologies are concentrated within three distinct regions of the two proteins where the identities are 47%, 54%...

Wasco, W.; Bupp, K.; Magendantz, M.; Gusella, J. F.; Tanzi, R. E.; Solomon, F.

1992-01-01

244

Non-Specificity of Binding of ?-Secretase Modulators to the Amyloid Precursor Protein  

Science.gov (United States)

Evidence has been presented (Kukar et al. 2008 Nature 453, 925–929) that certain ?–secretase modulators (GSMs) target the 99 residue C-terminal domain (C99) of the amyloid precursor protein, a substrate of ?-secretase, but not the protease complex itself. Here, NMR results demonstrate a lack of specific binding of these GSMs to monodisperse C99 in LMPG micelles. In addition, results indicate that C99 was likely to have been aggregated in some of the key experiments of the previous work, and that binding of GSMs to these C99 aggregates is also of a non-specific nature. PMID:19928774

Beel, Andrew J.; Barrett, Paul; Schnier, Paul D.; Hitchcock, Stephen A.; Bagal, Dhanashri; Sanders, Charles R.; Jordan, John B.

2009-01-01

245

Nonspecificity of binding of gamma-secretase modulators to the amyloid precursor protein.  

Science.gov (United States)

Evidence that certain gamma-secretase modulators (GSMs) target the 99-residue C-terminal domain (C99) of the amyloid precursor protein, a substrate of gamma-secretase, but not the protease complex itself has been presented [Kukar, T. L., et al. (2008) Nature 453, 925-929]. Here, NMR results demonstrate a lack of specific binding of these GSMs to monodisperse C99 in LMPG micelles. In addition, results indicate that C99 was likely to have been aggregated in some of the key experiments of the previous work and that binding of GSMs to these C99 aggregates is also of a nonspecific nature. PMID:19928774

Beel, Andrew J; Barrett, Paul; Schnier, Paul D; Hitchcock, Stephen A; Bagal, Dhanashri; Sanders, Charles R; Jordan, John B

2009-12-22

246

The Clathrin Assembly Protein AP180 Regulates the Generation of Amyloid-? Peptide  

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The overproduction and extracellular buildup of amyloid-? peptide (A?) is a critical step in the etiology of Alzheimer’s disease. Recent data suggest that intracellular trafficking is of central importance in the production of A?. Here we use a neuronal cell line to examine two structurally similar clathrin assembly proteins, AP180 and CALM. We show that RNA interference-mediated knockdown of AP180 reduces the generation of A? 1-40 and A? 1-42, whereas CALM knockdown has no effect on A...

Wu, Fangbai; Matsuoka, Yasuji; Mattson, Mark P.; Yao, Pamela J.

2009-01-01

247

Cellular prion protein participates in amyloid-? transcytosis across the blood–brain barrier  

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The blood–brain barrier (BBB) facilitates amyloid-? (A?) exchange between the blood and the brain. Here, we found that the cellular prion protein (PrPc), a putative receptor implicated in mediating A? neurotoxicity in Alzheimer's disease (AD), participates in A? transcytosis across the BBB. Using an in vitro BBB model, [125I]-A?1?40 transcytosis was reduced by genetic knockout of PrPc or after addition of a competing PrPc-specific antibody. Furthermore, we provide evidence that PrPc ...

Pflanzner, Thorsten; Petsch, Benjamin; Andre?-dohmen, Bettina; Mu?ller-schiffmann, Andreas; Tschickardt, Sabrina; Weggen, Sascha; Stitz, Lothar; Korth, Carsten; Pietrzik, Claus U.

2012-01-01

248

Cellular prion protein is essential for oligomeric amyloid-?-induced neuronal cell death  

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In Alzheimer disease (AD), amyloid-? (A?) oligomer is suggested to play a critical role in imitating neurodegeneration, although its pathogenic mechanism remains to be determined. Recently, the cellular prion protein (PrPC) has been reported to be an essential co-factor in mediating the neurotoxic effect of A? oligomer. However, these previous studies focused on the synaptic plasticity in either the presence or the absence of PrPC and no study to date has reported whether PrPC is required ...

Kudo, Wataru; Lee, Hyun-pil; Zou, Wen-quan; Wang, Xinglong; Perry, George; Zhu, Xiongwei; Smith, Mark A.; Petersen, Robert B.; Lee, Hyoung-gon

2012-01-01

249

Alzheimer Amyloid-? Oligomer Bound to Post-Synaptic Prion Protein Activates Fyn to Impair Neurons  

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Amyloid-beta (A?) oligomers are thought to trigger Alzheimer’s disease (AD) pathophysiology. Cellular Prion Protein (PrPC) selectively binds oligomeric A? and can mediate AD-related phenotypes. Here, we examined the specificity, distribution and signaling from A?/PrP complexes, seeking to explain how they might alter the function of NMDA receptors in neurons. PrPC is enriched in post-synaptic densities, and A?/PrPC interaction leads to Fyn kinase activation. Soluble A? assemblies deriv...

Um, Ji Won; Nygaard, Haakon B.; Heiss, Jacqueline K.; Kostylev, Mikhail A.; Stagi, Massimiliano; Vortmeyer, Alexander; Wisniewski, Thomas; Gunther, Erik C.; Strittmatter, Stephen M.

2012-01-01

250

Specificity of Amyloid Precursor-like Protein 2 Interactions with MHC Class I Molecules  

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The ubiquitously expressed amyloid precursor-like protein 2 (APLP2) has been previously found to regulate cell surface expression of the MHC class I molecule Kd and bind strongly to Kd. In the study reported here, we demonstrated that APLP2 binds, in varied degrees, to several other mouse MHC class I allotypes, and that the ability of APLP2 to affect cell surface expression of an MHC class I molecule is not limited to Kd. Ld, like Kd, was found associated with APLP2 in the Golgi, but Kd was a...

Tuli, Amit; Sharma, Mahak; Naslavsky, Naava; Caplan, Steve; Solheim, Joyce C.

2008-01-01

251

Amyloid Precursor Protein Regulates Brain Apolipoprotein E and Cholesterol Metabolism through Lipoprotein Receptor LRP1  

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Mutations in the amyloid precursor protein (APP) cause early-onset Alzheimer's disease (AD), but the only genetic risk factor for late-onset AD is the ?4 allele of apolipoprotein E (apoE), a major cholesterol carrier. Using Cre-lox conditional knockout mice, we demonstrate that lipoprotein receptor LRP1 expression regulates apoE and cholesterol levels within the CNS. We also found that deletion of APP and its homologue APLP2, or components of the ?-secretase complex, significantly enhanced ...

Liu, Qiang; Zerbinatti, Celina V.; Zhang, Juan; Hoe, Hyang-sook; Wang, Baiping; Cole, Sarah L.; Herz, Joachim; Muglia, Louis; Bu, Guojun

2007-01-01

252

Development of dual targeting inhibitors against aggregations of amyloid-? and tau protein.  

Science.gov (United States)

Aggregations of both amyloid-? (A?) and hyper-phosphorylated tau proteins are recognized as key pathological manifestations of Alzheimer's disease (AD). Agents that inhibit both those forms of aggregation show promise as drug candidates. Seventeen oligo heteroaromatic compounds were rapidly synthesized via a one-pot, 3- or 4-component coupling procedure. Evaluations showed that compounds E16 and E18 were the most potent inhibitors of A? and tau aggregations (E16: IC50s = 0.38, 0.29 ?M against A?, tau, respectively, E18: IC50s = 0.55, 0.30 ?M against A?, tau, respectively). PMID:25086914

Fuse, Shinichiro; Matsumura, Keisuke; Fujita, Yuki; Sugimoto, Hachiro; Takahashi, Takashi

2014-10-01

253

Structural heterogeneity in transmembrane amyloid precursor protein homodimer is a consequence of environmental selection.  

Science.gov (United States)

The 99 amino acid C-terminal fragment of amyloid precursor protein (C99), consisting of a single transmembrane (TM) helix, is known to form homodimers. Homodimers can be processed by ?-secretase to produce amyloid-? (A?) protein, which is implicated in Alzheimer's disease (AD). While knowledge of the structure of C99 homodimers is of great importance, experimental NMR studies and simulations have produced varying structural models, including right-handed and left-handed coiled-coils. In order to investigate the structure of this critical protein complex, simulations of the C99(15-55) homodimer in POPC membrane bilayer and DPC surfactant micelle environments were performed using a multiscale approach that blends atomistic and coarse-grained models. The C99(15-55) homodimer adopts a dominant right-handed coiled-coil topology consisting of three characteristic structural states in a bilayer, only one of which is dominant in the micelle. Our structural study, which provides a self-consistent framework for understanding a number of experiments, shows that the energy landscape of the C99 homodimer supports a variety of slowly interconverting structural states. The relative importance of any given state can be modulated through environmental selection realized by altering the membrane or micelle characteristics. PMID:24926593

Dominguez, Laura; Foster, Leigh; Meredith, Stephen C; Straub, John E; Thirumalai, D

2014-07-01

254

Tetraspanin12 regulates ADAM10-dependent cleavage of amyloid precursor protein  

Science.gov (United States)

Using mass spectrometry, we identified ADAM10 (a membrane-associated metalloproteinase) as a partner for TSPAN12, a tetraspanin protein. TSPAN12-ADAM10 interaction was confirmed by reciprocal coimmunoprecipitation in multiple tumor cell lines. TSPAN12, to a greater extent than other tetraspanins (CD81, CD151, CD9, and CD82), associated with ADAM10 but not with ADAM17. Overexpression of TSPAN12 enhanced ADAM10-dependent shedding of amyloid precursor protein (APP) in MCF7 (breast cancer) and SH-SY5Y (neuroblastoma) cell lines. Conversely, siRNA ablation of endogenous TSPAN12 markedly diminished APP proteolysis in both cell lines. Furthermore, TSPAN12 overexpression enhanced ADAM10 prodomain maturation, whereas TSPAN12 ablation diminished ADAM10 maturation. A palmitoylation-deficient TSPAN12 mutant failed to associate with ADAM10, inhibited ADAM10-dependent proteolysis of APP, and inhibited ADAM10 maturation, most likely by interfering with endogenous wild-type TSPAN12. In conclusion, TSPAN12 serves as a novel and robust partner for ADAM10 and promotes ADAM10 maturation, thereby facilitating ADAM10-dependent proteolysis of APP. This novel mode of regulating APP cleavage is of relevance to Alzheimer’s disease therapy.—Xu, D., Sharma, C., Hemler, M. E. Tetraspanin12 regulates ADAM10-dependent cleavage of amyloid precursor protein. PMID:19587294

Xu, Daosong; Sharma, Chandan; Hemler, Martin E.

2009-01-01

255

[The precursor of amyloid peptide in Alzheimer disease: a protein with multiple functions].  

Science.gov (United States)

Cellular metabolism of the amyloid precursor protein (APP) has been widely studied, but the function of the protein remains elusive. APP knock out mice do not show any phenotype, due to in vivo compensation by APLP genes, encoding proteins similar to APP. In order to study the neuronal metabolism of APP, human APP has been expressed in rat cortical neurons in culture. Following differentiation in culture, rat cortical neurons are organized into networks of connected cells, which show neuronal activity in the form of spontaneous and synchronous calcium oscillations. Expression of human APP in these neuronal networks inhibits calcium oscillations, while downregulation of endogenous APP expression increases the frequency and decreases the amplitude of oscillations. Therefore, APP controls neuronal calcium homeostasis and excitability. In the same experimental model, APP is also able to control the neuronal synthesis of cholesterol. Finally, the APP carboxy terminal domain is involved in the epigenetic control of gene expression. Modulation of neuronal expression of APP allows to identify several important functions of the precursor of the amyloid peptide found in senile plaques of Alzheimer disease. PMID:20218187

Octave, J N

2009-01-01

256

Endogenous Acute Phase Serum Amyloid A Lacks Pro-Inflammatory Activity, Contrasting the Two Recombinant Variants That Activate Human Neutrophils through Different Receptors.  

Science.gov (United States)

Most notable among the acute phase proteins is serum amyloid A (SAA), levels of which can increase 1000-fold during infections, aseptic inflammation, and/or trauma. Chronically elevated SAA levels are associated with a wide variety of pathological conditions, including obesity and rheumatic diseases. Using a recombinant hybrid of the two human SAA isoforms (SAA1 and 2) that does not exist in vivo, numerous in vitro studies have given rise to the notion that acute phase SAA is a pro-inflammatory molecule with cytokine-like properties. It is however unclear whether endogenous acute phase SAA per se mediates pro-inflammatory effects. We tested this in samples from patients with inflammatory arthritis and in a transgenic mouse model that expresses human SAA1. Endogenous human SAA did not drive production of pro-inflammatory IL-8/KC in either of these settings. Human neutrophils derived from arthritis patients displayed no signs of activation, despite being exposed to severely elevated SAA levels in circulation, and SAA-rich sera also failed to activate cells in vitro. In contrast, two recombinant SAA variants (the hybrid SAA and SAA1) both activated human neutrophils, inducing L-selectin shedding, production of reactive oxygen species, and production of IL-8. The hybrid SAA was approximately 100-fold more potent than recombinant SAA1. Recombinant hybrid SAA and SAA1 activated neutrophils through different receptors, with recombinant SAA1 being a ligand for formyl peptide receptor 2 (FPR2). We conclude that even though recombinant SAAs can be valuable tools for studying neutrophil activation, they do not reflect the nature of the endogenous protein. PMID:23626589

Christenson, Karin; Björkman, Lena; Ahlin, Sofie; Olsson, Maja; Sjöholm, Kajsa; Karlsson, Anna; Bylund, Johan

2013-01-01

257

Differential serum protein markers and the clinical severity of asthma  

Directory of Open Access Journals (Sweden)

Full Text Available Norbert Meyer,1,2 Sarah Janine Nuss,1 Thomas Rothe,1 Alexander Siebenhüner,1 Cezmi A Akdis,2 Günter Menz11Hochgebirgsklinik Davos, Davos-Wolfgang, Switzerland; 2Swiss Institute of Allergy and Asthma Research (SIAF, Davos Platz, SwitzerlandBackground: Asthma is a heterogeneous disease characterized by different clinical phenotypes and the involvement of multiple inflammatory pathways. During airway inflammation, many cytokines and chemokines are released and some are detectable in the sera.Objective: Serum chemokines and cytokines, involved in airway inflammation in asthma patients, were investigated.Methods: A total of 191 asthma patients were classified by hierarchical cluster analysis, including the following parameters: forced expiratory volume in 1 second (FEV1, eosinophil cationic protein (ECP serum levels, blood eosinophils, Junipers asthma symptom score, and the change in FEV1, ECP serum levels, and blood eosinophils after 3 weeks of asthma therapy. Serum proteins were measured by multiplex analysis. Receiver operating characteristic (ROC curves were used to evaluate the validity of serum proteins for discriminating between asthma clusters.Results: Classification of asthma patients identified one cluster with high ECP serum levels, increased blood eosinophils, low FEV1 values, and good FEV1 improvement in response to asthma therapy (n=60 and one cluster with low ECP serum levels, low numbers of blood eosinophils, higher FEV1 values, and no FEV1 improvement in response to asthma therapy (n=131. Serum interleukin (IL-8, eotaxin, vascular endothelial growth factor (VEGF, cutaneous T-cell-attracting chemokine (CTACK, growth-related oncogene (GRO-?, and hepatocyte growth factor (HGF were significantly different between the two clusters of asthma patients. ROC analysis for serum proteins calculated a sensitivity of 55.9% and specificity of 75.8% for discriminating between them.Conclusion: Serum cytokine and chemokine levels might be predictors for the severity of asthmatic inflammation, asthma control, and response to therapy, and therefore might be useful for treatment optimization.Keywords: asthma, cluster, phenotype, serum cytokines

Meyer N

2014-04-01

258

Effect of electroconvulsive therapy on serum myelin basic protein immunoreactivity.  

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A sensitive radioimmunoassay that can detect brain damage in cases of head injury and stroke was applied to blood samples from 13 patients before and after they received multiple treatments with electroconvulsive therapy for psychiatric disorder. None of the patients showed a significant increase in serum myelin basic protein immunoreactivity. As increased serum myelin basic protein immunoreactivity may reflect myelin damage it is apparent that in these patients electroconvulsive therapy did ...

Hoyle, N. R.; Pratt, R. T.; Thomas, D. G.

1984-01-01

259

Lysine 624 of the amyloid precursor protein (APP) is a critical determinant of amyloid ? peptide length: support for a sequential model of ?-secretase intramembrane proteolysis and regulation by the amyloid ? precursor protein (APP) juxtamembrane region.  

Science.gov (United States)

?-Secretase is a multiprotein intramembrane cleaving aspartyl protease (I-CLiP) that catalyzes the final cleavage of the amyloid ? precursor protein (APP) to release the amyloid ? peptide (A?). A? is the primary component of senile plaques in Alzheimer's disease (AD), and its mechanism of production has been studied intensely. ?-Secretase executes multiple cleavages within the transmembrane domain of APP, with cleavages producing A? and the APP intracellular domain (AICD), referred to as ? and ?, respectively. The heterogeneous nature of the ? cleavage that produces various A? peptides is highly relevant to AD, as increased production of A? 1-42 is genetically and biochemically linked to the development of AD. We have identified an amino acid in the juxtamembrane region of APP, lysine 624, on the basis of APP695 numbering (position 28 relative to A?) that plays a critical role in determining the final length of A? peptides released by ?-secretase. Mutation of this lysine to alanine (K28A) shifts the primary site of ?-secretase cleavage from 1-40 to 1-33 without significant changes to ? cleavage. These results further support a model where ? cleavage occurs first, followed by sequential proteolysis of the remaining transmembrane fragment, but extend these observations by demonstrating that charged residues at the luminal boundary of the APP transmembrane domain limit processivity of ?-secretase. PMID:21868378

Kukar, Thomas L; Ladd, Thomas B; Robertson, Paul; Pintchovski, Sean A; Moore, Brenda; Bann, Maralyssa A; Ren, Zhao; Jansen-West, Karen; Malphrus, Kim; Eggert, Simone; Maruyama, Hiroko; Cottrell, Barbara A; Das, Pritam; Basi, Guriqbal S; Koo, Edward H; Golde, Todd E

2011-11-18

260

On the subject of rigor in the study of amyloid ?-protein assembly.  

Science.gov (United States)

According to Thomas Kuhn, the success of 'normal science,' the science we all practice on a daily basis, depends on the adherence to, and practice of, a paradigm accepted by the scientific community. When great scientific upheavals occur, they involve the rejection of the current paradigm in favor of a new paradigm that better integrates the facts available and better predicts the behavior of a particular scientific system. In the field of Alzheimer's disease, a recent example of such a paradigm shift has been the apparent rejection of the 'amyloid cascade hypothesis,' promulgated by Hardy and Higgins in 1992 to explain the etiology of Alzheimer's disease, in favor of what has been referred to as the 'oligomer cascade hypothesis'. This paradigm shift has been breathtaking in its rapidity, its pervasiveness in the Alzheimer's disease field, and its adoption in an increasing number of other fields, including those of Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and the prionoses. However, these facts do not mean, a priori, that the experiments extant, and any re-interpretation of them, should be accepted by rote as support for the new paradigm. In the discussion that follows, I consider the foundational studies leading to the oligomer cascade hypothesis and evaluate the current state of the paradigm. I argue here that, more often than not, insufficient rigor has been applied in studies upon which this new paradigm has been based. Confusion, rather than clarity, has resulted. If the field is to make progress forward using as its paradigmatic basis amyloid ?-protein oligomerization, then an epistemological re-evaluation of the amyloid ?-protein oligomer system is required. PMID:23981712

Teplow, David B

2013-01-01

 
 
 
 
261

A mutation protective against Alzheimer's disease renders amyloid ? precursor protein incapable of mediating neurotoxicity.  

Science.gov (United States)

Expression of a familial Alzheimer's disease (AD)-linked mutant of amyloid ? precursor protein (APP) or the binding of transforming growth factor ?2 to wild-type (wt)-APP causes neuronal death by activating an intracellular death signal (a APP-mediated intracellular death signal) in the absence of the involvement of amyloid ? (A?) toxicity in vitro. These neuronal death models may therefore be regarded as A?-independent neuronal death models related to AD. A recent study has shown that the A673T mutation in the APP isoform APP770 , corresponding to the A598T mutation in the most prevalent neuronal APP isoform APP695 (an AD-protective mutant of APP), is linked to a reduction in the incidence rate of AD. Consistent with this, cells expressing the AD-protective mutant of APP produce less A? than cells expressing wt-APP. In this study, transforming growth factor ?2 caused death in cultured neuronal cells expressing wt-APP, but not in those expressing the AD-protective mutant of APP. This result suggests that the AD-protective mutation of APP reduces the incidence rate of AD by attenuating the APP-mediated intracellular death signal. In addition, a mutation that causes hereditary cerebral hemorrhage with amyloidosis-Dutch type also attenuated the APP-mediated intracellular death signal. The A598T mutation of amyloid precursor protein APP is linked to a reduction in the incidence rate of Alzheimer's disease (AD). This study shows that TGF?2 causes death in neuronal cells expressing wild-type APP, but not in those expressing the AD-protective mutant of APP, suggesting that the AD-protective mutation of APP reduces the incidence rate of AD by attenuating the APP-mediated intracellular death signal. PMID:24646423

Hashimoto, Yuichi; Matsuoka, Masaaki

2014-07-01

262

Deletion of the amyloid precursor-like protein 2 (APLP2) does not affect hippocampal neuron morphology or function  

Science.gov (United States)

Amyloid precursor protein (APP), the parent molecule to amyloid ? peptide, is part of larger gene family with two mammalian homologues, amyloid precursor-like protein 1 (APLP1) and amyloid precursor-like protein 2 (APLP2). Initial knock-out studies demonstrated that while single APP family gene deletions produced relatively mild phenotypes, deficiency of APLP2 and one other member of the gene family resulted in perinatal lethality, suggesting vital roles masked by functional redundancy of the other homologues. Because of the importance of APP in Alzheimer’s disease, the vast majority of studies to date have concentrated on the neuronal functions of APP, leaving limited data on its homologues. APLP2 is of particular interest as it contains high sequence homology with APP, is processed similarly, is expressed in overlapping spatial and temporal patterns, and is obligatory for lethality when combined with deficiency of either APLP1 or APP but does not contain the toxic amyloid ? sequence. Here we sought to test the role of APLP2 on neuronal structure and function using a combined approach involving in vitro and in vivo techniques in young and aged animals. Surprisingly, we found that unlike APP, APLP2 appears not to be essential for maintenance of dendritic structure, spiny density, or synaptic function. Thus, there is clear divergence in the functional redundancy between APP and APLP2. PMID:22353605

Midthune, Brea; Tyan, Sheue-Houy; Walsh, Jessica J.; Sarsoza, Floyd; Eggert, Simone; Hof, Patrick R.; Dickstein, Dara L.; Koo, Edward H.

2012-01-01

263

A quantitative method for detecting deposits of amyloid A protein in aspirated fat tissue of patients with arthritis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

OBJECTIVE—To describe a new, quantitative, and reproducible method for detecting deposits of amyloid A protein in aspirated fat tissue and to compare it with smears stained with Congo red.?METHODS—After extraction of at least 30 mg of abdominal fat tissue in guanidine, the amyloid A protein concentration was measured by a monoclonal antibody-based sandwich ELISA.?RESULTS—The concentrations in 24 patients with arthritis and AA amyloidosis (median 236, range 1.1-8530 ng/mg tissue) ...

Hazenberg, B.; Limburg, P.; Bijzet, J.; Rijswijk, M. H.

1999-01-01

264

Protein clearance mechanisms of alpha-synuclein and amyloid-Beta in lewy body disorders.  

Science.gov (United States)

Protein clearance is critical for the maintenance of the integrity of neuronal cells, and there is accumulating evidence that in most-if not all-neurodegenerative disorders, impaired protein clearance fundamentally contributes to functional and structural alterations eventually leading to clinical symptoms. Dysfunction of protein clearance leads to intra- and extraneuronal accumulation of misfolded proteins and aggregates. The pathological hallmark of Lewy body disorders (LBDs) is the abnormal accumulation of misfolded proteins such as alpha-synuclein (Asyn) and amyloid-beta (Abeta) in a specific subset of neurons, which in turn has been related to deficits in protein clearance. In this paper we will highlight common intraneuronal (including autophagy and unfolded protein stress response) and extraneuronal (including interaction of neurons with astrocytes and microglia, phagocytic clearance, autoimmunity, cerebrospinal fluid transport, and transport across the blood-brain barrier) protein clearance mechanisms, which may be altered across the spectrum of LBDs. A better understanding of the pathways underlying protein clearance-in particular of Asyn and Abeta-in LBDs may result in the identification of novel biomarkers for disease onset and progression and of new therapeutic targets. PMID:23133788

Deleidi, Michela; Maetzler, Walter

2012-01-01

265

Amyloid-clearing proteins and their epigenetic regulation as a therapeutic target in Alzheimer’s disease  

Directory of Open Access Journals (Sweden)

Full Text Available Abnormal elevation of amyloid ?-peptide (A? levels in the brain is the primary trigger for neuronal cell death specific to Alzheimer’s disease (AD. It is now evident that A? levels in the brain are manipulable due to a dynamic equilibrium between its production from the amyloid precursor protein (APP and removal by amyloid clearance proteins. Clearance can be either enzymic or non-enzymic (binding/transport proteins. Intriguingly several of the main amyloid-degrading enzymes (ADEs are members of the M13 peptidase family (neprilysin (NEP, NEP2 and the endothelin converting enzymes (ECE-1 and -2. A distinct metallopeptidase, insulin-degrading enzyme (IDE, also contributes to A? degradation in the brain. The ADE family currently embraces more than 20 members, both membrane-bound and soluble, and of differing cellular locations. NEP plays an important role in brain function terminating neuropeptide signals. Its decrease in specific brain areas with age or after hypoxia, ischaemia or stroke contribute significantly to the development of AD pathology. The recently discovered mechanism of epigenetic regulation of NEP (and other genes by the APP intracellular domain (AICD and its dependence on the cell type and APP isoform expression suggest possibilities for selective manipulation of NEP gene expression in neuronal cells. We have also observed that another amyloid-clearing protein, namely transthyretin (TTR, is also regulated in the neuronal cell by a mechanism similar to NEP. Dependence of amyloid clearance proteins on histone deacetylases and the ability of HDAC inhibitors to up-regulate their expression in the brain opens new avenues for developing preventive strategies in AD.

AnthonyJTurner

2014-09-01

266

Neuroinflammation in Lyme neuroborreliosis affects amyloid metabolism  

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Abstract Background The metabolism of amyloid precursor protein (APP) and ?-amyloid (A?) is widely studied in Alzheimer's disease, where A? deposition and plaque development are essential components of the pathogenesis. However, the physiological role of amyloid in the adult nervous system remains largely unknown. We have previously found altered cerebral amyloid metabolism in other neuroinflammatory conditions. To further elucidate this, we investigated amyloid metabolism...

Anckarsäter Henrik; Blennow Kaj; Anckarsäter Rolf; Mattsson Niklas; Hagberg Lars

2010-01-01

267

Effect of the beta secretase-1 inhibitor on the amyloid C-terminal fragment of amyloid precursor protein processing in a hyperphosphorylated tau rat model.  

Science.gov (United States)

The amyloid C-terminal fragment (?CTF) of the amyloid precursor protein (APP) is the cleaved component of APP by beta secretase-1 (BACE1), which shows similar neurotoxicity as amyloid beta (A?) in many ways. Evidence suggested that in addition to A?, ?CTF might also participate in the pathogenesis of Alzheimer's disease (AD). In recent years, the relationship between ?CTF processing and hyperphosphorylated tau has attracted increasing research attention. In this study, we established an animal model of tau hyperphosphorylation with okadaic acid (OA) treatment, and analyzed ?CTF processing in vivo. The ?CTF level was found to increase in neurons, which was most likely caused by the induction of OA and BACE1 overexpression. Furthermore, these results provide the first evidence that ?CTF can predominately accumulate in the axons of neurons in a hyperphosphorylated tau state in vivo, and suggested that the redistribution of ?CTF is involved in the pathogenesis of AD. These results indicate that BACE1 could be a therapeutic target of AD by affecting the processing of ?CTF. PMID:25158248

Chen, H Y; Wang, L; Liu, J F; Wang, W Z; Yu, C J

2014-01-01

268

Differential expression and redox proteomics analyses of an Alzheimer disease transgenic mouse model: effects of the amyloid-? peptide of amyloid precursor protein.  

Science.gov (United States)

Among the pathological factors known to be associated with Alzheimer disease (AD), oxidative stress induced by the amyloid-? peptide (A?) has been demonstrated to play a key role in human brain and animal models of AD. Recently, we reported elevated levels of oxidative damage in the brain of a transgenic (Tg) AD mouse model with Swedish and Indiana familial AD mutations in human amyloid precursor protein (APP) [PDAPP mice, line J20], as evidenced by increased levels of protein carbonyls, 3-nitrotyrosine, and protein-bound 4-hydroxy-2-nonenal. This oxidative damage was dependent on the methionine 35 residue within the A? peptide. Further insight into the molecular pathways affected in this Tg model of AD may be gained with discovery-based proteomics studies; therefore, two-dimensional gel-based expression proteomics was performed to compare differences in brain protein levels of J20 Tg mice with non-transgenic (NTg) littermate controls. Based on our studies, we identified six proteins that had significantly increased levels in J20 Tg relative to NTg mice: calcineurin subunit B type 1, ? GDP-dissociation inhibitor 1, T-complex protein 1 subunit ? A, ?-enolase, peptidyl-prolyl cis-trans isomerase (Pin-1), and ATP synthase subunit ? mitochondrial. Several of these proteins have previously been implicated in in vitro and in vivo models and subjects with AD. Additionally, using redox proteomics analyses we identified two oxidatively-modified proteins: phosphatidylethanolamine-binding protein 1 and Pin-1 with decreased levels of protein 3-nitrotyrosine in J20 Tg mice relative to NTg. Western blotting and immunoprecipitation analyses were used to validate proteomics results. Overall, these studies provide information about changes in the brain proteome as a result of A? deposition and clues with which to further direct studies on elucidating AD pathogenesis. PMID:21223993

Robinson, R A S; Lange, M B; Sultana, R; Galvan, V; Fombonne, J; Gorostiza, O; Zhang, J; Warrier, G; Cai, J; Pierce, W M; Bredesen, D E; Butterfield, D A

2011-03-17

269

Toward a molecular understanding of the detection of amyloid proteins with flexible conjugated oligothiophenes.  

Science.gov (United States)

Molecular and electronic structures and optical absorption properties of oligothiophenes used for spectral assignment of amyloid deposits have been investigated for a family of probes known as luminescent conjugated oligothiophenes (LCOs). Theoretical absorption spectra have been determined using conformational averaging, combining classical molecular dynamics (MD) simulations with quantum mechanical/molecular mechanics (QM/MM) time-dependent density functional theory (TD-DFT) spectrum calculations. Theoretical absorption spectra are in excellent agreement with experiments, showing average errors below 5 nm for absorption maxima. To couple observed properties to molecular structures, a measure of planarity is defined, revealing a strong correlation between the transition wavelength of the first and dominating electronically excited state and dihedral rotations. It is shown that from this correlation, predictions can be made of the absorption properties of probes based only on information from MD trajectories. We show experimentally that red shifts observed in the excitation maxima of LCOs when bound to amyloid protein aggregates are also evident in absorption spectra. We predict that these red shifts are due to conformational restriction of the LCO in a protein binding pocket, causing a planarization of the conjugated backbone. On the basis of our studies of planarity, it is shown that such shifts are both possible and realistic. PMID:25247879

Sjöqvist, Jonas; Maria, Jerôme; Simon, Rozalyn A; Linares, Mathieu; Norman, Patrick; Nilsson, K Peter R; Lindgren, Mikael

2014-10-23

270

Ketamine reduces amyloid ?-protein degradation by suppressing neprilysin expression in primary cultured astrocytes.  

Science.gov (United States)

Pathological accumulation of cortical amyloid ?-protein (A?) is an early and consistent feature of Alzheimer's disease (AD). A? levels in the brain are determined by production and catabolism. Previous studies have suggested that deficits in the brain expression of neprilysin (NEP) and the insulin-degrading enzyme (IDE), which are both proteases involved in amyloid degradation, may promote A? deposition in patients with sporadic late-onset AD. Because the incidence of AD increases after surgical intervention, we examined whether ketamine, which is a general anaesthetic with neuroprotective properties for excitotoxic ischaemic damage, is associated with A? degradation by inducing NEP and IDE expression. The non-competitive N-methyl-d-aspartate receptor antagonist ketamine and MK801 significantly decreased the expression of NEP, but not IDE, in a concentration- and time-dependent manner through the dephosphorylation of p38 mitogen-activated protein kinase (MAPK) in cultured rat astrocytes. Furthermore, NEP-reduced reagents significantly suppressed the degradation of exogenous A? in cultured astrocytes. These results suggested that ketamine suppresses the A? degradation of NEP by reducing p38 MAPK-mediated pathway activity. PMID:23624023

Yamamoto, Naoki; Arima, Hajime; Naruse, Kaori; Kasahara, Rika; Taniura, Hideo; Hirate, Hiroyuki; Sugiura, Takeshi; Suzuki, Kenji; Sobue, Kazuya

2013-06-17

271

AlphaB-crystallin, a small heat-shock protein, prevents the amyloid fibril growth of an amyloid beta-peptide and beta2-microglobulin.  

Science.gov (United States)

AlphaB-crystallin, a small heat-shock protein, exhibits molecular chaperone activity. We have studied the effect of alphaB-crystallin on the fibril growth of the Abeta (amyloid beta)-peptides Abeta-(1-40) and Abeta-(1-42). alphaB-crystallin, but not BSA or hen egg-white lysozyme, prevented the fibril growth of Abeta-(1-40), as revealed by thioflavin T binding, total internal reflection fluorescence microscopy and CD spectroscopy. Comparison of the activity of some mutants and chimaeric alpha-crystallins in preventing Abeta-(1-40) fibril growth with their previously reported chaperone ability in preventing dithiothreitol-induced aggregation of insulin suggests that there might be both common and distinct sites of interaction on alpha-crystallin involved in the prevention of amorphous aggregation of insulin and fibril growth of Abeta-(1-40). alphaB-crystallin also prevents the spontaneous fibril formation (without externally added seeds) of Abeta-(1-42), as well as the fibril growth of Abeta-(1-40) when seeded with the Abeta-(1-42) fibril seed. Sedimentation velocity measurements show that alphaB-crystallin does not form a stable complex with Abeta-(1-40). The mechanism by which it prevents the fibril growth differs from the known mechanism by which it prevents the amorphous aggregation of proteins. alphaB-crystallin binds to the amyloid fibrils of Abeta-(1-40), indicating that the preferential interaction of the chaperone with the fibril nucleus, which inhibits nucleation-dependent polymerization of amyloid fibrils, is the mechanism that is predominantly involved. We found that alphaB-crystallin prevents the fibril growth of beta2-microglobulin under acidic conditions. It also retards the depolymerization of beta2-microglobulin fibrils, indicating that it can interact with the fibrils. Our study sheds light on the role of small heat-shock proteins in protein conformational diseases, particularly in Alzheimer's disease. PMID:16053447

Raman, Bakthisaran; Ban, Tadato; Sakai, Miyo; Pasta, Saloni Y; Ramakrishna, Tangirala; Naiki, Hironobu; Goto, Yuji; Rao, Ch Mohan

2005-12-15

272

Interaction of Serum Proteins with Surface of Hemodialysis Fiber Membranes  

Science.gov (United States)

The poly(vinyl pyrrolidone)-covered hydrophilic surface of hollow-fiber membranes (fiber membrane, hereafter) for hemodialysis was mechanically probed using modified tips on an atomic force microscope (AFM) with covalent crosslinkers and several types of serum protein. The retraction part of many of the force extension (F-E) curves obtained with AFM tips coated with serum albumin had a long and smooth extension up to 200-300 nm indicating forced elongation of poly(vinyl pyrrolidone) chains. When fibrinogen-coated tips were used, long extension F-E curves up to 500 nm with multiple peaks were obtained in addition to smooth curves most likely reflecting the unfolding of fibrinogen molecules. The results indicated that individual polymer chains had a significant affinity toward serum proteins. The adhesion frequency of tips coated with serum proteins was lower on the poly(vinyl pyrrolidone) surface than on the uncoated hydrophobic polysulfone surface.

Afrin, Rehana; Shirako, Yuji; Kishimoto, Kikuo; Ikai, Atsushi

2012-08-01

273

How do membranes initiate Alzheimer's Disease? Formation of toxic amyloid fibrils by the amyloid ?-protein on ganglioside clusters.  

Science.gov (United States)

Alzheimer's disease (AD), a severe neurodegenerative disorder, causes more than half of dementia cases. According to the popular "A? hypothesis" to explain the mechanism of this disease, amyloid ?-peptides (A?) of 39-43 amino acid residues aggregate and deposit onto neurons, igniting the neurotoxic cascade of the disease. Therefore, researchers studying AD would like to elucidate the mechanisms by which essentially water-soluble but hydrophobic A? aggregates under pathological conditions. Most researchers have investigated the aggregation of A? in aqueous solution, and they concluded that the final aggregation product, the amyloid fibrils, were less toxic than the component peptide oligomers. They consequently shifted their interests to more toxic "soluble oligomers", structures that form as intermediates or off-pathway products during the aggregation process. Some researchers have also investigated artificial oligomers prepared under nonphysiological conditions. In contrast to these "in solution" studies, we have focused on "membrane-mediated" amyloidogenesis. In an earlier study, other researchers identified a specific form of A? that was bound to monosialoganglioside GM1, a sugar lipid, in brains of patients who exhibited the early pathological changes associated with AD. This Account summarizes 15 years of our research on this topic. We have found that A? specifically binds to GM1 that occurs in clusters, but not when it is uniformly distributed. Clustering is facilitated by cholesterol. Upon binding, A? changes its conformation from a random coil to an ?-helix-rich structure. A CH-? interaction between the aromatic side chains of A? and carbohydrate moieties appended to GM1 appears to be important for binding. In addition, as A? accumulates and reaches its first threshold concentration (A?/GM1 = ?0.013), aggregated ?-sheets of ?15 molecules appear and coexist with the helical form. However, this ?-structure is stable and does not form larger aggregates. When the disease progresses further and the A?/GM1 ratio exceeds ?0.044, the ?-structure converts to a second ?-structure that can seed aggregates. The seed recruits monomers from the aqueous phase to form toxic amyloid fibrils that have larger surface hydrophobicity and can contain antiparallel ?-sheets. In contrast, amyloid fibrils formed in aqueous solution are less toxic and have parallel ?-sheets. The less polar environments of GM1 clusters play an important role in the formation of these toxic fibrils. Membranes that contain GM1 clusters not only accelerate the aggregation of A? by locally concentrating A? molecules but also generate amyloid fibrils with unique structures and significant cytotoxicity. The inhibition of this aggregation cascade could be a promising strategy for the development of AD-modulating therapies. PMID:25029558

Matsuzaki, Katsumi

2014-08-19

274

Selective Vulnerability in Alzheimer's Disease: Amyloid Precursor Protein and p75NTR Interaction  

Science.gov (United States)

Objective Selective neuronal vulnerability in neurodegenerative diseases is poorly understood. In Alzheimer's disease, the basal forebrain cholinergic neurons are selectively vulnerable, putatively because of their expression of the cell death mediator p75NTR (the common neurotrophin receptor), and its interaction with proapoptotic ligands pro–nerve growth factor and amyloid-? peptide. However, the relation between amyloid precursor protein (APP) and p75NTR has not been described previously. Methods APP and p75NTR were assayed for interaction by coimmunoprecipitation in vitro and in vivo, yeast two-hybrid assay, bioluminescence resonance energy transfer, and confocal microscopy. Effects on APP processing and signaling were studied using immunoblotting, enzyme-linked immunosorbent assays, and luciferase reporter assays. Results The results of this study are as follows: (1) p75NTR and APP interact directly; (2) this interaction is modified by ligands nerve growth factor and ?-amyloid; (3) APP and p75NTR colocalization in vivo is modified in Alzheimer's model transgenic mice; (4) APP processing is altered by p75NTR, and to a lesser extent, p75NTR processing is altered by the presence of APP; (5) APP-dependent transcription mediated by Fe65 is blocked by p75NTR; and (6) coexpression of APP and p75NTR triggers cell death. Interpretation These results provide new insight into the emerging signaling network that mediates the Alzheimer's phenotype and into the mechanism of basal forebrain cholinergic neuronal selective vulnerability. In addition, the results argue that the interaction between APP and p75NTR may represent a therapeutic target in Alzheimer's disease. PMID:19334058

Fombonne, Joanna; Rabizadeh, Shahrooz; Banwait, Surita; Mehlen, Patrick; Bredesen, Dale E.

2011-01-01

275

Erythropoietin binding protein from mammalian serum  

Energy Technology Data Exchange (ETDEWEB)

Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described. 11 figs.

Clemons, G.K.

1997-04-29

276

Erythropoietin binding protein from mammalian serum  

Energy Technology Data Exchange (ETDEWEB)

Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described.

Clemons, Gisela K. (Berkeley, CA)

1997-01-01

277

Immunoglobulin light chains, glycosaminoglycans and amyloid.  

Energy Technology Data Exchange (ETDEWEB)

Immunoglobulin light chains are the precursor proteins for fibrils that are formed during primary amyloidosis and in amyloidosis associated with multiple myeloma. As found for the approximately 20 currently described forms of focal, localized, or systemic amyloidoses, light chain-related fibrils extracted from physiological deposits are invariably associated with glycosaminoglycans, predominantly heparan sulfate. Other amyloid-related proteins are either structurally normal, such as g2-microglobulin and islet amyloid polypeptide, fragments of normal proteins such as serum amyloid A protein or the precursor protein of the g peptide involved in Alzheimer's disease, or are inherited forms of single amino acid variants of a normal protein such as found in the familial forms of amyloid associated with transthyretin. In contrast, the primary structures of light chains involved in fibril formation exhibit extensive mutational diversity rendering some proteins highly amyloidogenic and others non-pathological. The interactions between light chains and glycosaminoglycans are also affected by amino acid variation and may influence the clinical course of disease by enhancing fibril stability and contributing to resistance to protease degradation. Relatively little is currently known about the mechanisms by which glycosaminoglycans interact with light chains and light-chain fibrils. It is probable that future studies of this uniquely diverse family of proteins will continue o shed light on the processes of amyloidosis, and contribute as well to a greater understanding of the normal physiological roles of glycosaminoglycans.

Stevens, F. J.; Kisilevsky, R.; Biosciences Division; Queen' s Univ.

2000-03-01

278

EGCG disaggregates amyloid-like fibrils formed by Plasmodium falciparum merozoite surface protein 2.  

Science.gov (United States)

Merozoite surface protein 2 (MSP2), one of the most abundant proteins on the surface of Plasmodium falciparum merozoites, is a promising malaria vaccine candidate. MSP2 is intrinsically unstructured and forms amyloid-like fibrils in solution. As this propensity of MSP2 to form fibrils in solution has the potential to impede its development as a vaccine candidate, finding an inhibitor that inhibits fibrillogenesis may enhance vaccine development. We have shown previously that EGCG inhibits the formation of MSP2 fibrils. Here we show that EGCG can alter the ?-sheet-like structure of the fibril and disaggregate pre-formed fibrils of MSP2 into soluble oligomers. The fibril remodelling effects of EGCG and other flavonoids were characterised using Thioflavin T fluorescence assays, electron microscopy and other biophysical methods. PMID:21784057

Chandrashekaran, Indu R; Adda, Christopher G; Macraild, Christopher A; Anders, Robin F; Norton, Raymond S

2011-09-15

279

Quantitating metabolites in protein precipitated serum using NMR spectroscopy.  

Science.gov (United States)

Quantitative NMR-based metabolite profiling is challenged by the deleterious effects of abundant proteins in the intact blood plasma/serum, which underscores the need for alternative approaches. Protein removal by ultrafiltration using low molecular weight cutoff filters thus represents an important step. However, protein precipitation, an alternative and simple approach for protein removal, lacks detailed quantitative assessment for use in NMR based metabolomics. In this study, we have comprehensively evaluated the performance of protein precipitation using methanol, acetonitrile, perchloric acid, and trichloroacetic acid and ultrafiltration approaches using 1D and 2D NMR, based on the identification and absolute quantitation of 44 human blood metabolites, including a few identified for the first time in the NMR spectra of human serum. We also investigated the use of a "smart isotope tag," (15)N-cholamine for further resolution enhancement, which resulted in the detection of a number of additional metabolites. (1)H NMR of both protein precipitated and ultrafiltered serum detected all 44 metabolites with comparable reproducibility (average CV, 3.7% for precipitation; 3.6% for filtration). However, nearly half of the quantified metabolites in ultrafiltered serum exhibited 10-74% lower concentrations; specifically, tryptophan, benzoate, and 2-oxoisocaproate showed much lower concentrations compared to protein precipitated serum. These results indicate that protein precipitation using methanol offers a reliable approach for routine NMR-based metabolomics of human blood serum/plasma and should be considered as an alternative to ultrafiltration. Importantly, protein precipitation, which is commonly used by mass spectrometry (MS), promises avenues for direct comparison and correlation of metabolite data obtained from the two analytical platforms to exploit their combined strength in the metabolomics of blood. PMID:24796490

Nagana Gowda, G A; Raftery, Daniel

2014-06-01

280

Insulin-like Growth Factor-1 (IGF-1)-induced Processing of Amyloid-? Precursor Protein (APP) and APP-like Protein 2 Is Mediated by Different Metalloproteinases*  

Digital Repository Infrastructure Vision for European Research (DRIVER)

?-Secretase cleavage of the amyloid precursor protein (APP) is of great interest because it prevents the formation of the Alzheimer-linked amyloid-? peptide. APP belongs to a conserved gene family including the two paralogues APP-like protein (APLP) 1 and 2. Insulin-like growth factor-1 (IGF-1) stimulates the shedding of all three proteins. IGF-1-induced shedding of both APP and APLP1 is dependent on phosphatidylinositol 3-kinase (PI3-K), whereas APLP2 shedding is independent of this signal...

Jacobsen, Kristin T.; Adlerz, Linda; Multhaup, Gerd; Iverfeldt, Kerstin

2010-01-01

 
 
 
 
281

Detection of human serum proteins using Raman and SERS spectroscopy  

Science.gov (United States)

The use of normal Raman (NR) spectroscopy and surface enhanced Raman scattering (SERS) spectroscopy to analyze the biochemical information of human serum proteins and hence distinguish between normal and primary hepatic carcinoma (PHC) serum samples was investigated. The serum samples were obtained from patients who were clinically diagnosed with PHC (n=20) and healthy volunteers (n=20). All spectra were collected in the spectral range of 400-1800 cm-1 and analyzed through the multivariate statistical methods of principal component analysis (PCA). The results showed that both NR and SERS combined with PCA had good performance in distinguishing the human serum proteins between PHC patients and healthy volunteers with high sensitivity and specificity of 100%. And we can get more detail information of component and conformation of human serum proteins by considering NR and SERS spectrum. Our results support the concept again that serum protein Raman and SERS spectroscopy combined with PCA analysis both can become noninvasive and rapid diagnostic tools to detect the primary hepatic carcinoma.

Ruan, Qiuyong; Liao, Fadian; Lin, Juqiang; Liu, Nenrong; Lin, Jinyong; Zeng, Yongyi; Li, Ling; Huang, Zufang; Chen, Rong

2014-09-01

282

Grizzly bear corticosteroid binding globulin: Cloning and serum protein expression.  

Science.gov (United States)

Serum corticosteroid levels are routinely measured as markers of stress in wild animals. However, corticosteroid levels rise rapidly in response to the acute stress of capture and restraint for sampling, limiting its use as an indicator of chronic stress. We hypothesized that serum corticosteroid binding globulin (CBG), the primary transport protein for corticosteroids in circulation, may be a better marker of the stress status prior to capture in grizzly bears (Ursus arctos). To test this, a full-length CBG cDNA was cloned and sequenced from grizzly bear testis and polyclonal antibodies were generated for detection of this protein in bear sera. The deduced nucleotide and protein sequences were 1218 bp and 405 amino acids, respectively. Multiple sequence alignments showed that grizzly bear CBG (gbCBG) was 90% and 83% identical to the dog CBG nucleotide and amino acid sequences, respectively. The affinity purified rabbit gbCBG antiserum detected grizzly bear but not human CBG. There were no sex differences in serum total cortisol concentration, while CBG expression was significantly higher in adult females compared to males. Serum cortisol levels were significantly higher in bears captured by leg-hold snare compared to those captured by remote drug delivery from helicopter. However, serum CBG expression between these two groups did not differ significantly. Overall, serum CBG levels may be a better marker of chronic stress, especially because this protein is not modulated by the stress of capture and restraint in grizzly bears. PMID:20347821

Chow, Brian A; Hamilton, Jason; Alsop, Derek; Cattet, Marc R L; Stenhouse, Gordon; Vijayan, Mathilakath M

2010-06-01

283

Biologically active protein fragments containing specific binding regions of serum albumin or related proteins  

Science.gov (United States)

In accordance with the present invention, biologically active protein fragments can be constructed which contain only those specific portions of the serum albumin family of proteins such as regions known as subdomains IIA and IIIA which are primarily responsible for the binding properties of the serum albumins. The artificial serums that can be prepared from these biologically active protein fragments are advantageous in that they can be produced much more easily than serums containing the whole albumin, yet still retain all or most of the original binding potential of the full albumin proteins. In addition, since the protein fragment serums of the present invention can be made from non-natural sources using conventional recombinant DNA techniques, they are far safer than serums containing natural albumin because they do not carry the potentially harmful viruses and other contaminants that will be found in the natural substances.

Carter, Daniel C. (Inventor)

1998-01-01

284

Serum amyloid A (SAA) as a biomarker of chronic infection due to boat strike trauma in a free-ranging Florida manatee (Trichechus manatus latirostris) with incidental polycystic kidneys  

Science.gov (United States)

Watercraft-related trauma is the predominant cause of human-induced mortality in manatees (Trichechus manatus latirostris), a federal- and state-listed endangered species. Pyothorax (documented in this case report) and other secondary infections are common sequelae of inhalation of water and the open wounds caused by boat propellers. These secondary infections can lead to the demise of the animal weeks to months after the traumatic incident when external wounds have healed. Diagnosis of underlying disease on physical examination during capture and restraint can be difficult. Acute phase proteins, including serum amyloid A, fibrinogen, and albumin can be used to diagnose inflammatory disease in manatees and improve quality of medical care and husbandry. We also provide the first report of polycystic kidneys in Sirenians.

Harr, Kendal E.; Rember, Renee; Ginn, Pamela E.; Lightsey, Jessica; Keller, Martha; Reid, James; Bonde, Robert K.

2011-01-01

285

NMDA receptor/amyloid precursor protein interactions: a comparison between wild-type and amyloid precursor protein mutations associated with familial Alzheimer's disease.  

Science.gov (United States)

Two recent reports showed that amyloid precursor protein (APP) may contribute to postsynaptic mechanisms via the regulation of the surface trafficking of excitatory N-methyl-D-aspartate (NMDA) receptors. Here we have investigated the interactions and surface trafficking of NR1-1a/NR2A and NR1-1a/NR2B NMDA receptor subtypes with three APP mutations linked to familial Alzheimer's disease, APP695(Indiana), APP695(London) and APP695(Swedish). Flag-tagged mutated APP695s were generated and shown to be expressed at equivalent levels to wild-type APP695 in mammalian cells. Each APP mutant co-precipitated with NR1-1a/NR2A and NR1-1a/NR2B receptors following co-expression in mammalian cells. Further, as found for wild-type APP695, each enhanced NMDA receptor surface expression with no concomitant increase in total NR1-1a, NR2A or NR2B subunit expression. Thus these three familial APP mutations behave as wild-type APP695 with respect to their association with assembled NMDA receptors and their APP695-enhanced receptor cell surface trafficking. PMID:22450047

Innocent, Neal; Cousins, Sarah L; Stephenson, F Anne

2012-05-01

286

ADAM10 is the physiologically relevant, constitutive ?-secretase of the amyloid precursor protein in primary neurons  

Science.gov (United States)

The amyloid precursor protein (APP) undergoes constitutive shedding by a protease activity called ?-secretase. This is considered an important mechanism preventing the generation of the Alzheimer's disease amyloid-? peptide (A?). ?-Secretase appears to be a metalloprotease of the ADAM family, but its identity remains to be established. Using a novel ?-secretase-cleavage site-specific antibody, we found that RNAi-mediated knockdown of ADAM10, but surprisingly not of ADAM9 or 17, completely suppressed APP ?-secretase cleavage in different cell lines and in primary murine neurons. Other proteases were not able to compensate for this loss of ?-cleavage. This finding was further confirmed by mass-spectrometric detection of APP-cleavage fragments. Surprisingly, in different cell lines, the reduction of ?-secretase cleavage was not paralleled by a corresponding increase in the A?-generating ?-secretase cleavage, revealing that both proteases do not always compete for APP as a substrate. Instead, our data suggest a novel pathway for APP processing, in which ADAM10 can partially compete with ?-secretase for the cleavage of a C-terminal APP fragment generated by ?-secretase. We conclude that ADAM10 is the physiologically relevant, constitutive ?-secretase of APP. PMID:20676056

Kuhn, Peer-Hendrik; Wang, Huanhuan; Dislich, Bastian; Colombo, Alessio; Zeitschel, Ulrike; Ellwart, Joachim W; Kremmer, Elisabeth; Rossner, Steffen; Lichtenthaler, Stefan F

2010-01-01

287

Lead exposure in pheochromocytoma cells induces persistent changes in amyloid precursor protein gene methylation patterns.  

Science.gov (United States)

It has been suggested that lead (Pb) exposure in early life may increase amyloid precursor protein (APP) expression and promote the pathogenesis of Alzheimer's disease in old age. The current study examined whether the DNA methylation patterns of APP gene in rat pheochromocytoma (PC12) cells changed after Pb acetate exposure. Undifferentiated PC12 cells were exposed to three doses of Pb acetate (50, 250, and 500 nM) and one control for 2 days or 1 week. The methylation patterns of APP promoter and global DNA methylation were analyzed. The DNA methyltransferase 1 (DNMT1) expression and the level of amyloid ? peptide (A?) were also investigated. The results showed that the exposure of the three concentrations of Pb acetate could make the APP promoter hypomethylated. The global DNA methylation level and the expression of DNMT1 were changed in the 500 nM group after 2 days exposure and in the 250 and 500 nM group after 7 days exposure. Thus, Pb may exert neurotoxic effects through mechanisms that alter the global and promoter methylation patterns of APP gene. © 2010 Wiley Periodicals, Inc. Environ Toxicol, 2012. PMID:22764079

Li, Yuan-Yuan; Chen, Tian; Wan, Yanjian; Xu, Shun-qing

2012-08-01

288

Structural Insights into Amyloid Oligomers of the Parkinson Disease-related Protein ?-Synuclein.  

Science.gov (United States)

The presence of intraneuronal deposits mainly formed by amyloid fibrils of the presynaptic protein ?-synuclein (AS) is a hallmark of Parkinson disease. Currently, neurotoxicity is attributed to prefibrillar oligomeric species rather than the insoluble aggregates, although their mechanisms of toxicity remain elusive. Structural details of the supramolecular organization of AS oligomers are critically needed to decipher the structure-toxicity relationship underlying their pathogenicity. In this study, we employed site-specific fluorescence to get a deeper insight into the internal architecture of AS oligomeric intermediates. We demonstrate that AS oligomers are ordered assemblies possessing a well defined pattern of intermolecular contacts. Some of these contacts involve regions that form the ?-sheet core in the fibrillar state, although their spatial arrangement may differ in the two aggregated forms. However, even though the two termini are excluded from the fibrillar core, they are engaged in a number of intermolecular interactions within the oligomer. Therefore, substantial structural remodeling of early oligomeric interactions is essential for fibril growth. The intermolecular contacts identified in AS oligomers can serve as targets for the rational design of anti-amyloid compounds directed at preventing oligomeric interactions/reorganizations. PMID:25143382

Gallea, J Ignacio; Celej, M Soledad

2014-09-26

289

Beta-secretase-cleaved amyloid precursor protein in Alzheimer brain: a morphologic study  

DEFF Research Database (Denmark)

beta-amyloid (Abeta) is the main constituent of senile plaques seen in Alzheimer's disease. Abeta is derived from the amyloid precursor protein (APP) via proteolytic cleavage by proteases beta- and gamma-secretase. In this study, we examined content and localization of beta-secretase-cleaved APP (beta-sAPP) in brain tissue sections from the frontal, temporal and occipital lobe. Strong granular beta-sAPP staining was found throughout the gray matter of all three areas, while white matter staining was considerably weaker. beta-sAPP was found to be localized in astrocytes and in axons. We found the beta-sAPP immunostaining to be stronger and more extensive in gray matter in Alzheimer disease (AD) cases than controls. The axonal beta-sAPP staining was patchy and unevenly distributed for the AD cases, indicating impaired axonal transport. beta-sAPP was also found surrounding senile plaques and cerebral blood vessels. The results presented here show altered beta-sAPP staining in the AD brain, suggestive of abnormalprocessing and transport of APP.

Sennvik, Kristina; Bogdanovic, N

2004-01-01

290

Purification and cloning of amyloid precursor protein beta-secretase from human brain.  

Science.gov (United States)

Proteolytic processing of the amyloid precursor protein (APP) generates amyloid beta (Abeta) peptide, which is thought to be causal for the pathology and subsequent cognitive decline in Alzheimer's disease. Cleavage by beta-secretase at the amino terminus of the Abeta peptide sequence, between residues 671 and 672 of APP, leads to the generation and extracellular release of beta-cleaved soluble APP, and a corresponding cell-associated carboxy-terminal fragment. Cleavage of the C-terminal fragment by gamma-secretase(s) leads to the formation of Abeta. The pathogenic mutation K670M671-->N670L671 at the beta-secretase cleavage site in APP, which was discovered in a Swedish family with familial Alzheimer's disease, leads to increased beta-secretase cleavage of the mutant substrate. Here we describe a membrane-bound enzyme activity that cleaves full-length APP at the beta-secretase cleavage site, and find it to be the predominant beta-cleavage activity in human brain. We have purified this enzyme activity to homogeneity from human brain using a new substrate analogue inhibitor of the enzyme activity, and show that the purified enzyme has all the properties predicted for beta-secretase. Cloning and expression of the enzyme reveals that human brain beta-secretase is a new membrane-bound aspartic proteinase. PMID:10591214

Sinha, S; Anderson, J P; Barbour, R; Basi, G S; Caccavello, R; Davis, D; Doan, M; Dovey, H F; Frigon, N; Hong, J; Jacobson-Croak, K; Jewett, N; Keim, P; Knops, J; Lieberburg, I; Power, M; Tan, H; Tatsuno, G; Tung, J; Schenk, D; Seubert, P; Suomensaari, S M; Wang, S; Walker, D; Zhao, J; McConlogue, L; John, V

1999-12-01

291

Systematic study of plasma and serum proteins in the pig  

International Nuclear Information System (INIS)

This work has been carried out in the framework of the determination of the physiological constants of a normal pig. The aim was to study the serum and plasma proteins of this animal species, the ultimate object being to discover whether the qualitative and quantitative changes in these proteins can make a significant contribution to the establishment of a biological dosimetry for irradiated pigs. The serum and plasma from a normal pig were analyzed first by various simple electrophoretic methods and then by immuno-electrophoresis. As a result of the particular characteristics of pig serum we have gradually been led to make numerous modifications to the techniques used for human serums or for those of small laboratory animals. Much careful work and patience were required in order to obtain reproducible results. (authors)

292

A folded and functional protein domain in an amyloid-like fibril.  

Science.gov (United States)

The effect of the polypeptide environment on polyalanine-induced fibril formation was investigated with amyloidogenic fragments from PAPBN1, a nuclear protein controlling polyadenylation. Mutation-caused extensions of the natural 10 alanine sequence up to maximally 17 alanines result in fibril formation of PABPN1 and the development of the disease oculopharyngeal muscular dystrophy (OPMD). We explored the influence of fibril formation on the structure and function of a one-domain protein linked to the fibril-forming part of PABPN1. The well-characterized, stably folded, one-domain protein, cold-shock protein CspB from Bacillus subtilis, was fused either to the C terminus of the entire N-terminal domain of PABPN1 or directly to peptides consisting of 10 or 17 alanine residues. The fusion protein between the N-terminal domain of PABPN1 and CspB formed fibrils in which the structure and activity of CspB were retained. In the fibrils formed by fusions in which the polyalanine sequence was directly linked to CspB, CspB was unfolded. These results indicate that the folded conformation and the function of a protein domain can be maintained in amyloid-like fibrils, and that the distance between this domain and the fibril plays an important role. PMID:18424511

Sackewitz, Mirko; von Einem, Sabrina; Hause, Gerd; Wunderlich, Michael; Schmid, Franz-Xaver; Schwarz, Elisabeth

2008-06-01

293

Non-chaperone Proteins Can Inhibit Aggregation and Cytotoxicity of Alzheimer Amyloid ? Peptide.  

Science.gov (United States)

Many factors are known to influence the oligomerization, fibrillation, and amyloid formation of the A? peptide that is associated with Alzheimer disease. Other proteins that are present when A? peptides deposit in vivo are likely to have an effect on these aggregation processes. To separate specific versus broad spectrum effects of proteins on A? aggregation, we tested a series of proteins not reported to have chaperone activity: catalase, pyruvate kinase, albumin, lysozyme, ?-lactalbumin, and ?-lactoglobulin. All tested proteins suppressed the fibrillation of Alzheimer A?(1-40) peptide at substoichiometric ratios, albeit some more effectively than others. All proteins bound non-specifically to A?, stabilized its random coils, and reduced its cytotoxicity. Surprisingly, pyruvate kinase and catalase were at least as effective as known chaperones in inhibiting A? aggregation. We propose general mechanisms for the broad-spectrum inhibition A? fibrillation by proteins. The mechanisms we discuss are significant for prognostics and perhaps even for prevention and treatment of Alzheimer disease. PMID:25100721

Luo, Jinghui; Wärmländer, Sebastian K T S; Gräslund, Astrid; Abrahams, Jan Pieter

2014-10-01

294

Lipoprotein receptor-related protein-1 mediates amyloid-beta-mediated cell death of cerebrovascular cells.  

Science.gov (United States)

Inefficient clearance of A beta, caused by impaired blood-brain barrier crossing into the circulation, seems to be a major cause of A beta accumulation in the brain of late-onset Alzheimer's disease patients and hereditary cerebral hemorrhage with amyloidosis Dutch type. We observed association of receptor for advanced glycation end products, CD36, and low-density lipoprotein receptor (LDLR) with cerebral amyloid angiopathy in both Alzheimer's disease and hereditary cerebral hemorrhage with amyloidosis Dutch type brains and increased low-density lipoprotein receptor-related protein-1 (LRP-1) expression by perivascular cells in cerebral amyloid angiopathy. We investigated if these A beta receptors are involved in A beta internalization and in A beta-mediated cell death of human cerebrovascular cells and astrocytes. Expression of both the LRP-1 and LDLR by human brain pericytes and leptomeningeal smooth muscle cells, but not by astrocytes, increased on incubation with A beta. Receptor-associated protein specifically inhibited A beta-mediated up-regulation of LRP-1, but not of LDLR, and receptor-associated protein also decreased A beta internalization and A beta-mediated cell death. We conclude that especially LRP-1 and, to a minor extent, LDLR are involved in A beta internalization by and A beta-mediated cell death of cerebral perivascular cells. Although perivascular cells may adapt their A beta internalization capacity to the levels of A beta present, saturated LRP-1/LDLR-mediated uptake of A beta results in degeneration of perivascular cells. PMID:18055545

Wilhelmus, Micha M M; Otte-Höller, Irene; van Triel, Jos J J; Veerhuis, Robert; Maat-Schieman, Marion L C; Bu, Guojun; de Waal, Robert M W; Verbeek, Marcel M

2007-12-01

295

Smoking and serum proteins in atomic bomb survivors in Hiroshima  

International Nuclear Information System (INIS)

Associations of smoking habit with serum levels of total protein as well as protein fractions were studied in a population consisting of 4,739 atomic bomb survivors and unexposed control subjects in Hiroshima who participated in the 1979-81 period of the Adult Health Study, an on-going health follow-up program of the RERF. Smoking was strongly related to serum protein concentration after correction for age, sex, and body mass index. Among current smokers as compared to nonsmokers, levels of total protein, ? globulin, and ? globulin were significantly lower (p1 and ?2 globulin were significantly higher (p1 globulin. Duration of smoking (years) was related to increased ?1 and ?2 globulin. Smoking duration was also associated with albumin level but the trend was not monotonic. The radiation exposure effect on serum protein level was significant in several instances but was in general much smaller than the smoking effect. Its inclusion in the regression models did not noticeably affect the association between smoking and serum proteins. (author)

296

An online nano-LC-ESI-FTICR-MS method for comprehensive characterization of endogenous fragments from amyloid ? and amyloid precursor protein in human and cat cerebrospinal fluid.  

Science.gov (United States)

Amyloid precursor protein (APP) is the precursor protein to amyloid ? (A?), the main constituent of senile plaques in Alzheimer's disease (AD). Endogenous A? peptides reflect the APP processing, and greater knowledge of different APP degradation pathways is important to understand the mechanism underlying AD pathology. When one analyzes longer A? peptides by low-energy collision-induced dissociation tandem mass spectrometry (MS/MS), mainly long b-fragments are observed, limiting the possibility to determine variations such as amino acid variants or post-translational modifications (PTMs) within the N-terminal half of the peptide. However, by using electron capture dissociation (ECD), we obtained a more comprehensive sequence coverage for several APP/A? peptide species, thus enabling a deeper characterization of possible variants and PTMs. Abnormal APP/A? processing has also been described in the lysosomal storage disease Niemann-Pick type C and the major large animal used for studying this disease is cat. By ECD MS/MS, a substitution of Asp7 ? Glu in cat A? was identified. Further, sialylated core 1 like O-glycans at Tyr10, recently discovered in human A? (a previously unknown glycosylation type), were identified also in cat cerebrospinal fluid (CSF). It is therefore likely that this unusual type of glycosylation is common for (at least) species belonging to the magnorder Boreoeutheria. We here describe a detailed characterization of endogenous APP/A? peptide species in CSF by using an online top-down MS-based method. PMID:22576872

Brinkmalm, Gunnar; Portelius, Erik; Öhrfelt, Annika; Mattsson, Niklas; Persson, Rita; Gustavsson, Mikael K; Vite, Charles H; Gobom, Johan; Månsson, Jan-Eric; Nilsson, Jonas; Halim, Adnan; Larson, Göran; Rüetschi, Ulla; Zetterberg, Henrik; Blennow, Kaj; Brinkmalm, Ann

2012-05-01

297

Genetic and environmental influences of surfactant protein D serum levels  

DEFF Research Database (Denmark)

The collectin surfactant protein D (SP-D) is an important component of the pulmonary innate immune system, but SP-D is also present on extrapulmonary epithelial surfaces and in serum, where it has been used as a biomarker for pulmonary disease states. In this study, we investigate the mechanisms defining the constitutional serum level of SP-D and determine the magnitude of the genetic contribution to serum SP-D in the adult population. Recent studies have demonstrated that serum SP-D concentrations in children are genetically determined and that a single nucleotide polymorphism (SNP) located in the NH(2)-terminal region (Met11Thr) of the mature protein is significantly associated with the serum SP-D levels. A classic twin study was performed on a twin population including 1,476 self-reported healthy adults. The serum SP-D levels increased with male sex, age, and smoking status. The intraclass correlation was significantly higher for monozygotic (MZ) twin pairs than for dizygotic (DZ) twin pairs. Serum SP-D variance was influenced by nonshared environmental effects and additive genetic effects. Multivariate analysis of MZ and DZ covariance matrixes showed significant genetic correlation among serum SP-D and metabolic variables. The Met11Thr variant explained a significant part of the heritability indicating that serum SP-D variance could be decomposed into non-shared environmental effects (e(2) = 0.19), additive genetic effects (h(2) = 0.42), and the effect of the Met11Thr variations (q(2) = 0.39).

Sorensen, GL; Hjelmborg, JV

2006-01-01

298

Oxidative stress induces macroautophagy of amyloid beta-protein and ensuing apoptosis  

DEFF Research Database (Denmark)

There is increasing evidence for the toxicity of intracellular amyloid beta-protein (Abeta) to neurons and the involvement of lysosomes in this process in Alzheimer disease (AD). We have recently shown that oxidative stress, a recognized determinant of AD, enhances macroautophagy and leads to intralysosomal accumulation of Abeta in cultured neuroblastoma cells. We hypothesized that oxidative stress promotes AD by stimulating macroautophagy of Abeta that further may induce cell death by destabilizing lysosomal membranes. To investigate such possibility, we compared the effects of hyperoxia (40% ambient oxygen) in cultured HEK293 cells that were transfected with an empty vector (Vector), wild-type APP (APPwt), or Swedish mutant APP (APPswe). Exposure to hyperoxia for 5 days increased the number of cells with Abeta-containing lysosomes, as well as the number of apoptotic cells, compared to normoxic conditions. The rate of apoptosis in all three cell lines demonstrated dependence on intralysosomal Abeta content (Vector

Benedikz, Eirikur

2009-01-01

299

Characterization of brevin, a serum protein that shortens actin filaments  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We have purified a protein from rabbit serum with a molecular weight of 90,000 that inhibits the polymerization of actin measured viscometrically and that we have named “brevin” (from the Latin breviare, to shorten). From the extent of purification, we estimate that this inhibitor constitutes 0.3% of the total protein in plasma and serum. Brevin is also present in sera from humans and rats. Almost all of the activity in blood is extracellular; only 1% is present in platelets or other cell...

Harris, David A.; Schwartz, James H.

1981-01-01

300

Synaptic and sub-synaptic localization of amyloid-? protein precursor in the rat hippocampus.  

Science.gov (United States)

Amyloid-? protein precursor (A?PP) is a large transmembrane protein highly expressed in the central nervous system and cleavage of it can produce amyloid-? peptides (A?) involved in synaptic dysfunction and loss associated with cognitive impairment in Alzheimer's disease (AD). Surprisingly, little is known about the synaptic and sub-synaptic distribution of A?PP in different types of nerve terminals. We used total, synaptic, sub-synaptic, and astrocytic membrane preparations obtained from the hippocampus of adult rats to define the localization of A?PP, using two different antibodies against different A?PP epitopes. Western blot analysis revealed that A?PP was not significantly enriched in synaptosomal as compared to total membranes. Within synapses, A?PP immunoreactivity was more abundant in pre- (60 ± 4%) than post- (30 ± 5%) or extra-synaptic fractions (10 ± 2%). Immunocytochemical analysis of purified nerve terminals indicated that A?PP was more frequently associated with glutamatergic (present in 31 ± 4% of glutamatergic terminals) rather than with GABAergic (16 ± 3%) or cholinergic terminals (4 ± 1%, n = 4). We also observed a general lack of co-localization of A?PP and GFAP immunoreactivities in the hippocampus of sections of adult rat brain, albeit we could detect the presence of A?PP in gliosomes (vesicular specializations of astrocytic membranes), suggesting that A?PP has a heterogeneous localization restricted to certain regions of astrocytes. These results provide the first direct demonstration that A?PP is mostly distributed among glutamatergic rather than GABAergic or cholinergic terminals of the adult rat hippocampus, in remarkable agreement with the particular susceptibility to dysfunction and degeneration of glutamatergic synapses in early AD. PMID:24531160

Rodrigues, Diana I; Gutierres, Jessié; Pliássova, Anna; Oliveira, Catarina R; Cunha, Rodrigo A; Agostinho, Paula

2014-01-01

 
 
 
 
301

Relevance of amyloid precursor-like protein 2 C-terminal fragments in pancreatic cancer cells  

Science.gov (United States)

In some cellular systems, particularly neurons, amyloid precursor-like protein 2 (APLP2), and its highly homologous family member amyloid precursor protein (APP), have been linked to cellular growth. APLP2 and APP undergo regulated intramembrane proteolysis to produce C-terminal fragments. In this study, we found comprehensive expression of APLP2 C-terminal fragments in a panel of pancreatic cancer cell lines; however, APP C-terminal fragments were notably limited to the BxPC3 cell line. Extensive glycosaminoglycan modification on APLP2 was also found in the majority of pancreatic cancer cell lines. Glycosaminoglycan-modified and -unmodified APLP2, and particularly APLP2 C-terminal fragments, also demonstrated increased expression in oncogene-transformed pancreatic ductal cells. Additionally, elevated APLP2 levels were confirmed in human pancreatic cancer tissue. Downregulation of APLP2 and APP expression, alone or in combination, caused a decrease in the growth of a pancreatic cancer cell line with representatively low APP C-terminal fragment expression, the S2-013 cell line. Furthermore, we found that treatment with ?-secretase inhibitors to block formation of APLP2 C-terminal fragments decreased the growth and viability of S2-013 cells, without affecting the survival of a non-transformed pancreatic ductal cell line. In conclusion, our studies demonstrate that abundant APLP2, but not APP, C-terminal fragment expression is conserved in pancreatic cancer cell lines; however, APP and APLP2 equally regulated the growth of S2-013 pancreatic cancer cells. Chiefly, our discoveries establish a role for APLP2 in the growth of pancreatic cancer cells and show that inhibitors preventing APLP2 cleavage reduce the viability of pancreatic cancer cells. PMID:22797723

PETERS, HALEY L.; TULI, AMIT; WANG, XIAOJIAN; LIU, CUILING; PAN, ZENGGANG; OUELLETTE, MICHEL M.; HOLLINGSWORTH, MICHAEL A.; MacDONALD, RICHARD G.; SOLHEIM, JOYCE C.

2012-01-01

302

Amyloid precursor protein regulates migration and metalloproteinase gene expression in prostate cancer cells.  

Science.gov (United States)

Amyloid precursor protein (APP) is a type I transmembrane protein, and one of its processed forms, ?-amyloid, is considered to play a central role in the development of Alzheimer's disease. We previously showed that APP is a primary androgen-responsive gene in prostate cancer and that its increased expression is correlated with poor prognosis for patients with prostate cancer. APP has also been implicated in several human malignancies. Nevertheless, the mechanism underlying the pro-proliferative effects of APP on cancers is still not well-understood. In the present study, we explored a pathophysiological role for APP in prostate cancer cells using siRNA targeting APP (siAPP). The proliferation and migration of LNCaP and DU145 prostate cancer cells were significantly suppressed by siAPP. Differentially expressed genes in siAPP-treated cells compared to control siRNA-treated cells were identified by microarray analysis. Notably, several metalloproteinase genes, such as ADAM10 and ADAM17, and epithelial-mesenchymal transition (EMT)-related genes, such as VIM, and SNAI2, were downregulated in siAPP-treated cells as compared to control cells. The expression of these genes was upregulated in LNCaP cells stably expressing APP when compared with control cells. APP-overexpressing LNCaP cells exhibited enhanced migration in comparison to control cells. These results suggest that APP may contribute to the proliferation and migration of prostate cancer cells by modulating the expression of metalloproteinase and EMT-related genes. PMID:25218471

Miyazaki, Toshiaki; Ikeda, Kazuhiro; Horie-Inoue, Kuniko; Inoue, Satoshi

2014-09-26

303

Independent Relationship between Amyloid Precursor Protein (APP) Dimerization and ?-Secretase Processivity  

Science.gov (United States)

Altered production of ?-amyloid (A?) from the amyloid precursor protein (APP) is closely associated with Alzheimer’s disease (AD). APP has a number of homo- and hetero-dimerizing domains, and studies have suggested that dimerization of ?-secretase derived APP carboxyl terminal fragment (CTF?, C99) impairs processive cleavage by ?-secretase increasing production of long A?s (e.g., A?1-42, 43). Other studies report that APP CTF? dimers are not ?-secretase substrates. We revisited this issue due to observations made with an artificial APP mutant referred to as 3xK-APP, which contains three lysine residues at the border of the APP ectodomain and transmembrane domain (TMD). This mutant, which dramatically increases production of long A?, was found to form SDS-stable APP dimers, once again suggesting a mechanistic link between dimerization and increased production of long A?. To further evaluate how multimerization of substrate affects both initial ?-secretase cleavage and subsequent processivity, we generated recombinant wild type- (WT) and 3xK-C100 substrates, isolated monomeric, dimeric and trimeric forms of these proteins, and evaluated both ?-cleavage site utilization and A? production. These show that multimerization significantly impedes ?-secretase cleavage, irrespective of substrate sequence. Further, the monomeric form of the 3xK-C100 mutant increased long A? production without altering the initial ?-cleavage utilization. These data confirm and extend previous studies showing that dimeric substrates are not efficient ?-secretase substrates, and demonstrate that primary sequence determinants within APP substrate alter ?-secretase processivity. PMID:25350374

Jung, Joo In; Premraj, Sasha; Cruz, Pedro E.; Ladd, Thomas B.; Kwak, Yewon; Koo, Edward H.; Felsenstein, Kevin M.; Golde, Todd E.; Ran, Yong

2014-01-01

304

Expression of the C terminus of the amyloid precursor protein alters growth factor responsiveness in stably transfected PC12 cells.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The amyloid precursor protein (APP) is a molecule centrally involved in Alzheimer disease pathology, but whose normal function is still poorly understood. To investigate the consequences of increased intracellular production of various regions of APP on cellular physiology, we stably transfected PC12 cells with the C-terminal 100 amino acids of the human APP. In eight transfected clones that express the APP(C100) protein, exposure to nerve growth factor (NGF) did not promote differentiation. ...

Sandhu, F. A.; Kim, Y.; Lapan, K. A.; Salim, M.; Aliuddin, V.; Zain, S. B.

1996-01-01

305

Serum protein inhibition of thyrotropin binding to human thyroid tissue  

International Nuclear Information System (INIS)

We used a modificaton of the TSH radioreceptor assay to detect TSH-binding inhibition (TBI) activity in serum and serum fractions from normal subjects and patients with Graves' disease. TBI activity is present in normal IgG prepared by DEAE-Sephadex chromatography and in normal globulins prepared by precipitation at 1.6 M ammonium sulfate. Other normal serum proteins also had TBI activity when large concentrations were tested. Gel filtration chromatography and powder block electrophoresis were used to prepare fractions of normal and Graves' disease sera. In these fractions from normal serum, TBI activity was found in both ?-globulin and ?-globulin-albumin fractions electrophoretically and in both 7S and 4S peaks from gel filtration. TBI activity from Graves' disease patients' sera was similarly distributed, but relatively more TBI accompanied the electrophoretic ?-globulins. Sepharose Protein-A and anti-IgG were used as immunoabsorbents to isolate and purify IgG from normal and Graves' disease sera. TBI activity in IgG was proportional to the IgG concentration, indicating that the TBI which migrates as a ?-globulin electrophoretically is an IgG and thus may possibly be an antibody. Inhibitory activity found in normal serum globulins and in the non-IgG fractions of both normal and abnormal sera seriously interferes with attempts to use the TSH radioreceptor assay to study the hypothesized anti-TSH receptor antibody in the serum of patients with Graves' disease of patients with Graves' disease

306

P-glycoprotein efflux and other factors limit brain amyloid beta reduction by beta-site amyloid precursor protein-cleaving enzyme 1 inhibitors in mice.  

Science.gov (United States)

Alzheimer's disease (AD) is a progressive neurodegenerative disease. Amyloid beta (Abeta) peptides are hypothesized to cause the initiation and progression of AD based on pathologic data from AD patients, genetic analysis of mutations that cause early onset forms of AD, and preclinical studies. Based on this hypothesis, beta-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1) inhibitors are an attractive therapeutic approach for AD because cleavage of the APP by BACE1 is required to form Abeta. In this study, three potent BACE1 inhibitors are characterized. All three inhibitors decrease Abeta formation in cultured cells with IC(50) values less than 10 nM. Analysis of APP C-terminal fragments by immunoblotting and Abeta peptides by mass spectrometry showed that these inhibitors decreased Abeta by inhibiting BACE1. An assay for Abeta1-40 in mice was developed and used to show that these BACE1 inhibitors decreased plasma Abeta1-40, but not brain Abeta1-40, in wild-type mice. Because these BACE1 inhibitors were substrates for P-glycoprotein (P-gp), a member of the ATP-binding cassette superfamily of efflux transporters, these inhibitors were administered to P-gp knockout (KO) mice. These studies showed that all three BACE1 inhibitors decreased brain Abeta1-40 in P-gp KO mice, demonstrating that P-gp is a major limitation for development of BACE1 inhibitors to test the amyloid hypothesis. A comparison of plasma Abeta1-40 and brain Abeta1-40 dose responses for these three compounds revealed differences in relative ED(50) values, indicating that factors other than P-gp can also contribute to poor brain activity by BACE1 inhibitors. PMID:18499745

Meredith, Jere E; Thompson, Lorin A; Toyn, Jeremy H; Marcin, Lawrence; Barten, Donna M; Marcinkeviciene, Jovita; Kopcho, Lisa; Kim, Young; Lin, Alan; Guss, Valerie; Burton, Catherine; Iben, Lawrence; Polson, Craig; Cantone, Joe; Ford, Michael; Drexler, Dieter; Fiedler, Tracey; Lentz, Kimberley A; Grace, James E; Kolb, Janet; Corsa, Jason; Pierdomenico, Maria; Jones, Kelli; Olson, Richard E; Macor, John E; Albright, Charles F

2008-08-01

307

Binding of an oxindole alkaloid from Uncaria tomentosa to amyloid protein (Abeta1-40).  

Science.gov (United States)

The primary aim of this work was to determine the interactions of an oxindole alkaloid (mitraphylline) isolated from Uncaria tomentosa with beta-amyloid 1-40 (Abeta1-40 protein) applying the capillary electrophoresis (CE) method. Specifically the Hummel-Dreyer method and Scatchard analysis were performed to study the binding of oxindole alkaloids with Abeta1-40 protein. Prior to these studies extraction of the alkaloid of interest was carried out. Identification of the isolated alkaloid was performed by the use of thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC) combined with electrospray ionization mass spectrometry (ESI-MS). The proposed approach was proved to be an efficient and accurate method for specific compound isolation and identification purposes. Moreover, analytical information from the CE approach can be considered as the valuable tool for binding constant determination. The binding constant of mitraphylline with Abeta1-40 protein determined by the Hummel-Dreyer method and Scatchard analysis equals K = 9.95 x 10(5) M(-1). The results obtained showed the significant binding of the tested compound with Abeta1-40 protein. These results are discussed and interpreted in the view of developing a strategy for identification of novel compounds of great importance in Alzheimer disease therapy. PMID:17294693

Frackowiak, Teresa; Baczek, Tomasz; Roman, Kaliszana; Zbikowska, Beata; Gle?sk, Micha?; Fecka, Izabela; Cisowski, Wojciech

2006-01-01

308

Sorting by the cytoplasmic domain of the amyloid precursor protein binding receptor SorLA.  

DEFF Research Database (Denmark)

SorLA/LR11 (250 kDa) is the largest and most composite member of the Vps10p-domain receptors, a family of type 1 proteins preferentially expressed in neuronal tissue. SorLA binds several ligands, including neurotensin, platelet-derived growth factor-bb, and lipoprotein lipase, and via complex-formation with the amyloid precursor protein it downregulates generation of Alzheimer's disease-associated Abeta-peptide. The receptor is mainly located in vesicles, suggesting a function in protein sorting and transport. Here we examined SorLA's trafficking using full-length and chimeric receptors and find that its cytoplasmic tail mediates efficient Golgi body-endosome transport, as well as AP-2 complex-dependent endocytosis. Functional sorting sites were mapped to an acidic cluster-dileucine-like motif and to a GGA binding site in the C terminus. Experiments in permanently or transiently AP-1 mu1-chain-deficient cells established that the AP-1 adaptor complex is essential to SorLA's transport between Golgi membranes and endosomes. Our results further implicate the GGA proteins in SorLA trafficking and provide evidence that SNX1 and Vps35, as parts of the retromer complex or possibly in a separate context, are engaged in retraction of the receptor from endosomes. Udgivelsesdato: 2007-Oct

Nielsen, Morten S; Gustafsen, Camilla

2007-01-01

309

Serum protein signatures differentiating autoimmune pancreatitis versus pancreatic cancer.  

Science.gov (United States)

Autoimmune pancreatitis (AIP) is defined by characteristic lymphoplasmacytic infiltrate, ductal strictures and a pancreatic enlargement or mass that can mimic pancreatic cancer (PaCa). The distinction between this benign disease and pancreatic cancer can be challenging. However, an accurate diagnosis may pre-empt the misdiagnosis of cancer, allowing the appropriate medical treatment of AIP and, consequently, decreasing the number of unnecessary pancreatic resections. Mass spectrometry (MS) and two-dimensional differential gel electrophoresis (2D-DIGE) have been applied to analyse serum protein alterations associated with AIP and PaCa, and to identify protein signatures indicative of the diseases. Patients' sera were immunodepleted from the 20 most prominent serum proteins prior to further 2D-DIGE and image analysis. The identity of the most-discriminatory proteins detected, was performed by MS and ELISAs were applied to confirm their expression. Serum profiling data analysis with 2D-DIGE revealed 39 protein peaks able to discriminate between AIP and PaCa. Proteins were purified and further analysed by MALDI-TOF-MS. Peptide mass fingerprinting led to identification of eleven proteins. Among them apolipoprotein A-I, apolipoprotein A-II, transthyretin, and tetranectin were identified and found as 3.0-, 3.5-, 2-, and 1.6-fold decreased in PaCa sera, respectively, whereas haptoglobin and apolipoprotein E were found to be 3.8- and 1.6-fold elevated in PaCa sera. With the exception of haptoglobin the ELISA results of the identified proteins confirmed the 2D-DIGE image analysis characteristics. Integration of the identified serum proteins as AIP markers may have considerable potential to provide additional information for the diagnosis of AIP to choose the appropriate treatment. PMID:24349355

Felix, Klaus; Hauck, Oliver; Fritz, Stefan; Hinz, Ulf; Schnölzer, Martina; Kempf, Tore; Warnken, Uwe; Michel, Angelika; Pawlita, Michael; Werner, Jens

2013-01-01

310

Deletion of Serum Amyloid A3 Improves High Fat High Sucrose Diet-Induced Adipose Tissue Inflammation and Hyperlipidemia in Female Mice  

Science.gov (United States)

Serum amyloid A (SAA) increases in response to acute inflammatory stimuli and is modestly and chronically elevated in obesity. SAA3, an inducible form of SAA, is highly expressed in adipose tissue in obese mice where it promotes monocyte chemotaxis, providing a mechanism for the macrophage accumulation that occurs with adipose tissue expansion in obesity. Humans do not express functional SAA3 protein, but instead express SAA1 and SAA2 in hepatic as well as extrahepatic tissues, making it difficult to distinguish between liver and adipose tissue-specific SAA effects. SAA3 does not circulate in plasma, but may exert local effects that impact systemic inflammation. We tested the hypothesis that SAA3 contributes to chronic systemic inflammation and adipose tissue macrophage accumulation in obesity using mice deficient for Saa3 (Saa3?/?). Mice were rendered obese by feeding a pro-inflammatory high fat, high sucrose diet with added cholesterol (HFHSC). Both male and female Saa3?/? mice gained less weight on the HFHSC diet compared to Saa3+/+ littermate controls, with no differences in body composition or resting metabolism. Female Saa3?/? mice, but not males, had reduced HFHSC diet-induced adipose tissue inflammation and macrophage content. Both male and female Saa3?/? mice had reduced liver Saa1 and Saa2 expression in association with reduced plasma SAA. Additionally, female Saa3?/? mice, but not males, showed improved plasma cholesterol, triglycerides, and lipoprotein profiles, with no changes in glucose metabolism. Taken together, these results suggest that the absence of Saa3 attenuates liver-specific SAA (i.e., SAA1/2) secretion into plasma and blunts weight gain induced by an obesogenic diet. Furthermore, adipose tissue-specific inflammation and macrophage accumulation are attenuated in female Saa3?/? mice, suggesting a novel sexually dimorphic role for this protein. These results also suggest that Saa3 influences liver-specific SAA1/2 expression, and that SAA3 could play a larger role in the acute phase response than previously thought. PMID:25251243

den Hartigh, Laura J.; Wang, Shari; Goodspeed, Leela; Ding, Yilei; Averill, Michelle; Subramanian, Savitha; Wietecha, Tomasz; O'Brien, Kevin D.; Chait, Alan

2014-01-01

311

Dynamic behavior of small heat shock protein inhibition on amyloid fibrillization of a small peptide (SSTSAA) from RNase A  

International Nuclear Information System (INIS)

Highlights: ? Mechanism of small heat shock protein inhibition on fibril formation was studied. ? Peptide SSTSAA with modified ends was used for amyloid fibril formation. ? FRET signal was followed during the fibril formation. ? Mj HSP16.5 inhibits fibril formation when introduced in the lag phase. ? Mj HSP16.5 slows down fibril formation when introduced after the lag phase. -- Abstract: Small heat shock proteins, a class of molecular chaperones, are reported to inhibit amyloid fibril formation in vitro, while the mechanism of inhibition remains unknown. In the present study, we investigated the mechanism by which Mj HSP16.5 inhibits amyloid fibril formation of a small peptide (SSTSAA) from RNase A. A model peptide (dansyl-SSTSAA-W) was designed by introducing a pair of fluorescence resonance energy transfer (FRET) probes into the peptide, allowing for the monitoring of fibril formation by this experimental model. Mj HSP16.5 completely inhibited fibril formation of the model peptide at a molar ratio of 1:120. The dynamic process of fibril formation, revealed by FRET, circular dichroism, and electron microscopy, showed a lag phase of about 2 h followed by a fast growth period. The effect of Mj HSP16.5 on amyloid fibril formation was investigated by adding it into the incubation solution during different growth phases. Adding Mj HSP16.5 to the incubating peptide before or during the lag phase completely inhibited fibril formation. However, introducing Mj HSP16.5 after the lag phase only slowed down the fibril formation process by adhering to the already formed fibrils. These findings provide insight into the inhibitory roles of small heat shock proteins on amyloid fibril formation at the molecular level.

312

Minocycline alleviates beta-amyloid protein and tau pathology via restraining neuroinflammation induced by diabetic metabolic disorder  

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Full Text Available Zhiyou Cai,1 Yong Yan,2 Yonglong Wang2 1Department of Neurology, the Lu’an Affiliated Hospital of Anhui Medical University, Lu’an People’s Hospital, Lu’an, Anhui Province, People’s Republic of China; 2Department of Neurology, the First Affiliated Hospital of Chongqing Medical University, Chongqing Key Laboratory of Neurology, Chongqing, People’s Republic of China Background: Compelling evidence has shown that diabetic metabolic disorder plays a critical role in the pathogenesis of Alzheimer’s disease, including increased expression of ?-amyloid protein (A? and tau protein. Evidence has supported that minocycline, a tetracycline derivative, protects against neuroinflammation induced by neurodegenerative disorders or cerebral ischemia. This study has evaluated minocycline influence on expression of A? protein, tau phosphorylation, and inflammatory cytokines (interleukin-1? and tumor necrosis factor-? in the brain of diabetic rats to clarify neuroprotection by minocycline under diabetic metabolic disorder. Method: An animal model of diabetes was established by high fat diet and intraperitoneal injection of streptozocin. In this study, we investigated the effect of minocycline on expression of A? protein, tau phosphorylation, and inflammatory cytokines (interleukin-1? and tumor necrosis factor-? in the hippocampus of diabetic rats via immunohistochemistry, western blotting, and enzyme-linked immunosorbent assay. Results: These results showed that minocycline decreased expression of A? protein and lowered the phosphorylation of tau protein, and retarded the proinflammatory cytokines, but not amyloid precursor protein. Conclusion: On the basis of the finding that minocycline had no influence on amyloid precursor protein and beta-site amyloid precursor protein cleaving enzyme 1 which determines the speed of A? generation, the decreases in A? production and tau hyperphosphorylation by minocycline are through inhibiting neuroinflammation, which contributes to A? production and tau hyperphosphorylation. Minocycline may also lower the self-perpetuating cycle between neuroinflammation and the pathogenesis of tau and A? to act as a neuroprotector. Therefore, the ability of minocycline to modulate inflammatory reactions may be of great importance in the selection of neuroprotective agents, especially in chronic conditions like diabetes and Alzheimer’s disease. Keywords: diabetes mellitus, minocycline, tau protein, ?-amyloid protein

Cai Z

2013-08-01

313

Posiphen as a candidate drug to lower CSF amyloid precursor protein, amyloid-? peptide and ? levels: target engagement, tolerability and pharmacokinetics in humans  

Science.gov (United States)

Aim A first in human study to evaluate tolerability and pharmacokinetics followed by an early proof of mechanism (POM) study to determine whether the small orally, available molecule, Posiphen tartrate (Posiphen), lowers secreted (s) amyloid-? precursor protein (APP) ? and -?, amyloid-? peptide (A?), tau (?) and inflammatory markers in CSF of patients with mild cognitive impairment (MCI). Study design Posiphen single and multiple ascending dose phase 1 randomised, double blind, placebo-controlled safety, tolerance, pharmacokinetic studies were undertaken in a total of 120 healthy volunteers to define a dose that was then used in a small non-randomised study of five MCI subjects, used as their own controls, to define target engagement. Main outcome measures Pharmacodynamic: sAPP?, sAPP?, A?42, ? (total (t) and phosphorylated (p)) and inflammatory marker levels were time-dependently measured over 12?h and compared prior to and following 10?days of oral Posiphen treatment in four MCI subjects who completed the study. Pharmacokinetic: plasma and CSF drug and primary metabolite concentrations with estimated brain levels extrapolated from steady-state drug administration in rats. Results Posiphen proved well tolerated and significantly lowered CSF levels of sAPP?, sAPP?, t-?, p-? and specific inflammatory markers, and demonstrated a trend to lower CSF A?42. Conclusions These results confirm preclinical POM studies, demonstrate that pharmacologically relevant drug/metabolite levels reach brain and support the continued clinical optimisation and evaluation of Posiphen for MCI and Alzheimer's disease. PMID:22791904

Maccecchini, Maria L; Chang, Mee Young; Pan, Catherine; John, Varghese; Zetterberg, Henrik

2012-01-01

314

Rapid determination of thyroxine binding proteins of human serum  

Directory of Open Access Journals (Sweden)

Full Text Available A simple method is described for determing thyroxine binding proteins in human serum by electrophoresis at pH 8.6, using cellulose acetate membrane as the supporting medium. The procedure had high reliability in sera of normal subjects, pregnant women and patients with decreased thyroxine binding capacity of thyroxine binding globulin.

Arima,Terukatsu

1976-02-01

315

Blood serum components and serum protein test of Hybro-PG broilers of different ages  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Blood serum samples of HYBRO PG broilers were analyzed, with 30 samples collected from 21-day-old broilers (G1), 30 from 35-day-old birds (G2), and 30 from 42-day-old birds (G3), with the aim of establishing normal values of some blood serum parameters. The activities of the enzymes gamma-glutamyl-t [...] ransferase (GGT), aspartate aminotransferase (AST), creatine kinase (CK), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH), serum levels of total calcium, calcium ion, phosphorus, sodium, potassium, magnesium, chlorides, creatinine, uric acid, triglycerides, cholesterol, total protein, albumin, total and indirect and direct bilirubin, and electrophoretic profile of serum proteins in acrylamide (SDS-PAGE) and agarose gel were determined. There was no influence of age on total bilirubin and albumin levels. All the other evaluated parameters presented differences in at least one age group. Protein electrophoretic profile also changed as a function of age. The obtained results can be considered as normal for the studied ages, and therefore be used as references for the interpretation of laboratory exams of broilers of this genetic line in the evaluated ages.

PRL, Silva; OC, Freitas Neto; AC, Laurentiz; OM, Junqueira; JJ, Fagliari.

2007-12-01

316

Deletion of an endosomal/lysosomal targeting signal promotes the secretion of Alzheimer's disease amyloid precursor protein (APP).  

Science.gov (United States)

Alzheimer's disease amyloid precursor protein (APP) generates a beta-amyloid protein (A beta) that is a main component of the senile plaques found in the brains of Alzheimer's disease patients. APP is thought to undergo proteolysis via two different pathways, the amyloidogenic pathway which produces A beta, and the non-amyloidogenic pathway which releases a large N-terminal fragment into the medium. The proteases that mediate these processes remain unidentified. The physiological function of APP is not clear yet. Therefore, the cytoplasmic region of APP has attracted much interest, because this region is highly conserved among species, and members of the amyloid precursor-like protein (APLP) family. Several potentially functional sequences exist in the region, including signal sequences for protein sorting and a G0-protein binding sequence. We constructed two mutants, 695 deltaNPTY and 695 deltaGYEN. They lack potential endosome/lysosome targeting signals, NPTY and GY, in the cytoplasmic domain of APP695, respectively. The mutant APPs had longer half-lives and were secreted more easily into the medium than the wild type, suggesting that these sequences are important for the secretion and metabolism of APP. PMID:9133629

Ono, Y; Kinouchi, T; Sorimachi, H; Ishiura, S; Suzuki, K

1997-03-01

317

Tau/Amyloid Beta 42 Peptide Test (Alzheimer Biomarkers)  

Science.gov (United States)

... Formal name: Tau Protein and Amyloid Beta 42 Peptide Related tests: Phosporylated Tau (P-tau), APOE Genotyping , ... cerebrospinal fluid (CSF) . Amyloid beta 42 is a peptide (protein fragment). Increased production of amyloid beta 42 ...

318

Mechanism of cytotoxicity mediated by the C31 fragment of the amyloid precursor protein.  

Science.gov (United States)

The cytoplasmic tail of the amyloid precursor protein (APP) contains two putatively cytotoxic peptides, Jcasp and C31, derived by caspase cleavage of APP. Jcasp is a fragment starting from the epsilon-secretase site to position 664, while C31 is a fragment from position 665 to the C-terminus. Our studies now showed that compared to C31, Jcasp appeared to play a minor role in cytotoxicity. In particular, inhibition of Jcasp generation by treatment of gamma-secretase inhibitor did not lead to any attenuation of C31-induced toxicity. Secondly, because C31 toxicity is largely absent in cells lacking endogenous APP, we determined, using a split beta-galactosidase complementary assay to monitor protein-protein interactions, the presence of APP associated complexes. Our results demonstrated that both APP homomeric and C31/APP heteromeric complexes were correlated with cell death, indicating that C31 complexes with APP to recruit the interacting partners that initiate the signals related to cellular toxicity. PMID:19679105

Park, Sun Ah; Shaked, Gideon M; Bredesen, Dale E; Koo, Edward H

2009-10-16

319

Significant association between renal function and amyloid-positive area in renal biopsy specimens in AL amyloidosis  

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Full Text Available Abstract Background The kidney is a major target organ for systemic amyloidosis that often affects the kidney including proteinura, and elevated serum creatinine (Cr. The correlation between amount of amyloid deposits and clinical parameters is not known. The aim of this study was to clarify correlation the amyloid area in all renal biopsy specimen and clinical parameters. Methods Fifty-eight patients with an established diagnosis of AL amyloidosis participated in the study. All patients showed amyloid deposits in renal biopsies. We retrospectively investigated the correlation between clinical data and amyloid occupied area in whole renal biopsy specimens. Results The area occupied by amyloid was less than 10% in 57 of the 58 patients, and was under 2% in 40. For statistical analyses, %amyloid-positive areas were transformed to common logarithmic values (Log10%amyloid. Cr showed significant correlation with Log10%amyloid and estimated glomerular filtration rate (eGFR showed the significant negative correlation. Patient age, cleatinine clearance (Ccr, blood urea nitorogen, and urinary protein was not significantly correlated with Log10%amyloid. The correlation with other clinical factors such as sex, and serum concentrations of total protein, albumin, immunoglobulins, compliments was evaluated. None of these factors significantly correlated with Log10%amyloid. According to sex- and age- adjusted multiple linear regression analysis, Log10%amyloid had significant positive association with Cr and significant negative association with eGFR. Conclusion There is significant association between amyloid-positive area in renal tissue and renal function, especially Cr and eGFR. The level of Cr and eGFR may be a marker of amount of amyloid in renal tissue.

Kuroda Takeshi

2012-09-01

320

Identification of novel small molecule inhibitors of amyloid precursor protein synthesis as a route to lower Alzheimer's disease amyloid-beta peptide.  

Science.gov (United States)

A wealth of independent research with transgenic mice, antibodies, and vaccines has pointed to a causative role of the amyloid-beta peptide (A beta) in Alzheimer's disease (AD). Based on these and earlier associative studies, A beta represents a promising target for development of therapeutics focused on AD disease progression. Interestingly, a cholinesterase inhibitor currently in clinical trials, phenserine, has been shown to inhibit production of both amyloid precursor protein (APP) and A beta. We have shown that this inhibition occurs at the post-transcriptional level with a specific blocking of the synthesis of APP relative to total protein synthesis (Shaw et al., 2001). However, the dose of phenserine necessary to block APP production is far higher than that needed to elicit its anticholinesterase activity, and it is these latter actions that are dose limiting in vivo. The focus of this study was to screen 144 analogs of phenserine to identify additional small molecules that inhibit APP protein synthesis, and thereby A beta production, without possessing potent acetylcholinesterase (AChE) inhibitory activity. An enzyme-linked immunosorbent assay was used to identify analogs capable of suppressing APP production following treatment of human neuroblastoma cells with 20 muM of compound. Eight analogs were capable of dose dependently reducing APP and A beta production without causing cell toxicity in further studies. Several of these analogs had little to no AChE activities. Translation of APP and A beta actions to mice was demonstrated with one agent. They thus represent interesting lead molecules for assessment in animal models, to define their tolerance and utility as potential AD therapeutics. PMID:16690718

Utsuki, Tada; Yu, Qian-Sheng; Davidson, Diane; Chen, Demao; Holloway, Harold W; Brossi, Arnold; Sambamurti, Kumar; Lahiri, Debomoy K; Greig, Nigel H; Giordano, Tony

2006-08-01

 
 
 
 
321

Amyloid-like Fibrils from a Domain-swapping Protein Feature a Parallel, in-Register Conformation without Native-like Interactions*  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The formation of amyloid-like fibrils is characteristic of various diseases, but the underlying mechanism and the factors that determine whether, when, and how proteins form amyloid, remain uncertain. Certain mechanisms have been proposed based on the three-dimensional or runaway domain swapping, inspired by the fact that some proteins show an apparent correlation between the ability to form domain-swapped dimers and a tendency to form fibrillar aggregates. Intramolecular ?-sheet contacts pr...

Li, Jun; Hoop, Cody L.; Kodali, Ravindra; Sivanandam, V. N.; Wel, Patrick C. A.

2011-01-01

322

Comparative transcriptome profiling of amyloid precursor protein family members in the adult cortex  

Science.gov (United States)

Background The ?-amyloid precursor protein (APP) and the related ?-amyloid precursor-like proteins (APLPs) undergo complex proteolytic processing giving rise to several fragments. Whereas it is well established that A? accumulation is a central trigger for Alzheimer's disease, the physiological role of APP family members and their diverse proteolytic products is still largely unknown. The secreted APPs? ectodomain has been shown to be involved in neuroprotection and synaptic plasticity. The ?-secretase-generated APP intracellular domain (AICD) functions as a transcriptional regulator in heterologous reporter assays although its role for endogenous gene regulation has remained controversial. Results To gain further insight into the molecular changes associated with knockout phenotypes and to elucidate the physiological functions of APP family members including their proposed role as transcriptional regulators, we performed DNA microarray transcriptome profiling of prefrontal cortex of adult wild-type (WT), APP knockout (APP-/-), APLP2 knockout (APLP2-/-) and APPs? knockin mice (APP?/?) expressing solely the secreted APPs? ectodomain. Biological pathways affected by the lack of APP family members included neurogenesis, transcription, and kinase activity. Comparative analysis of transcriptome changes between mutant and wild-type mice, followed by qPCR validation, identified co-regulated gene sets. Interestingly, these included heat shock proteins and plasticity-related genes that were both down-regulated in knockout cortices. In contrast, we failed to detect significant differences in expression of previously proposed AICD target genes including Bace1, Kai1, Gsk3b, p53, Tip60, and Vglut2. Only Egfr was slightly up-regulated in APLP2-/- mice. Comparison of APP-/- and APP?/? with wild-type mice revealed a high proportion of co-regulated genes indicating an important role of the C-terminus for cellular signaling. Finally, comparison of APLP2-/- on different genetic backgrounds revealed that background-related transcriptome changes may dominate over changes due to the knockout of a single gene. Conclusion Shared transcriptome profiles corroborated closely related physiological functions of APP family members in the adult central nervous system. As expression of proposed AICD target genes was not altered in adult cortex, this may indicate that these genes are not affected by lack of APP under resting conditions or only in a small subset of cells. PMID:21435241

2011-01-01

323

Comparative transcriptome profiling of amyloid precursor protein family members in the adult cortex  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The ?-amyloid precursor protein (APP and the related ?-amyloid precursor-like proteins (APLPs undergo complex proteolytic processing giving rise to several fragments. Whereas it is well established that A? accumulation is a central trigger for Alzheimer's disease, the physiological role of APP family members and their diverse proteolytic products is still largely unknown. The secreted APPs? ectodomain has been shown to be involved in neuroprotection and synaptic plasticity. The ?-secretase-generated APP intracellular domain (AICD functions as a transcriptional regulator in heterologous reporter assays although its role for endogenous gene regulation has remained controversial. Results To gain further insight into the molecular changes associated with knockout phenotypes and to elucidate the physiological functions of APP family members including their proposed role as transcriptional regulators, we performed DNA microarray transcriptome profiling of prefrontal cortex of adult wild-type (WT, APP knockout (APP-/-, APLP2 knockout (APLP2-/- and APPs? knockin mice (APP?/? expressing solely the secreted APPs? ectodomain. Biological pathways affected by the lack of APP family members included neurogenesis, transcription, and kinase activity. Comparative analysis of transcriptome changes between mutant and wild-type mice, followed by qPCR validation, identified co-regulated gene sets. Interestingly, these included heat shock proteins and plasticity-related genes that were both down-regulated in knockout cortices. In contrast, we failed to detect significant differences in expression of previously proposed AICD target genes including Bace1, Kai1, Gsk3b, p53, Tip60, and Vglut2. Only Egfr was slightly up-regulated in APLP2-/- mice. Comparison of APP-/- and APP?/? with wild-type mice revealed a high proportion of co-regulated genes indicating an important role of the C-terminus for cellular signaling. Finally, comparison of APLP2-/- on different genetic backgrounds revealed that background-related transcriptome changes may dominate over changes due to the knockout of a single gene. Conclusion Shared transcriptome profiles corroborated closely related physiological functions of APP family members in the adult central nervous system. As expression of proposed AICD target genes was not altered in adult cortex, this may indicate that these genes are not affected by lack of APP under resting conditions or only in a small subset of cells.

Eils Roland

2011-03-01

324

Interactions of lecithinized superoxide dismutase with serum proteins and cells.  

Science.gov (United States)

Superoxide dismutase covalently bound to four lecithin molecules (PC-SOD) is known to be retained in circulating blood for a prolonged period and has a high affinity for cells, resulting in beneficial therapeutic effects in animal disease models. In this study, we evaluated the interaction of PC-SOD with biological components, such as serum proteins and cells, to clarify the mechanism underlying the improved pharmacokinetics of SOD induced by lecithin chemical modification (lecithinization). PC-SOD was distributed in the plasma but not in blood cells after being added to the blood. PC-SOD formed a complex with serum protein(s) such as albumin, whereas unmodified SOD did not. The cellular content of PC-SOD was markedly higher than that of unmodified SOD, and was distributed in lysosomes. The pathway associated with the cellular uptake was found to involve clathrin-/caveolae-independent and cholesterol-sensitive endocytosis. Overall, our data indicated that the increased hydrophobicity of lecithinized SOD enhanced its association to both serum protein(s) and plasma membrane microdomains. The former inhibited SOD excretion and promoted long-term retention in circulating blood, whereas the latter enhanced internalization into cells via endocytosis. PMID:24867597

Ishihara, Tsutomu; Nara, Shunsuke; Mizushima, Tohru

2014-07-01

325

Serum proteins, trace metals and phosphatases in psoriasis  

Directory of Open Access Journals (Sweden)

Full Text Available Serum proteins, zinc, copper, acid phosphatase (AcPase and alkaline phosphatase (AlPase were studied in both active and remission phases of psoriasis. Data were compared with healthy controls, ?1, ? and ? globulins were high in active phase while ?1 and ? globulins were at par in remission phase. Serum copper was low but zinc and alkaline phosphatase were significantly high in both active and remission phases of the disease. Acid phosphatase level was at par in all the experimental groups. Study suggests a positive correlation of globulin, zinc and Alpase in active and remission phase of psoriasis.

Bhatnagar M

1994-01-01

326

Effects of Methylthiouracil on the Serum Protein Fractions in Dog  

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Full Text Available The effects of a prolonged oral administration of methylthiouracil was studied on the electrophoretically - separated serum protein fractions in 24 dogs. Though the statistical mean of changes in serum albumin and alpha2 globulin were highly significant but the individual alterations were not consistent and therefore on the basis of the available data there does not seem to be a characteristic electrophoretic pattern in the experimentally produced hypothyroidism in the dog. This finding is in agreement with the clinical electrophoretic pattern reported in the human spontaneous hypothyroidism.

A- Pousti N- Guiti

1970-07-01

327

Phosphorylation of Alzheimer disease amyloid precursor peptide by protein kinase C and Ca2+/calmodulin-dependent protein kinase II  

International Nuclear Information System (INIS)

The amino acid sequence of the Alzheimer disease amyloid precursor (ADAP) has been deduced from the corresponding cDNA, and hydropathy analysis of the sequence suggest a receptor-like structure with a single transmembrane domain. The putative cytoplasmic domain of ADAP contains potential sites for serine and threonine phosphorylation. In the present study, synthetic peptides derived from this domain were used as model substrates for various purified protein kinases. Protein kinase C rapidly catalyzed the phosphorylation of a peptide corresponding to amino acid residues 645-661 of ADAP. Ca2+/calmodulin-dependent protein kinase II phosphorylated ADAP peptide (645-661) on Thr-654 and Ser-655. Using rat cerebral cortex synaptosomes prelabeled with 32Pi, a 32P-labeled phosphoprotein of ?135 kDa was immunoprecipitated by using antisera prepared against ADAP peptide(597-624), consistent with the possibility that the holoform of ADAP in rat brain is a phosphoprotein. Based on analogy with the effect of phosphorylation by protein kinase C of juxtamembrane residues in the cytoplasmic domain of the epidermal growth factor receptor and the interleukin 2 receptor, phosphorylation of ADAP may target it for internalization

328

Acute serum amyloid A induces migration, angiogenesis, and inflammation in synovial cells in vitro and in a human rheumatoid arthritis/SCID mouse chimera model.  

LENUS (Irish Health Repository)

Serum amyloid A (A-SAA), an acute-phase protein with cytokine-like properties, is expressed at sites of inflammation. This study investigated the effects of A-SAA on chemokine-regulated migration and angiogenesis using rheumatoid arthritis (RA) cells and whole-tissue explants in vitro, ex vivo, and in vivo. A-SAA levels were measured by real-time PCR and ELISA. IL-8 and MCP-1 expression was examined in RA synovial fibroblasts, human microvascular endothelial cells, and RA synovial explants by ELISA. Neutrophil transendothelial cell migration, cell adhesion, invasion, and migration were examined using transwell leukocyte\\/monocyte migration assays, invasion assays, and adhesion assays with or without anti-MCP-1\\/anti-IL-8. NF-kappaB was examined using a specific inhibitor and Western blotting. An RA synovial\\/SCID mouse chimera model was used to examine the effects of A-SAA on cell migration, proliferation, and angiogenesis in vivo. High expression of A-SAA was demonstrated in RA patients (p < 0.05). A-SAA induced chemokine expression in a time- and dose-dependent manner (p < 0.05). Blockade with anti-scavenger receptor class B member 1 and lipoxin A4 (A-SAA receptors) significantly reduced chemokine expression in RA synovial tissue explants (p < 0.05). A-SAA induced cell invasion, neutrophil-transendothelial cell migration, monocyte migration, and adhesion (all p < 0.05), effects that were blocked by anti-IL-8 or anti-MCP-1. A-SAA-induced chemokine expression was mediated through NF-kappaB in RA explants (p < 0.05). Finally, in the RA synovial\\/SCID mouse chimera model, we demonstrated for the first time in vivo that A-SAA directly induces monocyte migration from the murine circulation into RA synovial grafts, synovial cell proliferation, and angiogenesis (p < 0.05). A-SAA promotes cell migrational mechanisms and angiogenesis critical to RA pathogenesis.

Connolly, Mary

2010-06-01

329

Investigation of copper(II) binding to the protein precursor of Non-Amyloid-Beta Component of Alzheimer Disease Amyloid Plaque  

Science.gov (United States)

The Non-Amyloid-Beta Component Precursor (NACP) is a natively unfolded synaptic protein that is implicated in Alzheimers and Parkinsons diseases. Its aggregation into fibrillar structures is accelerated by the binding of copper(II). Experimental studies suggest that the dominant copper binding site is located at the histidine residue in NACP. Based on this evidence we assembled a model fragment of the binding site and used DFT to analyze the conformational details of the most probable binding motifs. We investigated the overall conformational effects with classical MD by constraining the copper binding site to the most energetically favorable geometry obtained from the DFT calculations. These results are compared and contrasted with those of the unbound NACP.

Rose, Francis; Hodak, Miroslav; Bernholc, Jerry

2007-03-01

330

Immunohistochemical characterization of alterations in the distribution of amyloid precursor proteins and beta-amyloid peptide after experimental brain injury in the rat.  

Science.gov (United States)

Recent reports suggest a relationship between traumatic brain injury and the precocious development of neurodegenerative cascades, including diffuse deposits of beta-amyloid peptides (A beta) in the injured brain. Because the lateral fluid-percussion (FP) model of experimental brain injury produces clinically relevant neuropathological sequelae in the rat brain, we used this model together with a series of antibodies specific for amyloid precursor proteins (APPs), APP-like proteins (APLPs), or A beta to identify acute neurodegenerative changes after brain trauma. Male Sprague-Dawley rats were anesthetized and subjected to lateral FP brain injury of moderate to high severity. At 1 hr, 2 hr, 48 hr, 1 week, or 2 weeks after injury, animals were killed and their brains were removed for immunohistochemical analysis. APP/APLP immunoreactivity increased in specific brain regions as early as 1 hr after injury and persisted for at least 2 weeks. Axons in the thalamus and subcortical white matter showed the greatest APP/APLP accumulation. Injured cortex, striatum, cingulum, and hippocampus also demonstrated significant axonal accumulations of APP/APLP. Accumulation of APP/APLPs occurred primarily ipsilateral to the injury, although bilateral changes were observed in some brain regions. No deposition of A beta was observed in any brain region at any time point examined. These results demonstrate a pattern of widespread axonal pathology after lateral FP brain injury in the rat, characterized by intra-axonal accumulations of APP/APLP immunoreactivity in the absence of plaque-like deposits of A beta in the traumatized brain. PMID:8558237

Pierce, J E; Trojanowski, J Q; Graham, D I; Smith, D H; McIntosh, T K

1996-02-01

331

Characterizing the location and trafficking routes of the neuronal retromer and its role in amyloid precursor protein transport  

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The retromer complex plays an important role in intracellular transport, is highly expressed in the hippocampus, and has been implicated in the trafficking of the amyloid precursor protein (APP). Nevertheless, the trafficking routes of the neuronal retromer and the role it plays in APP transport in neuronal processes remains unknown. Here we use hippocampal neuronal cultures to address these issues. Using fluorescence microscopy, we find that Vps35, the core element of the retromer complex, i...

Bhalla, Akhil; Vetanovetz, Christopher P.; Morel, Etienne; Chamoun, Zeina; Paolo, Gilbert Di; Small, Scott A.

2012-01-01

332

Amyloid ?-Protein Is Degraded by Cellular Angiotensin-converting Enzyme (ACE) and Elevated by an ACE Inhibitor*  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Human genetic data have associated angiotensin-converting enzyme (ACE) with Alzheimer disease (AD), and purified ACE has been reported to cleave synthetic amyloid ?-protein (A?) in vitro. Whether deficiency in ACE activity, arising from genetic alteration or pharmacological inhibition, can decrease A? degradation and allow A? accumulation in intact cells is unknown. We cloned ACE from human neuroblastoma cells and showed that it had posttranslational processing and enzymatic activity typi...

Hemming, Matthew L.; Selkoe, Dennis J.

2005-01-01

333

ADAM9 Inhibition Increases Membrane Activity of ADAM10 and Controls ?-Secretase Processing of Amyloid Precursor Protein*  

Science.gov (United States)

Prodomains of A disintegrin and metalloproteinase (ADAM) metallopeptidases can act as highly specific intra- and intermolecular inhibitors of ADAM catalytic activity. The mouse ADAM9 prodomain (proA9; amino acids 24–204), expressed and characterized from Escherichia coli, is a competitive inhibitor of human ADAM9 catalytic/disintegrin domain with an overall inhibition constant of 280 ± 34 nm and high specificity toward ADAM9. In SY5Y neuroblastoma cells overexpressing amyloid precursor protein, proA9 treatment reduces the amount of endogenous ADAM10 enzyme in the medium while increasing membrane-bound ADAM10, as shown both by Western and activity assays with selective fluorescent peptide substrates using proteolytic activity matrix analysis. An increase in membrane-bound ADAM10 generates higher levels of soluble amyloid precursor protein ? in the medium, whereas soluble amyloid precursor protein ? levels are decreased, demonstrating that inhibition of ADAM9 increases ?-secretase activity on the cell membrane. Quantification of physiological ADAM10 substrates by a proteomic approach revealed that substrates, such as epidermal growth factor (EGF), HER2, osteoactivin, and CD40-ligand, are increased in the medium of BT474 breast tumor cells that were incubated with proA9, demonstrating that the regulation of ADAM10 by ADAM9 applies for many ADAM10 substrates. Taken together, our results demonstrate that ADAM10 activity is regulated by inhibition of ADAM9, and this regulation may be used to control shedding of amyloid precursor protein by enhancing ?-secretase activity, a key regulatory step in the etiology of Alzheimer disease. PMID:21956108

Moss, Marcia L.; Powell, Gary; Miller, Miles A.; Edwards, Lori; Qi, Bin; Sang, Qing-Xiang Amy; De Strooper, Bart; Tesseur, Ina; Lichtenthaler, Stefan F.; Taverna, Mara; Zhong, Julia Li; Dingwall, Colin; Ferdous, Taheera; Schlomann, Uwe; Zhou, Pei; Griffith, Linda G.; Lauffenburger, Douglas A.; Petrovich, Robert; Bartsch, Jorg W.

2011-01-01

334

No Evidence for a Role of Adipose Tissue-Derived Serum Amyloid A in the Development of Insulin Resistance or Obesity-Related Inflammation in hSAA1+/? Transgenic Mice  

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Obesity is associated with a low-grade inflammation including moderately increased serum levels of the acute phase protein serum amyloid A (SAA). In obesity, SAA is mainly produced from adipose tissue and serum levels of SAA are associated with insulin resistance. SAA has been described as a chemoattractant for inflammatory cells and adipose tissue from obese individuals contains increased numbers of macrophages. However, whether adipose tissue-derived SAA can have a direct impact on macrophage infiltration in adipose tissue or the development of insulin resistance is unknown. The aim of this study was to investigate the effects of adipose tissue-derived SAA1 on the development of insulin resistance and obesity-related inflammation. We have previously established a transgenic mouse model expressing human SAA1 in the adipose tissue. For this report, hSAA1+/? transgenic mice and wild type mice were fed with a high fat diet or normal chow. Effects of hSAA1 on glucose metabolism were assessed using an oral glucose tolerance test. Real-time PCR was used to measure the mRNA levels of macrophage markers and genes related to insulin sensitivity in adipose tissue. Cytokines during inflammation were analyzed using a Proinflammatory 7-plex Assay. We found similar insulin and glucose levels in hSAA1 mice and wt controls during an oral glucose tolerance test and no decrease in mRNA levels of genes related to insulin sensitivity in adipose tissue in neither male nor female hSAA1 animals. Furthermore, serum levels of proinflammatory cytokines and mRNA levels of macrophage markers in adipose tissue were not increased in hSAA1 mice. Hence, in this model we find no evidence that adipose tissue-derived hSAA1 influences the development of insulin resistance or obesity-related inflammation. PMID:23967285

Ahlin, Sofie; Olsson, Maja; Olsson, Bob; Svensson, Per-Arne; Sjoholm, Kajsa

2013-01-01

335

No evidence for a role of adipose tissue-derived serum amyloid a in the development of insulin resistance or obesity-related inflammation in hSAA1(+/-) transgenic mice.  

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Obesity is associated with a low-grade inflammation including moderately increased serum levels of the acute phase protein serum amyloid A (SAA). In obesity, SAA is mainly produced from adipose tissue and serum levels of SAA are associated with insulin resistance. SAA has been described as a chemoattractant for inflammatory cells and adipose tissue from obese individuals contains increased numbers of macrophages. However, whether adipose tissue-derived SAA can have a direct impact on macrophage infiltration in adipose tissue or the development of insulin resistance is unknown. The aim of this study was to investigate the effects of adipose tissue-derived SAA1 on the development of insulin resistance and obesity-related inflammation. We have previously established a transgenic mouse model expressing human SAA1 in the adipose tissue. For this report, hSAA1(+/-) transgenic mice and wild type mice were fed with a high fat diet or normal chow. Effects of hSAA1 on glucose metabolism were assessed using an oral glucose tolerance test. Real-time PCR was used to measure the mRNA levels of macrophage markers and genes related to insulin sensitivity in adipose tissue. Cytokines during inflammation were analyzed using a Proinflammatory 7-plex Assay. We found similar insulin and glucose levels in hSAA1 mice and wt controls during an oral glucose tolerance test and no decrease in mRNA levels of genes related to insulin sensitivity in adipose tissue in neither male nor female hSAA1 animals. Furthermore, serum levels of proinflammatory cytokines and mRNA levels of macrophage markers in adipose tissue were not increased in hSAA1 mice. Hence, in this model we find no evidence that adipose tissue-derived hSAA1 influences the development of insulin resistance or obesity-related inflammation. PMID:23967285

Ahlin, Sofie; Olsson, Maja; Olsson, Bob; Svensson, Per-Arne; Sjöholm, Kajsa

2013-01-01

336

Stereoselective serum protein binding of ketoprofen in liver diseases.  

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The stereoselective binding of ketoprofen (KP) to human serum was studied in patients with liver diseases and was compared with that of normal volunteers. The serum protein binding of racemic KP was found to be decreased in hepatic patients whereas, interestingly, the stereoselectivity of KP was increased. The ratio of unbound concentrations of the KP enantiomers (FR/FS) showed positive linear correlation with albumin concentrations. Inhibition of KP binding to human serum albumin (HSA) induced by lithocholate, lithocholate sulfate and bilirubin was studied. The data presented in this paper strongly suggest that the variation of stereoselective binding of KP in patients suffering from liver diseases was mainly caused by the decrease of HSA levels and secondly by the stereoselective inhibition induced by the increased concentrations of bile acid and its metabolite. PMID:10586515

Sakai, T; Maruyama, T; Sako, T; Ahmed, S; Zuidema, X; Fujiyama, S; Otagiri, M

1999-01-01

337

Protein ?-mediated effects on rat hippocampal choline transporters CHT1 and ?-amyloid ? interactions.  

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It is suggested that intracellular tau protein (?), when released extracellularly upon neuron degeneration, could evoke direct toxic effects on the cholinergic neurotransmitter system through muscarinic receptors and thus contribute to the pathogenesis of Alzheimer's disease. In this study, we evaluated the in vitro effects of six naturally occurring monomeric ? isoforms on rat hippocampal synaptosomal choline transporters CHT1 (large transmembrane proteins associated with high-affinity choline transport and vulnerable to actions of amyloid ? peptides (A?) applied in vitro or in vivo). Some ? isoforms at nM concentrations inhibited choline transport in a dose- and time-dependent saturable manner (352 = 441 > 410 = 383 > 381 = 412) and effects were associated with changes in the Michaelis constant rather than in maximal velocity. Moreover, the actions of ? 352/441 were not influenced by previous depolarisation of synaptosomes or by previous depletion of membrane cholesterol. Specific binding of [3H]hemicholinium-3 was not significantly altered by ? 352/441 at higher nM concentrations. Results of in vitro tests on CHT1 transporters from cholesterol-depleted synaptosomes supported interactions between A? 1-40 and ? 352. In addition, we developed surface plasmon resonance biosensors to monitor complexes of A? 1-42 and ? 352 using a sandwich detection format. It seems, therefore, that protein ?, similar to A? peptides, can contribute to the pathogenesis of Alzheimer's disease through its actions on CHT1 transporters. However, the interaction mechanisms are quite different (? probably exerts its effects through direct interactions of microtubule binding repeats with extracellular portions of the CHT1 protein without influencing the choline recognition site, A? rather through lipid rafts in the surrounding membranes). An N-terminal insert of ? is not necessary but the N-terminal projection domain plays a role. The developed biosensor will be used to detect A?-? complexes in cerebrospinal fluid in order to evaluate them as prospective biomarkers of Alzheimer's disease. PMID:23824558

Kristofikova, Zdena; Ripova, Daniela; Hegnerová, Katerina; Sirova, Jana; Homola, Jiri

2013-09-01

338

Turnover of Amyloid Precursor Protein Family Members Determines Their Nuclear Signaling Capability  

Science.gov (United States)

The amyloid precursor protein (APP) as well as its homologues, APP-like protein 1 and 2 (APLP1 and APLP2), are cleaved by ?-, ?-, and ?-secretases, resulting in the release of their intracellular domains (ICDs). We have shown that the APP intracellular domain (AICD) is transported to the nucleus by Fe65 where they jointly bind the histone acetyltransferase Tip60 and localize to spherical nuclear complexes (AFT complexes), which are thought to be sites of transcription. We have now analyzed the subcellular localization and turnover of the APP family members. Similarly to AICD, the ICD of APLP2 localizes to spherical nuclear complexes together with Fe65 and Tip60. In contrast, the ICD of APLP1, despite binding to Fe65, does not translocate to the nucleus. In addition, APLP1 predominantly localizes to the plasma membrane, whereas APP and APLP2 are detected in vesicular structures. APLP1 also demonstrates a much slower turnover of the full-length protein compared to APP and APLP2. We further show that the ICDs of all APP family members are degraded by the proteasome and that the N-terminal amino acids of ICDs determine ICD degradation rate. Together, our results suggest that different nuclear signaling capabilities of APP family members are due to different rates of full-length protein processing and ICD proteasomal degradation. Our results provide evidence in support of a common nuclear signaling function for APP and APLP2 that is absent in APLP1, but suggest that APLP1 has a regulatory role in the nuclear translocation of APP family ICDs due to the sequestration of Fe65. PMID:23874953

Trutzel, Annette; Bundschuh, Diana; Nitsch, Roger M.; Konietzko, Uwe

2013-01-01

339

Turnover of amyloid precursor protein family members determines their nuclear signaling capability.  

Science.gov (United States)

The amyloid precursor protein (APP) as well as its homologues, APP-like protein 1 and 2 (APLP1 and APLP2), are cleaved by ?-, ?-, and ?-secretases, resulting in the release of their intracellular domains (ICDs). We have shown that the APP intracellular domain (AICD) is transported to the nucleus by Fe65 where they jointly bind the histone acetyltransferase Tip60 and localize to spherical nuclear complexes (AFT complexes), which are thought to be sites of transcription. We have now analyzed the subcellular localization and turnover of the APP family members. Similarly to AICD, the ICD of APLP2 localizes to spherical nuclear complexes together with Fe65 and Tip60. In contrast, the ICD of APLP1, despite binding to Fe65, does not translocate to the nucleus. In addition, APLP1 predominantly localizes to the plasma membrane, whereas APP and APLP2 are detected in vesicular structures. APLP1 also demonstrates a much slower turnover of the full-length protein compared to APP and APLP2. We further show that the ICDs of all APP family members are degraded by the proteasome and that the N-terminal amino acids of ICDs determine ICD degradation rate. Together, our results suggest that different nuclear signaling capabilities of APP family members are due to different rates of full-length protein processing and ICD proteasomal degradation. Our results provide evidence in support of a common nuclear signaling function for APP and APLP2 that is absent in APLP1, but suggest that APLP1 has a regulatory role in the nuclear translocation of APP family ICDs due to the sequestration of Fe65. PMID:23874953

Gersbacher, Manuel T; Goodger, Zoë V; Trutzel, Annette; Bundschuh, Diana; Nitsch, Roger M; Konietzko, Uwe

2013-01-01

340

Duplication of amyloid precursor protein (APP), but not prion protein (PRNP) gene is a significant cause of early onset dementia in a large UK series.  

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Amyloid precursor protein gene (APP) duplications have been identified in screens of selected probands with early onset familial Alzheimer's disease (FAD). A causal role for copy number variation (CNV) in the prion protein gene (PRNP) in prion dementias is not known. We aimed to determine the prevalence of copy number variation in APP and PRNP in a large referral series, test a screening method for detection of the same, and expand knowledge of clinical phenotype. We used a 3-tiered screening...

2012-01-01

 
 
 
 
341

Identification and expression of the first nonmammalian amyloid-ß precursor-like protein APLP2 in the amphibian Xenopus laevis  

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The Alzheimer's disease-linked amyloid- precursor protein (APP) belongs to a superfamily of proteins, which also comprises the amyloid- precursor-like proteins, APLP1 and APLP2. Whereas APP has been identified in both lower and higher vertebrates, thus far, APLP1 and 2 have been characterized only in human and rodents. Here we identify the first nonmammalian APLP2 protein in the South African claw-toed frog Xenopus laevis. The identity between the Xenopus and mammalian APLP2 proteins is 75°...

Collin, R. W. J.; Strien, D.; Leunissen, J. A. M.; Martens, G. J. M.

2004-01-01

342

Distinctive serum protein profiles involving abundant proteins in lung cancer patients based upon antibody microarray analysis  

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Full Text Available Abstract Background Cancer serum protein profiling by mass spectrometry has uncovered mass profiles that are potentially diagnostic for several common types of cancer. However, direct mass spectrometric profiling has a limited dynamic range and difficulties in providing the identification of the distinctive proteins. We hypothesized that distinctive profiles may result from the differential expression of relatively abundant serum proteins associated with the host response. Methods Eighty-four antibodies, targeting a wide range of serum proteins, were spotted onto nitrocellulose-coated microscope slides. The abundances of the corresponding proteins were measured in 80 serum samples, from 24 newly diagnosed subjects with lung cancer, 24 healthy controls, and 32 subjects with chronic obstructive pulmonary disease (COPD. Two-color rolling-circle amplification was used to measure protein abundance. Results Seven of the 84 antibodies gave a significant difference (p Conclusion Our results suggest that a distinctive serum protein profile involving abundant proteins may be observed in lung cancer patients relative to healthy subjects or patients with chronic disease and may have utility as part of strategies for detecting lung cancer.

Rom William N

2005-08-01

343

Expression of the amyloid protein precursor of Alzheimer's disease in the developing rat olfactory system.  

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The expression of the amyloid protein precursor (APP) of Alzheimer's disease (AD) was examined in the olfactory system of the developing rat. Two monoclonal antibodies were used to detect APP: Alz-90, which specifically recognizes APP, and 22C11 which recognizes both APP and the structurally related protein APLP-2. Very similar patterns of immunoreactivity were observed with both antibodies. APP immunoreactivity was first detected in a subpopulation of olfactory epithelial cells at embryonic day 16 (E16), at a time when primary sensory olfactory axons are first beginning to pierce the glia limitans of the olfactory bulb. At E16, there were more olfactory receptor neurons which expressed APP than the olfactory marker protein (OMP), indicating that some APP-containing neurons were not fully mature. Between E16 and postnatal day 8 (P8), there was a marked increase in the number of primary sensory olfactory neurons expressing APP. In the olfactory bulb, APP was first detected in the mitral cell layer at E18, at a time when synapses are first beginning to form between the dendrites of these cells and primary sensory axons. The level of APP detected within mitral cell perikarya decreased after birth and could no longer be detected between P3 and P8. This indicated that once synaptic connections had been initiated within olfactory glomeruli, the expression of APP within the mitral cells was down-regulated. High levels of APP were, however, detected within the olfactory nerve fiber layer and glomeruli between P3 and P8. The results demonstrate that APP expression in the olfactory system is coordinately regulated with the major periods of synaptogenesis.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7493410

Clarris, H J; Key, B; Beyreuther, K; Masters, C L; Small, D H

1995-08-28

344

Curli functional amyloid systems are phylogenetically widespread and display large diversity in operon and protein structure.  

Science.gov (United States)

Escherichia coli and a few other members of the Enterobacteriales can produce functional amyloids known as curli. These extracellular fibrils are involved in biofilm formation and studies have shown that they may act as virulence factors during infections. It is not known whether curli fibrils are restricted to the Enterobacteriales or if they are phylogenetically widespread. The growing number of genome-sequenced bacteria spanning many phylogenetic groups allows a reliable bioinformatic investigation of the phylogenetic diversity of the curli system. Here we show that the curli system is phylogenetically much more widespread than initially assumed, spanning at least four phyla. Curli fibrils may consequently be encountered frequently in environmental as well as pathogenic biofilms, which was supported by identification of curli genes in public metagenomes from a diverse range of habitats. Identification and comparison of curli subunit (CsgA/B) homologs show that these proteins allow a high degree of freedom in their primary protein structure, although a modular structure of tightly spaced repeat regions containing conserved glutamine, asparagine and glycine residues has to be preserved. In addition, a high degree of variability within the operon structure of curli subunits between bacterial taxa suggests that the curli fibrils might have evolved to fulfill specific functions. Variations in the genetic organization of curli genes are also seen among different bacterial genera. This suggests that some genera may utilize alternative regulatory pathways for curli expression. Comparison of phylogenetic trees of Csg proteins and the 16S rRNA genes of the corresponding bacteria showed remarkably similar overall topography, suggesting that horizontal gene transfer is a minor player in the spreading of the curli system. PMID:23251478

Dueholm, Morten S; Albertsen, Mads; Otzen, Daniel; Nielsen, Per Halkjær

2012-01-01

345

Glial expression of the {beta}-Amyloid Precursor Protein (APP) in global ischemia  

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The {beta}-amyloid precursor protein (APP) bears characteristics of an acute-phase protein and therefore is likely to be involved in the glial response to brain injury. In the brain, APP is rapidly synthesized by activated glial cells in response to comparatively mild neuronal lesions, e.g., a remote peripheral nerve injury. Perfusion deficits in the brain result largely in neuronal necrosis and are a common condition in elderly patients. This neuronal necrosis is accompanied by a pronounced reaction of astrocytes and microglia, which can also be observed in animal models. We have therefore studied in the rat, immunocytochemically, the induction of APP after 30 min of global ischemia caused by four-vessel occlusion. The postischemic brain injuries were examined at survival times from 12 h to 7 days. From day 3 onward, APP immunoreactivity was strongly induced in the CA{sub 1} and CA{sub 4} regions of the rat dorsal hippocampus as well as in the dorsolateral striatum. In these areas, the majority of APP-immunoreactive cells were reactive glial fibrillary acidic protein (GFAP)-positive astrocytes, as shown by double-immunofluorescence labeling for GFAP and APP. Additionally, small ramified cells, most likely activated microglia, expressed APP immunoreactivity. In contrast, in the parietal cortex, APP immunoreactivity occurred focally in clusters of activated microglia rather than in astrocytes, as demonstrated by double-immunofluorescence labeling for APP and the microglia-binding lectin Griffonia simplicifolia isolectin B{sub 4}. In conclusion, following global ischemia, APP is induced in reactive glial cells with spatial differences in the distribution pattern of APP induction in actrocytes and microglia. 51 refs., 4 figs.

Banati, R.B.; Gehrmann, J.; Kreutzberg, G.W. [Max Planck Institute of Psychiarty, Martinsried (Germany)]|[Max Planck Institute for Neurological Research, Koeln (Germany)]|[Univ. Hospital, Zurich (Switzerland)

1995-07-01

346

The amyloid beta-protein precursor and its mammalian homologues. Evidence for a zinc-modulated heparin-binding superfamily.  

Science.gov (United States)

The Alzheimer beta-amyloid precursor protein (APP) contains an ectodomain zinc binding site that has been reported to modulate the heparin affinity and protease-inhibitory properties of the molecule. This motif, GVEFVCCP, is highly conserved in amyloid precursor-like proteins 1 and 2 (APLP1 and APLP2), as well as in the Drosophila and Caenorhabditis elegans APP-like proteins (APPL and APL-1). To determine whether the function of this domain is preserved in the human APP-like proteins, the effect of zinc in modulating the elution profile of these proteins upon heparin-Sepharose chromatography was studied. Both APLP1 and APLP2 bound heparin-Sepharose and had NaCl elution profiles similar to that of APP. As previously reported for APP, zinc increased the recovery of APLP1 and APLP2 upon heparin-Sepharose chromatography. APP, APLP1, and APLP2 all bind zinc-chelating Sepharose, indicating that the zinc binding motif may be functionally conserved in these proteins. Additionally, APP, APLP1, and APLP2 migrate at higher molecular sizes (approximately 40 kDa) on SDS-polyacrylamide gel electrophoresis than their predicted molecular sizes. We report data that compare the physicochemical properties of APP to its novel APLP homologues and indicate that these molecules behave as a family of zinc-modulated, heparin-binding proteins. PMID:7929392

Bush, A I; Pettingell, W H; de Paradis, M; Tanzi, R E; Wasco, W

1994-10-28

347

Injury severity and serum amyloid A correlate with plasma oxidation-reduction potential in multi-trauma patients: a retrospective analysis  

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Full Text Available Abstract Background In critical injury, the occurrence of increased oxidative stress or a reduced antioxidant status has been observed. The purpose of this study was to correlate the degree of oxidative stress, by measuring the oxidation-reduction potential (ORP of plasma in the critically injured, with injury severity and serum amyloid A (SAA levels. Methods A total of 140 subjects were included in this retrospective study comprising 3 groups: healthy volunteers (N = 21, mild to moderate trauma (ISS Results ORP maxima were reached on day 3 (± 0.4 SEM and day 5 (± 0.5 SEM for the ISS Conclusion The results suggest the presence of an oxidative environment in the plasma of the critically injured as measured by ORP. More importantly, ORP can differentiate the degree of oxidative stress based on the severity of the trauma and degree of inflammation.

Mains Charles W

2009-11-01

348

The effect of zinc on amyloid ?-protein assembly and toxicity: A mechanistic investigation  

Science.gov (United States)

Neurotoxic assemblies of amyloid ?-protein (A?) are widely believed to be the cause for Alzheimer's disease (AD). Therefore, understanding the factors and mechanisms that control, modulate, and inhibit formation of these assemblies is crucial for the development of therapeutic intervention of AD. This information also can contribute significantly to our understanding of the mechanisms of other amyloidosis diseases, such as Parkinson's disease, Huntington's disease, type 2 diabetes, amyotrophic lateral sclerosis (Lou Gehrig's disease) and prion diseases (e.g. Mad Cow disease). We have developed a multidisciplinary experimental strategy to study structural and dynamic mechanistic aspects that underlie the A? assembly process. Utilizing this strategy, we explored the molecular basis leading to the perturbation of the A? assembly process by divalent metal ions, mainly Zn2+ ions. Using Zn2+ as reaction physiological relevant probes, it was demonstrated that Zn2+ rapidly (milliseconds) induce self-assembly of A? aggregates and stabilize them in a manner that prevents formation of A? fibrils. Importantly, the early-formed intermediates are substantially more neurotoxic than fibrils. Our results suggest that relevant A? modulators should be targeted against the rapidly evolved intermediate states of A? assembly. The design of such modulators is challenging, as they have to compete with different natural mediators (such as Zn2+) of A? aggregation, which diverse A? assemblies in both specific and nonspecific manners.

Solomonov, Inna; Sagi, Irit

2014-10-01

349

Substrate sequence influences ?-secretase modulator activity, role of the transmembrane domain of the amyloid precursor protein.  

Science.gov (United States)

A subset of non-steroidal anti-inflammatory drugs modulates the ? cleavage site in the amyloid precursor protein (APP) to selectively reduce production of A?42. It is unclear precisely how these ?-secretase modulators (GSMs) act to preferentially spare A?40 production as well as Notch processing and signaling. In an effort to determine the substrate requirements in NSAID/GSM activity, we determined the effects of sulindac sulfide and flurbiprofen on ?-cleavage of artificial constructs containing several ?-secretase substrates. Using FLAG-tagged constructs that expressed extracellularly truncated APP, Notch-1, or CD44, we found that these substrates have different sensitivities to sulindac sulfide. ?-Secretase cleavage of APP was altered by sulindac sulfide, but CD44 and Notch-1 were either insensitive or only minimally altered by this compound. Using chimeric APP constructs, we observed that the transmembrane domain (TMD) of APP played a pivotal role in determining drug sensitivity. Substituting the APP TMD with that of APLP2 retained the sensitivity to ?-cleavage modulation, but replacing TMDs from Notch-1 or ErbB4 rendered the resultant molecules insensitive to drug treatment. Specifically, the GXXXG motif within APP appeared to be critical to GSM activity. Consequently, the modulatory effects on ?-cleavage appears to be substrate-dependent. We hypothesize that the substrate present in the ?-secretase complex influences the conformation of the complex so that the binding site of GSMs is either stabilized or less favorable to influence the cleavage of the respective substrates. PMID:21868380

Sagi, Sarah A; Lessard, Christian B; Winden, Kellen D; Maruyama, Hiroko; Koo, Jeremy C; Weggen, Sascha; Kukar, Thomas L; Golde, Todd E; Koo, Edward H

2011-11-18

350

Memory loss caused by beta-amyloid protein is rescued by a beta(3)-adrenoceptor agonist.  

Science.gov (United States)

Accumulation of the neurotoxic beta-amyloid protein (Abeta) in the brain is a key step in the pathogenesis of Alzheimer's disease (AD). Although transgenic mouse models of AD have been developed, there is a clear need for a validated animal model of Abeta-induced amnesia which can be used for toxicity testing and drug development. Intracranial injections of Abeta(1-42) impaired memory in a single trial discriminative avoidance learning task in chicks. Memory inhibition was closely associated with the state of aggregation of the Abeta peptide, and a scrambled-sequence of Abeta(1-42) peptide failed to impair memory. Abeta had little effect on labile (short-term and intermediate) memory, but blocked consolidation of memory into long-term storage mimicking the type of anterograde amnesia that occurs in early AD. Since noradrenaline exerts a modulatory influence on labile memory in the chick, we examined the effects of two beta-adrenoceptor (AR) agonists on Abeta-induced amnesia. A beta(3)-AR agonist (CL316243), but not a beta(2)-AR agonist, rescued Abeta-induced memory loss, suggesting the need for further studies on the role of beta(3)-ARs in AD. PMID:18632189

Gibbs, Marie E; Maksel, Danuta; Gibbs, Zoe; Hou, Xu; Summers, Roger J; Small, David H

2010-04-01

351

Lipid rafts: linking prion protein to zinc transport and amyloid-? toxicity in Alzheimer's disease  

Science.gov (United States)

Dysregulation of neuronal zinc homeostasis plays a major role in many processes related to brain aging and neurodegenerative diseases, including Alzheimer's disease (AD). Yet, despite the critical role of zinc in neuronal function, the cellular mechanisms underpinning its homeostatic control are far from clear. We reported that the cellular prion protein (PrPC) is involved in the uptake of zinc into neurons. This PrPC-mediated zinc influx required the metal-binding octapeptide repeats in PrPC and the presence of the zinc permeable AMPA channel with which PrPC directly interacted. Together with the observation that PrPC is evolutionarily related to the ZIP family of zinc transporters, these studies indicate that PrPC plays a key role in neuronal zinc homeostasis. Therefore, PrPC could contribute to cognitive health and protect against age-related zinc dyshomeostasis but PrPC has also been identified as a receptor for amyloid-? oligomers which accumulate in the brains of those with AD. We propose that the different roles that PrPC has are due to its interaction with different ligands and/or co-receptors in lipid raft-based signaling/transport complexes. PMID:25364748

Watt, Nicole T.; Griffiths, Heledd H.; Hooper, Nigel M.

2014-01-01

352

Substrate Sequence Influences ?-Secretase Modulator Activity, Role of the Transmembrane Domain of the Amyloid Precursor Protein*  

Science.gov (United States)

A subset of non-steroidal anti-inflammatory drugs modulates the ? cleavage site in the amyloid precursor protein (APP) to selectively reduce production of A?42. It is unclear precisely how these ?-secretase modulators (GSMs) act to preferentially spare A?40 production as well as Notch processing and signaling. In an effort to determine the substrate requirements in NSAID/GSM activity, we determined the effects of sulindac sulfide and flurbiprofen on ?-cleavage of artificial constructs containing several ?-secretase substrates. Using FLAG-tagged constructs that expressed extracellularly truncated APP, Notch-1, or CD44, we found that these substrates have different sensitivities to sulindac sulfide. ?-Secretase cleavage of APP was altered by sulindac sulfide, but CD44 and Notch-1 were either insensitive or only minimally altered by this compound. Using chimeric APP constructs, we observed that the transmembrane domain (TMD) of APP played a pivotal role in determining drug sensitivity. Substituting the APP TMD with that of APLP2 retained the sensitivity to ?-cleavage modulation, but replacing TMDs from Notch-1 or ErbB4 rendered the resultant molecules insensitive to drug treatment. Specifically, the GXXXG motif within APP appeared to be critical to GSM activity. Consequently, the modulatory effects on ?-cleavage appears to be substrate-dependent. We hypothesize that the substrate present in the ?-secretase complex influences the conformation of the complex so that the binding site of GSMs is either stabilized or less favorable to influence the cleavage of the respective substrates. PMID:21868380

Sagi, Sarah A.; Lessard, Christian B.; Winden, Kellen D.; Maruyama, Hiroko; Koo, Jeremy C.; Weggen, Sascha; Kukar, Thomas L.; Golde, Todd E.; Koo, Edward H.

2011-01-01

353

Lipid rafts: linking prion protein to zinc transport and amyloid-? toxicity in Alzheimer's disease.  

Science.gov (United States)

Dysregulation of neuronal zinc homeostasis plays a major role in many processes related to brain aging and neurodegenerative diseases, including Alzheimer's disease (AD). Yet, despite the critical role of zinc in neuronal function, the cellular mechanisms underpinning its homeostatic control are far from clear. We reported that the cellular prion protein (PrP(C)) is involved in the uptake of zinc into neurons. This PrP(C)-mediated zinc influx required the metal-binding octapeptide repeats in PrP(C) and the presence of the zinc permeable AMPA channel with which PrP(C) directly interacted. Together with the observation that PrP(C) is evolutionarily related to the ZIP family of zinc transporters, these studies indicate that PrP(C) plays a key role in neuronal zinc homeostasis. Therefore, PrP(C) could contribute to cognitive health and protect against age-related zinc dyshomeostasis but PrP(C) has also been identified as a receptor for amyloid-? oligomers which accumulate in the brains of those with AD. We propose that the different roles that PrP(C) has are due to its interaction with different ligands and/or co-receptors in lipid raft-based signaling/transport complexes. PMID:25364748

Watt, Nicole T; Griffiths, Heledd H; Hooper, Nigel M

2014-01-01

354

Overexpression of amyloid precursor protein increases copper content in HEK293 cells  

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Amyloid precursor protein (APP) is a transmembrane glycoprotein widely expressed in mammalian tissues and plays a central role in Alzheimer's disease. However, its physiological function remains elusive. Cu{sup 2+} binding and reduction activities have been described in the extracellular APP135-156 region, which might be relevant for cellular copper uptake and homeostasis. Here, we assessed Cu{sup 2+} reduction and {sup 64}Cu uptake in two human HEK293 cell lines overexpressing APP. Our results indicate that Cu{sup 2+} reduction increased and cells accumulated larger levels of copper, maintaining cell viability at supra-physiological levels of Cu{sup 2+} ions. Moreover, wild-type cells exposed to both Cu{sup 2+} ions and APP135-155 synthetic peptides increased copper reduction and uptake. Complementation of function studies in human APP751 transformed Fre1 defective Saccharomyces cerevisiae cells rescued low Cu{sup 2+} reductase activity and increased {sup 64}Cu uptake. We conclude that Cu{sup 2+} reduction activity of APP facilitates copper uptake and may represent an early step in cellular copper homeostasis.

Suazo, Miriam; Hodar, Christian; Morgan, Carlos [INTA, Laboratorio de Bioinformatica y Expresion Genica, Universidad de Chile, El Libano 5524, Macul, Santiago (Chile); Cerpa, Waldo [Centro de Envejecimiento y Regeneracion (CARE), Centro de Regulacion Celular y Patologia ' Joaquin V. Luco' (CRCP), MIFAB, Departamento de Biologia Celular y Molecular, Facultad de Ciencias Biologicas, Pontificia Universidad Catolica de Chile, Santiago (Chile); Cambiazo, Veronica [INTA, Laboratorio de Bioinformatica y Expresion Genica, Universidad de Chile, El Libano 5524, Macul, Santiago (Chile); Millenium Nucleus CGC, Universidad de Chile (Chile); Inestrosa, Nibaldo C. [Centro de Envejecimiento y Regeneracion (CARE), Centro de Regulacion Celular y Patologia ' Joaquin V. Luco' (CRCP), MIFAB, Departamento de Biologia Celular y Molecular, Facultad de Ciencias Biologicas, Pontificia Universidad Catolica de Chile, Santiago (Chile); Gonzalez, Mauricio, E-mail: mgonzale@inta.cl [INTA, Laboratorio de Bioinformatica y Expresion Genica, Universidad de Chile, El Libano 5524, Macul, Santiago (Chile)

2009-05-15

355

Phenotypical difference of Amyloid Precursor Protein (APP V717L mutation in Japanese family  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Alzheimer’s disease (AD is the most common form of dementia. Mutations in genes such as those encoding amyloid precursor protein (APP, presenilin 1 and presenilin 2, are responsible for early-onset familial AD. Case presentation In this study, we report a 275341 G > C (Val717Leu mutation in the APP gene in a Japanese family with early onset AD by genetic screening. This mutation has previously been detected in European families. In the Japanese family we screened, the age at onset of AD was 47.1 ± 3.1 years old (n = 9; range, 42–52. The symptoms in the affected members included psychiatric vulnerability and focal signs such as pyramidal signs, epileptic seizures, and myoclonic discharges. An MR imaging study showed relatively mild atrophic changes in the bilateral hippocampus and cerebral cortices in all affected members compared with their clinical presentations. Conclusion We conclude that the clinical features of Alzheimer’s disease can be different even when caused by the same mutation in the APP gene. Further clinical and genetic studies are required to clarify the relationship between phenotypes and genotypes.

Abe Masao

2012-06-01

356

Novel mutations in the amyloid precursor protein gene within Moroccan patients with Alzheimer's disease.  

Science.gov (United States)

In Morocco, Alzheimer's disease (AD) affects almost 30,000 individuals, and this number could increase to 75,000 by 2020. To our knowledge, the genes predisposing individuals to AD and predicting disease incidence remain elusive. In this study, we aimed to evaluate the genetic contribution of mutations in the amyloid precursor protein (APP) gene exons 16 and 17 to familial and sporadic AD cases. Seventeen sporadic cases and eight family cases were seen at the memory clinic of the University of Casablanca Neurology Department. These patients underwent standard somatic neurological examination, cognitive function assessment, brain imaging, and laboratory tests. Direct sequencing of exons 16 and 17 of the APP gene was performed on genomic DNA of AD patients. In this original Moroccan study, we identified seven novel frameshift mutations in exons 16 and 17 of the APP gene. Interestingly, only one novel splice mutation was detected in a family case. There is a strong correlation between clinical symptoms and genetic factors in Moroccan patients with a family history of AD. Therefore, mutations in APP gene exons 16 and 17 may eventually become genetic markers for AD predisposition. PMID:24627227

El Kadmiri, Nadia; Zaid, Nabil; Hachem, Ahmed; Zaid, Younes; Dubé, Marie-Pierre; Hamzi, Khalil; El Moutawakil, Bouchra; Slassi, Ilham; Nadifi, Sellama

2014-06-01

357

Solid-state NMR analysis of interaction sites of curcumin and 42-residue amyloid ?-protein fibrils.  

Science.gov (United States)

Aggregation of 42-residue amyloid ?-protein (A?42) plays a pivotal role in the etiology of Alzheimer's disease (AD). Curcumin, the yellow pigment in the rhizome of turmeric, attracts considerable attention as a food component potentially preventing the pathogenesis of AD. This is because curcumin not only inhibits the aggregation of A?42 but also binds to its aggregates (fibrils), resulting in disaggregation. However, the mechanism of interaction between curcumin and the A?42 fibrils remains unclear. In this study, we analyzed the binding mode of curcumin to the A?42 fibrils by solid-state NMR using dipolar-assisted rotational resonance (DARR). To improve the quality of 2D spectra, 2D DARR data were processed with the covariance NMR method, which enabled us to detect weak cross peaks between carbons of curcumin and those of the A?42 fibrils. The observed (13)C-(13)C cross peaks indicated that curcumin interacts with the 12th and 17-21st residues included in the ?-sheet structure in the A?42 fibrils. Interestingly, aromatic carbons adjacent to the methoxy and/or hydroxy groups of curcumin showed clear cross peaks with the A?42 fibrils. This suggested that these functional groups of curcumin play an important role in its interaction with the A?42 fibrils. PMID:21924918

Masuda, Yuichi; Fukuchi, Masashi; Yatagawa, Tatsuya; Tada, Masato; Takeda, Kazuyuki; Irie, Kazuhiro; Akagi, Ken-Ichi; Monobe, Youko; Imazawa, Takayoshi; Takegoshi, K

2011-10-15

358

Solid-state NMR analysis of the ?-strand orientation of the protofibrils of amyloid ?-protein  

International Nuclear Information System (INIS)

Highlights: ? The supramolecular structure of A?42 protofibrils was analyzed by solid-state NMR. ? The Ala-21 residue in the A?42 protofibrils is included in a slightly disordered ?-strand. ? The A?42 protofibrils do not form intermolecular in-register parallel ?-sheets. -- Abstract: Alzheimer’s disease (AD) is caused by abnormal deposition (fibrillation) of a 42-residue amyloid ?-protein (A?42) in the brain. During the process of fibrillation, the A?42 takes the form of protofibrils with strong neurotoxicity, and is thus believed to play a crucial role in the pathogenesis of AD. To elucidate the supramolecular structure of the A?42 protofibrils, the intermolecular proximity of the Ala-21 residues in the A?42 protofibrils was analyzed by 13C–13C rotational resonance experiments in the solid state. Unlike the A?42 fibrils, an intermolecular 13C–13C correlation was not found in the A?42 protofibrils. This result suggests that the ?-strands of the A?42 protofibrils are not in an in-register parallel orientation. A?42 monomers would assemble to form protofibrils with the ?-strand conformation, then transform into fibrils by forming intermolecular parallel ?-sheets.

359

Activated protein C inhibits amyloid ? production via promoting expression of ADAM-10.  

Science.gov (United States)

Inhibition of A? production and clearance of senile plaques have been considered as potential strategies in the treatment of Alzheimer's disease (AD). Activated protein C (APC) is an important factor in the anticoagulant system. However, whether APC can influence the condition of a chronic neurodegenerative process, such as that present in AD, is unknown. In this study, we found that administration of APC on AD Tg2576 mice significantly reduced amyloid ? production and helped to facilitate cognitive improvement. APC could also reduce levels of A?40 and A?42 produced in APPswe cells, an AD cell model. Further results demonstrated that APC did not change the levels of A?-degrading enzymes, insulin-degrading enzyme (IDE), or neprilysin (NEP). Next, we found that APC promoted sAPP? and CTF? release and inhibited sAPP? and CTF? release, thereby indicating that APC could regulate A? secretion by shifting APP processing from the amyloidogenic pathway toward the nonamyloidogenic pathway. Correspondingly, further study revealed that ADAM-10 expression was increased by APC, suggesting that APC inhibits A? secretion through stimulating activity of ?-secretase. These findings support the idea that APC could hold therapeutic potential in the treatment of AD. PMID:24333930

Li, Bing; Yu, Dawei; Xu, Zhiying

2014-01-30

360

3,3',4',5'-tetrahydroxyflavone induces formation of large aggregates of amyloid ? protein.  

Science.gov (United States)

Amyloid ? protein (A?) self-assembles into insoluble fibrils, and forms the senile plaques associated with Alzheimer's disease. 3,3',4',5'-Tetrahydroxyflavone, a synthetic analogue of the natural flavonoid fisetin, has been found to potently inhibit A? fibril formation. In the present study, we investigated how inhibition of A? fibril formation by this flavonoid affects A? conformation and neurotoxicity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of A?1-42 (20?µM) incubated with or without 3,3',4',5'-tetrahydroxyflavone demonstrated that 3,3',4',5'-tetrahydroxyflavone (100?µM) rapidly caused formation of atypical A? conformers, which appeared as a very broad, smear-like band in the high molecular weight region and were distinguishable from soluble A? oligomers or mature A? fibrils. Transmission electron microscopy (TEM) revealed that large spherical A? aggregates were preferentially formed in the presence of 3,3',4',5'-tetrahydroxyflavone. The SDS-resistant, smear-like band on SDS-PAGE and the large spherical aggregates in TEM both disappeared after heat treatment (100°C, 10?min). Furthermore, a neurotoxicity assay with cultured rat hippocampal neurons demonstrated that A? incubated with 3,3',4',5'-tetrahydroxyflavone was significantly less toxic than A? incubated without the flavonoid. These results suggest that the newly synthesized fisetin analogue 3,3',4',5'-tetrahydroxyflavone directly produces atypical, large A? aggregates and reduces A? toxicity. PMID:24789998

Ushikubo, Hiroko; Tanimoto, Yui; Abe, Kazuho; Asakawa, Tomohiro; Kan, Toshiyuki; Akaishi, Tatsuhiro

2014-01-01

 
 
 
 
361

Dimerization of the transmembrane domain of amyloid precursor proteins and familial Alzheimer's disease mutants  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Amyloid precursor protein (APP is enzymatically cleaved by ?-secretase to form two peptide products, either A?40 or the more neurotoxic A?42. The A?42/40 ratio is increased in many cases of familial Alzheimer's disease (FAD. The transmembrane domain (TM of APP contains the known dimerization motif GXXXA. We have investigated the dimerization of both wild type and FAD mutant APP transmembrane domains. Results Using synthetic peptides derived from the APP-TM domain, we show that this segment is capable of forming stable transmembrane dimers. A model of a dimeric APP-TM domain reveals a putative dimerization interface, and interestingly, majority of FAD mutations in APP are localized to this interface region. We find that FAD-APP mutations destabilize the APP-TM dimer and increase the population of APP peptide monomers. Conclusion The dissociation constants are correlated to both the A?42/A?40 ratio and the mean age of disease onset in AD patients. We also show that these TM-peptides reduce A? production and A?42/A?40 ratios when added to HEK293 cells overexpressing the Swedish FAD mutation and ?-secretase components, potentially revealing a new class of ?-secretase inhibitors.

Fraser Paul E

2008-01-01

362

HAPTOGLOBIN AND SERUM AMYLOID A IN SUBACUTE RUMINAL ACIDOSIS IN GOATS / HAPTOGLOBINA Y PROTEÍNA AMILÓIDE SÉRICA A EN ACIDOSIS RUMINAL SUBAGUADA EN CABRAS  

Scientific Electronic Library Online (English)

Full Text Available SciELO Colombia | Language: English Abstract in spanish La acidosis ruminal es un trastorno frecuente en cabras como consecuencia de errores en el manejo alimentario en animales no adaptados a dietas que contienen carbohidratos fácilmente fermentables. La forma subaguda de la enfermedad es de difícil diagnóstico toda vez que no muestra evidencia de signo [...] s clínicos claros y los parámetros ácido-básicos pueden permanecer en el rango normal. El presente estudio tuvo por objetivo probar la hipótesis de que la haptoglobina y la proteína amilóide sérica A, las dos proteínas de fase aguda más importantes en rumiantes, pueden ser útiles como marcadores de acidosis subaguda en cabras. Se indujo acidosis ruminal a seis cabras de la raza Murciano-Granadina, no adaptadas al consumo de concentrado, mediante el suministro de una dieta con 60% de concentrado y 40% de heno de alfalfa durante 5 días. Dos cabras fueron sometidas a fistulación ruminal para comprobar el efecto del tratamiento sobre el pH del rumen. A todos los animales se les tomaron muestras de sangre y orina el día anterior a la inducción, durante el período de inducción y hasta 18 días después de la inducción (período de recuperación). El pH ruminal cayó a menos de 5,5 durante el período de inducción de acidosis en las cabras fistuladas, mientras que la mitad de las cabras tuvieron diarrea al tercer día de la inducción de acidosis. Los parámetros gasométricos indicaron que los mecanismos compensatorios fueron eficientes para mantener el equilibrio ácido-básico. La haptoglobina sérica presentó un aumento moderado durante el período de inducción de acidosis, mientras que la amilóide sérica A no presentó cambios. Los resultados sugieren que la haptoglobina puede utilizarse como un potencial indicador de acidosis ruminal en cabras. Abstract in english Ruminal acidosis is a frequent disorder that occurs in goats as a consequence of feeding mistakes in animals not adapted to a diet of easily fermentable carbohydrates. The subacute form of the disease is difficult to diagnose because no apparent signs are shown and the acid-base parameters may remai [...] n within the normal range. The present study aimed at testing the hypothesis that haptoglobin (Hp) and serum amyloid A (SAA), the two major acute phase proteins in ruminants, may be useful as markers of subacute acidosis in goats. A subacute acidosis was induced in six Murciano-Granadina goats through a diet of 60% mixed feed-40% alfalfa hay offered during 5 days to goats not adapted to eat mixed feed. Two goats were rumen-fistulated to investigate the effect of feeding on ruminal pH. Sampling of blood and urine of all animals was done before the induction of the acidosis, during 5 days after the onset of induction and for 18 days after the induction (recovery period). Ruminal pH in the fistulated goats dropped to less than 5.5 during the induction period, and half of the goats had diarrhea on the third day after the induction of acidosis. Acid-base parameters showed that the acid-base compensatory mechanisms were efficient in maintaining the equilibrium. Serum Hp had a moderate increase during the induction period, while SAA did not change. These results suggest that Hp might be a potential marker for ruminal acidosis in goats.

F.H.D, González; F.H, Ruipérez; J.M, Sánchez; J.C, Souza; S, Martínez-Subiela; J.J, Cerón.

2010-12-01

363

Molecular Mechanisms of Alzheimer Disease Protection by the A673T Allele of Amyloid Precursor Protein.  

Science.gov (United States)

Pathogenic mutations in the amyloid precursor protein (APP) gene have been described as causing early onset familial Alzheimer disease (AD). We recently identified a rare APP variant encoding an alanine-to-threonine substitution at residue 673 (A673T) that confers protection against development of AD (Jonsson, T., Atwal, J. K., Steinberg, S., Snaedal, J., Jonsson, P. V., Bjornsson, S., Stefansson, H., Sulem, P., Gudbjartsson, D., Maloney, J., Hoyte, K., Gustafson, A., Liu, Y., Lu, Y., Bhangale, T., Graham, R. R., Huttenlocher, J., Bjornsdottir, G., Andreassen, O. A., Jönsson, E. G., Palotie, A., Behrens, T. W., Magnusson, O. T., Kong, A., Thorsteinsdottir, U., Watts, R. J., and Stefansson, K. (2012) Nature 488, 96-99). The Ala-673 residue lies within the ?-secretase recognition sequence and is part of the amyloid-? (A?) peptide cleavage product (position 2 of A?). We previously demonstrated that the A673T substitution makes APP a less favorable substrate for cleavage by BACE1. In follow-up studies, we confirm that A673T APP shows reduced cleavage by BACE1 in transfected mouse primary neurons and in isogenic human induced pluripotent stem cell-derived neurons. Using a biochemical approach, we show that the A673T substitution modulates the catalytic turnover rate (Vmax) of APP by the BACE1 enzyme, without affecting the affinity (Km) of the APP substrate for BACE1. We also show a reduced level of A?(1-42) aggregation with A2T A? peptides, an observation not conserved in A?(1-40) peptides. When combined in a ratio of 1:9 A?(1-42)/A?(1-40) to mimic physiologically relevant mixtures, A2T retains a trend toward slowed aggregation kinetics. Microglial uptake of the mutant A?(1-42) peptides correlated with their aggregation level. Cytotoxicity of the mutant A? peptides was not dramatically altered. Taken together, our findings demonstrate that A673T, a protective allele of APP, reproducibly reduces amyloidogenic processing of APP and also mildly decreases A? aggregation. These effects could together have an additive or even synergistic impact on the risk of developing AD. PMID:25253696

Maloney, Janice A; Bainbridge, Travis; Gustafson, Amy; Zhang, Shuo; Kyauk, Roxanne; Steiner, Pascal; van der Brug, Marcel; Liu, Yichin; Ernst, James A; Watts, Ryan J; Atwal, Jasvinder K

2014-11-01

364

Calcium-enhanced aggregation of serum amyloid P component and its inhibition by the ligands heparin and heparan sulphate. An electron microscopic and immunoelectrophoretic study.  

DEFF Research Database (Denmark)

Serum amyloid P component (SAP) is a pentraxin found in the circulation and in all forms of amyloid deposits. Its physiological and pathophysiological functions are largely unknown. Electron microscopy showed purified human SAP to consist of double pentameric discs compatible with the results of size chromatography. The formation of double pentamers did not require calcium ions. The outer diameter of the discs arranged face-to-face was 11.6 nm a