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Sample records for serum amyloid protein

  1. Heterogeneity of human serum amyloid A proteins

    OpenAIRE

    1980-01-01

    Serum amyloid A proteins (SAA), presumed precursors of the tissue amyloid A proteins (AA) characteristic of secondary amyloidosis, have been isolated from the plasma high-density lipoproteins (HDL) of normals after etiocholanolone-induced inflammation and from patients with Wegener's granulomatosis, systemic lupus erythematosis, juvenile rheumatoid arthritis, Waldenstrom's macroglobulinemia, and Goodpasture's syndrome. At least six polymorphic forms of SAA wer identified among the low molecul...

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

    International Nuclear Information System (INIS)

    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

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

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    Golam Sadik

    2000-01-01

    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.

  4. The Acute-Phase Proteins Serum Amyloid A and C Reactive Protein in Transudates and Exudates

    Directory of Open Access Journals (Sweden)

    2006-01-01

    Full Text Available The distinction between exudates and transudates is very important in the patient management. Here we evaluate whether the acute-phase protein serum amyloid A (SAA, in comparison with C reactive protein (CRP and total protein (TP, can be useful in this discrimination. CRP, SAA, and TP were determined in 36 exudate samples (27 pleural and 9 ascitic and in 12 transudates (9 pleural and 3 ascitic. CRP, SAA, and TP were measured. SAA present in the exudate corresponded to 10 % of the amount found in serum, that is, the exudate/serum ratio (E/S was 0.10 ± 0.13 . For comparison, the exudate/serum ratio for CRP and TP was 0.39 ± 0.37 and 0.68 ± 0.15 , respectively. There was a strong positive correlation between serum and exudate SAA concentration ( r = 0.764 ; p < 0.0001 . The concentration of SAA in transudates was low and did not overlap with that found in exudates (0.02-0.21 versus 0.8–360.5un g/mL. SAA in pleural and ascitic exudates results mainly from leakage of the serum protein via the inflamed membrane. A comparison of the E/S ratio of SAA and CRP points SAA as a very good marker in discriminating between exudates and transudates

  5. Serum amyloid A is a retinol binding protein that transports retinol during bacterial infection

    OpenAIRE

    Derebe, Mehabaw G.; Zlatkov, Clare M.; Gattu, Sureka; Ruhn, Kelly A.; Vaishnava, Shipra; Diehl, Gretchen E.; Macmillan, John B.; Williams, Noelle S.; Hooper, Lora V.

    2014-01-01

    Retinol plays a vital role in the immune response to infection, yet proteins that mediate retinol transport during infection have not been identified. Serum amyloid A (SAA) proteins are strongly induced in the liver by systemic infection and in the intestine by bacterial colonization, but their exact functions remain unclear. Here we show that mouse and human SAAs are retinol binding proteins. Mouse and human SAAs bound retinol with nanomolar affinity, were associated with retinol in vivo, an...

  6. A comparison of serum amyloid A (SAA) synthesis with that of the pentraxins: Serum amyloid P (SAP) and C-reactive protein (CRP)

    International Nuclear Information System (INIS)

    Serum amyloid A (SAA) and serum amyloid P (SAP) were detected in cultures of hepatocytes which had been isolated from normal CBA/J mice by the collagenase perfusion technique. SAP production in 24 h cultures was more resistant than SAA and total protein synthesis to inhibition by actinomycin D, but was more sensitive to inhibition by 48 h. However, the production of SAP was more sensitive to cycloheximide than SAA and total protein throughout the 48 hr incubation period. SAP and SAA levels in the culture media were suppressed by treatment of liver cells with 10-6 M of colchicine for 48 h. Inhibition of SAP production by colchicine was the same regardless of culture condition, but the effect of colchicine on SAA synthesis varied according to the presence of serum of monokine. These observations also support the concept that the two amyloid proteins are produced under different regulatory mechanisms. When C-reactive protein (CRP) was not detected in the sera of patients with severe chronic liver diseases, the SAA levels were very low. When CRP was detected, SAA values were within the normal range. Thus, in order to produce SAA, liver cells in these patients not only were viable but also maintained their specialized function

  7. Amyloidosis and the serum amyloid A protein response to muramyl dipeptide analogs and different mycobacterial species.

    OpenAIRE

    Mcadam, K. P.; Foss, N. T.; Garcia, C.; Delellis, R.; Chedid, L.; Rees, R. J.; Wolff, S. M.

    1983-01-01

    Serum amyloid A protein (SAA) elevation accompanies induction of secondary amyloidosis in mice given Mycobacterium butyricum in Freund adjuvant. The synthesis of SAA by cultured hepatocytes is induced by a macrophage-derived mediator, which has been identified as interleukin 1. In these studies, SAA synthesis has been used as an index of macrophage activation to examine the in vivo response of mice to challenge with seven different mycobacteria and with synthetic analogs of the immunoadjuvant...

  8. Effect of colchicine on the acute phase serum amyloid A protein response and splenic amyloid deposition during experimental murine inflammation

    International Nuclear Information System (INIS)

    We investigated the effects of colchicine on the acute phase serum amyloid A protein (SAA) response and splenic amyloid A protein (AA) deposition in CBA/J mice undergoing chronic inflammatory stimulation with silver nitrate (AgNO3), and on accelerated amyloid deposition induced by amyloidenhancing factor (AEF). Colchicine (10 microg daily) significantly lowered splenic AA levels after 25 days of inflammation, as determined by radioimmunoassay. Pretreatment (3 days) with colchicine decreased SAA levels 24 h after AgNO3. It was (unexpectedly) observed that brief pretreatment (12 h) with colchicine augmented the acute phase SAA response to AgNO3 at 24 h. Colchicine stimulated production of both the SAA inducer and lymphocyte-activating factor (LAF) activities of interleukin 1 (IL 1) by macrophages. Decreased SAA levels did not appear to be the mechanism by which colchicine inhibited amyloidosis, since SAA levels fell both in colchicinetreated and control mice after 25 days of inflammation. Colchicine only partially lowered AA deposition after injection of AEF. This effect could be explained by decreased acute phase SAA levels. It is postulated that colchicine inhibits amyloidosis in the pre-deposition period by altering the production of factors (e.g., AEF) required in the deposition phase

  9. Docosahexaenoic Acid Enhances Hepatic Serum Amyloid A Expression via Protein Kinase A-dependent Mechanism*

    OpenAIRE

    Tai, Chen C.; Chen, Ching Y.; Lee, Hsuan S.; Wang, Ya C.; Li, Tsai K.; Mersamm, Harry J.; Ding, Shih T.; Wang, Pei H.

    2009-01-01

    Serum amyloid A (SAA) reduces fat deposition in adipocytes and hepatoma cells. Human SAA1 mRNA is increased by docosahexaenoic acid (DHA) treatment in human cells. These studies asked whether DHA decreases fat deposition through SAA1 and explored the mechanisms involved. We demonstrated that DHA increased human SAA1 and C/EBP? mRNA expression in human hepatoma cells, SK-HEP-1. Utilizing a promoter deletion assay, we found that a CCAAT/enhancer-binding protein ? (C/EBP?)-binding site in the...

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

    International Nuclear Information System (INIS)

    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 same similarity with the corresponding AA protein as previously reported for human SAA/AA

  11. Solid-phase immunoradiometric assay for serum amyloid A protein using magnetisable cellulose particles.

    Science.gov (United States)

    De Beer, F C; Dyck, R F; Pepys, M B

    1982-10-29

    An immunoradiometric assay for human serum amyloid A protein (SAA) was developed using magnetisable cellulose particles as the solid phase. Rabbit antiserum to to SAA was raised by immunization with SAA isolated from acute-phase serum by gel filtration in formic acid. The antiserum was rendered monospecific for SAA by solid-phase immunoabsorption with normal human serum, which contains only traces of SAA, and some was coupled covalently to the cellulose particles. Immunopurified anti-SAA antibodies were isolated from the monospecific anti-SAA serum by binding to, and elution from insolubilized acute-phase serum and were radiolabelled with 125I. The assay was calibrated with an acute phase serum which contained 6000 times more SAA than normal sera with the lowest detectable level of SAA, and an arbitrary value of 6000 U/l was assigned to this standard. Sera were tested in the native, undenatured state and there was no increase in SAA immunoreactivity following alkali treatment or heating. The assay range was from 1-2000 U/l so that all SAA levels above 6 U/l could be measured on a single (1:6) dilution of serum. The intra- and interassay coefficients of variation were 11.7 and 15.0% respectively. Among 100 healthy normal subjects (50 male, 50 female) the median SAA level was 9 U/l, range less than 1-100, with 93% below 20 U/l and only 2% below the lower limit of sensitivity of the assay (1 U/l). PMID:7175191

  12. Solid-phase immunoradiometric assay for serum amyloid A protein using magnetisable cellulose particles

    International Nuclear Information System (INIS)

    An immunoradiometric assay for human serum amyloid A protein (SAA) was developed using magnetisable cellulose particles as the solid phase. Rabbit antiserum to SAA was raised by immunization with SAA isolated from acute-phase serum by gel filtration in formic acid. The antiserum was rendered monospecific for SAA by solid-phase immunoabsorption with normal human serum, which contains only traces of SAA, and some was coupled covalently to the cellulose particles. Immunopurified anti-SAA antibodies were isolated from the monospecific anti-SAA serum by binding to, and elution from insolubilized acute-phase serum and were radiolabelled with 125I. The assay was calibrated with an acute phase serum which contained 6000 times more SAA than normal sera with the lowest detectable level of SAA, and an arbitrary value of 6000 U/l was assigned to this standard. Sera were tested in the native, undenatured state and there was no increase in SAA immunoreactivity following alkali treatment or heating. The assay range was from 1-2000 U/l so that all SAA levels above 6 U/l could be measured on a single (1:6) dilution of serum. The intra- and interassay coefficients of variation were 11.7 and 15.0% respectively. Among 100 healthy normal subjects (50 male, 50 female) the median SAA level was 9 U/l, range <1-100, with 93% below 20 U/l and only 2% below the lower limit of sensitivity of the assay (1 U/l). (Auth.)

  13. Human serum amyloid A protein. Behaviour in aqueous and urea-containing solutions and antibody production.

    Science.gov (United States)

    Strachan, A F; Shephard, E G; Bellstedt, D U; Coetzee, G A; van der Westhuyzen, D R; de Beer, F C

    1989-10-15

    Human serum amyloid A protein (apo-SAA) can be prepared by gel filtration of delipidated acute-phase high-density lipoprotein in the presence of urea. The resultant apo-SAA is soluble (greater than 90% solubility) in a wide range of buffer solutions, with all of the six major isoforms of apo-SAA being equally soluble. In urea-containing solutions the isoforms behave qualitatively differently in various urea concentrations, probably reflecting subtle primary-structure variations. The higher-pI isoforms are only completely unfolded at greater than 7 M-urea. By immunizing with apo-SAA adsorbed to acid-treated bacteria (Salmonella minnesota R595), high-titre antibodies can easily be elicited in rabbits. PMID:2597108

  14. Goodpasture antigen-binding protein/ceramide transporter binds to human serum amyloid P-component and is present in brain amyloid plaques.

    Science.gov (United States)

    Mencarelli, Chiara; Bode, Gerard H; Losen, Mario; Kulharia, Mahesh; Molenaar, Peter C; Veerhuis, Robert; Steinbusch, Harry W M; De Baets, Marc H; Nicolaes, Gerry A F; Martinez-Martinez, Pilar

    2012-04-27

    Serum amyloid P component (SAP) is a non-fibrillar glycoprotein belonging to the pentraxin family of the innate immune system. SAP is present in plasma, basement membranes, and amyloid deposits. This study demonstrates, for the first time, that the Goodpasture antigen-binding protein (GPBP) binds to human SAP. GPBP is a nonconventional Ser/Thr kinase for basement membrane type IV collagen. Also GPBP is found in plasma and in the extracellular matrix. In the present study, we demonstrate that GPBP specifically binds SAP in its physiological conformations, pentamers and decamers. The START domain in GPBP is important for this interaction. SAP and GPBP form complexes in blood and partly colocalize in amyloid plaques from Alzheimer disease patients. These data suggest the existence of complexes of SAP and GPBP under physiological and pathological conditions. These complexes are important for understanding basement membrane, blood physiology, and plaque formation in Alzheimer disease. PMID:22396542

  15. Serum amyloid A and C-reactive protein levels may predict microalbuminuria and macroalbuminuria in newly diagnosed type 1 diabetic patients

    DEFF Research Database (Denmark)

    Overgaard, Julie; McGuire, James N

    2012-01-01

    In this study we evaluated the association of baseline levels of six different candidate proteins for the development of microalbuminuria and macroalbuminuria in type 1 diabetic patients, who were followed for approximately 30years. Two of the proteins are markers of inflammation: serum amyloid A (SAA) and C-reactive protein (CRP), three are involved in lipid metabolism: apolipoprotein A1, apolipoprotein E and adiponectin and the last protein, fibronectin, is related to structural changes.

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

    DEFF Research Database (Denmark)

    Christensen, Michelle BrØnniche; Langhorn, Rebecca

    2014-01-01

    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.

  17. Calumenin interacts with serum amyloid P component.

    DEFF Research Database (Denmark)

    Vorum, H; Jacobsen, Christian

    2000-01-01

    We recently reported the identification of human calumenin, a novel Ca(2+) binding, transformation-sensitive and secreted protein [Vorum et al. (1998) Biochim. Biophys. Acta 1386, 121-131; Vorum et al. (1999) Exp. Cell Res. 248, 473-481] belonging to the family of multiple EF-hand proteins of the secretory pathway that include reticulocalbin, ERC-55, Cab45 and crocalbin. In order to further investigate the extracellular functions of calumenin we immobilized the recombinant protein to a column. After application of a placental tissue extract we were able to elute one protein that interacts with calumenin in the presence of Ca(2+). Amino acid sequencing identified this protein as serum amyloid P component (SAP). Furthermore, we verified and characterized the calumenin-SAP interaction by the surface plasmon resonance technique. The findings indicate that calumenin may participate in the immunological defense system and could be involved in the pathological process of amyloidosis that leads to formation of amyloid deposits seen in different types of tissues. Udgivelsesdato: 2000-Jan-14

  18. 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

  19. Serum amyloid A and C-reactive protein positive nodule in alcoholic liver cirrhosis, hard to make definite diagnosis.

    Science.gov (United States)

    Kim, Soo Ryang; Kondo, Fukuo; Otono, Yumi; Imoto, Susumu; Ando, Kenji; Hirakawa, Makoto; Fukuda, Katsumi; Sasaki, Madoka; Kim, Soo Ki; Komaki, Takamitsu; Tsuchida, Shinobu; Kobayashi, Sawako; Matsuoka, Toshiyuki; Kudo, Masatoshi

    2014-05-01

    We describe a case of serum amyloid A (SAA) and C-reactive protein (CRP) positive nodule detected by immunohistochemical analysis in a 37-year-old woman with alcohol-related cirrhosis. Imaging studies at first admission pointed to hepatocellular carcinoma (HCC), a dysplastic nodule, an inflammatory pseudotumor or focal nodular hyperplasia (FNH). Ultrasonography-guided biopsy in Segment 2 showed minimal atypical changes, except for a slight increase in cell density and micronodular cirrhosis in the non-nodular portion. gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid-enhanced magnetic resonance imaging carried out after a year and a half revealed hypervascularity in the arterial phase and isointensity in the hepatobiliary phase. Three years thereafter, however, the imaging displayed a change from isointensity to a defect in the hepatobiliary phase, and the nodule demonstrated minimal histological atypia. Immunohistochemical staining of the nodule was positive for SAA, CRP, liver fatty acid-binding protein and glutamine synthetase, but negative for ?-catenin, heat shock protein 70 and Glypican 3. Organic anion transporter (OATP)8 staining was weaker in the nodule than in the non-nodular portion of the alcohol-related micronodular cirrhosis. The nodule was diagnosed as an SAA and CRP positive nodule, and HCC was ruled out. Despite the change from isointensity to a defect in the hepatobiliary phase, no evidence of HCC was found in the biopsy specimen. The change may be explained more by the weak OATP8 staining compared with that of alcohol-related liver cirrhosis than by malignant transformation into HCC. PMID:23607539

  20. Detection of high molecular weight amyloid serum protein complexes using Biological On-Line Tracer Sedimentation (BOLTS)

    OpenAIRE

    Kingsbury, Jonathan S.; Laue, Thomas M.; Chase, Susan F.; Connors, Lawreen H.

    2012-01-01

    The systemic amyloidoses are a rare, but deadly class of protein folding disorders with significant unmet diagnostic and therapeutic needs. The current model for symptomatic amyloid progression includes a causative role for soluble toxic aggregates as well as for the fibrillar tissue deposits. Although much research is focused on elucidating the potential mechanism of aggregate toxicity, evidence to support their existence in vivo has been limited. We report the use of a technique we have ter...

  1. Serum amyloid P inhibits dermal wound healing

    Science.gov (United States)

    The repair of open wounds depends on granulation tissue formation and contraction, which is primarily mediated by myofibroblasts. A subset of myofibroblasts originates from bone-marrow-derived monocytes which differentiate into fibroblast-like cells called fibrocytes. Serum amyloid P (SAP) inhibits ...

  2. Evaluation of intrathecal serum amyloid P (SAP) and C-reactive protein (CRP) synthesis in Alzheimer's disease with the use of index values.

    Science.gov (United States)

    Mulder, Sandra D; Hack, C Erik; van der Flier, Wiesje M; Scheltens, Philip; Blankenstein, Marinus A; Veerhuis, Robert

    2010-01-01

    Serum amyloid P (SAP) and C-reactive protein (CRP) are proteins involved in innate immunity. The expression of SAP and CRP is increased in Alzheimer's disease (AD) brain tissue, compared to healthy controls. Although both proteins are found in cerebrospinal fluid (CSF), their origin is unclear. We investigated if increased local production of SAP and CRP in AD brain results in higher levels in CSF with the use of index values. To study this, SAP, CRP, and albumin levels were determined in CSF and serum samples of 30 control (65 ± 11 years; 57% female) and 140 AD subjects (65 ± 9 years; 53% female). To correct for inter-individual differences in protein diffusion from blood to CSF, quotients (Q =CSF/serum) of SAP, CRP, and albumin and index values (Qprotein/Qalb) were calculated. The results showed no significant differences in SAP and CRP index values between control and AD subjects, although eight percent of individual AD patients showed evidence of intrathecal SAP or CRP production using the Reiber hyperbolic model. Interestingly, the SAP index value was much lower than expected, based on its molecular size. In conclusion, these data suggest that local production of SAP and CRP in the AD brain does not substantially contribute to the CSF levels. PMID:20930309

  3. Effects of Chinese herbal medicine Guanxinkang on lipid metabolism and serum C-reactive protein, amyloid A protein, and fibrinogen in apolipoprotein E-knockout mice with atherosclerosis

    Directory of Open Access Journals (Sweden)

    Mei-jiao Mao

    2011-03-01

    Full Text Available Objective: To observe the effects of Guanxinkang (GXK decoction, a compound traditional Chinese herbal medicine, on serum lipids and apolipoprotein A ? (ApoA ?, apolipoprotein B (ApoB, apolipoprotein E (ApoE, C-reactive protein (CRP, serum amyloid A protein (SAA and fibrinogen (Fbg concentrations of ApoE-knockout mice with atherosclerosis, and to explore the mechanism of GXK decoction in anti-atherosclerosis.Methods: Seventy 6-week-old ApoE-knockout mice receiving a high-cholesterol diet were used to induce atherosclerosis and were randomly divided into 5 groups: untreated group, simvastatin group and low- (drug concentration is 0.864 g/mL, medium- (1.728 g/mL, and high-dose (3.456 g/mL GXK groups. Another fourteen 6-week-old C57BL/6J mice were used as the normal control. Two 12-week-old mice were randomly selected from the normal control and the ApoE-knockout mice respectively to observe vulnerable plaque in the mouse’s aortic by hematoxylin-eosin staining. Blood was collected from venous plexus of eye socket after gavage of corresponding drugs once daily for 8 weeks continuously, and then the serum was separated. Triglyceride (TAG and total cholesterol (TC were measured by enzyme-coupled assay; low-density lipoprotein cholesterol (LDL-C and high-density lipoprotein cholesterol (HDL-C were measured by selective precipitation method. Serum levels of ApoA ? and ApoB were determined by turbidimetry. Double-antibody sandwich enzyme-linked immunosorbent assay was used to detect ApoE, CRP, SAA and Fbg concentrations in serum.Results: Compared with the normal control group, the levels of serum TC, TAG, LDL-C, ApoB, CRP, SAA and Fbg in the untreated group were increased (P<0.05, and the serum concentrations of HDL-C, ApoA ? and ApoE in the untreated group were decreased (P<0.05. After treatment, GXK decoction and simvastatin improved the dyslipidemia by increasing the concentrations of ApoA ? and HDL-C and decreasing the concentrations of TC, TAG, LDL-C, ApoB, CRP, SAA and Fbg (P<0.05. The high-dose GXK decoction had the most marked effects on SAA and Fbg and the serum lipids compared with the low-dose and medium-dose GXK and simvastatin.Conclusion: GXK decoction may not only provide an active effect on hyperlipidemia, but also down-regulate the levels of serum CRP, SAA and Fbg. GXK decoction exerts an anti-atherosclerosis effect in ApoE-knockout mice.

  4. 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.)

  5. A prospective study of the sensitivity, specificity and diagnostic performance of soluble intercellular adhesion molecule 1, highly sensitive C-reactive protein, soluble E-selectin and serum amyloid A in the diagnosis of neonatal infection

    OpenAIRE

    Gallimore J Ruth; Gabriel Vanessa; Edgar J David M; McMillan Stanley A; Grant Judith

    2010-01-01

    Abstract Background Diagnosis of neonatal infection is difficult, because of it's non-specific clinical presentation and the lack of reliable diagnostic tests. The purpose of this study was to examine the potential diagnostic value of serum soluble intercellular adhesion molecule-1 (sICAM-1), soluble E-selectin (sE-selectin), highly sensitive C-reactive protein (hsCRP) and serum amyloid A (SAA) measurements, both individually and in combination in the setting of a neonatal intensive care unit...

  6. Native human serum amyloid P component is a single pentamer

    DEFF Research Database (Denmark)

    SØrensen, Inge Juul; Andersen, Ove

    1995-01-01

    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 ofdecamers. However, the decamer form of SAP reversed to single pentamers when purified SAP was incorporated into SAP-depleted serum.

  7. Amyloid Aggregation and Membrane Disruption by Amyloid Proteins

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    Ramamoorthy, Ayyalusamy

    2013-03-01

    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.

  8. An in vitro study on neuroprotective effects of serum containing Gengnianchun decoction and its main monomers against amyloid beta protein-induced cellular toxicity

    Directory of Open Access Journals (Sweden)

    Wen-jun WANG

    2010-01-01

    Full Text Available Obiective: To observe the effects of serum containing Gengnianchun (GNC decoction, a compound traditional Chinese herbal medicine, and its monomers (paeoniflorin, berberine, timosaponin A-? and icariin on neurotoxicity in PC12 cells induced by amyloid beta-protein (A?.?Methods: Injury of PC12 cells was induced by in incubating with A?25-35 in vitro.? Ovariectomized rats were intragastrically administered with GNC decoction twice daily for 5 days and then sera were obtained. Different concentrations of serum containing GNC decoction and its main monomers including paeoniflorin, berberine, timosaponin and icariine and the monomer mixtures were cultured with PC12 cells to determine the best concentration of the drugs by methyl thiazolyl tetrazolium (MTT method. The effective concentration of A?25-35 was detemined by culturing PC12 cells with different concentrations of A?25-35. Then, the activity of PC12 cells with A?25-35-induced injury was observed with MTT method. Cellular morphological change was observed by phase contrast microscopy. Flow cytometry and fluorescence microscopy were employed to observe the A?25-35-induced early apoptosis of PC12 cells.?Results: After A?25-35 induction, the PC12 cells were fewer in number, less viable with shrinked cel1 body, many fragments, adhered less and nuclei shrinked. The cell proliferation was inhibited by A?25-35 concentration- and time?dependently. A?25-35 at concentration of 20 ?mol/ L was selected to construct the Alzheimer's disease model ?in vitro.? The sera containing GNC decoction could reinforce PC12 cell activity, and concentration at 20% was better than other concentrations after 24-, 48- and 72-hour culture. The 20% serum containing GNC decoction, 0.1 ?mol/L berberin and monomer mixture 3 including 1 mg/mL paeoniflorin, 1 ?mol/L berberine, 1 ?mol/L timosaponin and 1 ng/mL icariine could antagonize neurotoxicity induced by A?25-35. Moreover, they could inhibit A?25-35-induced early apoptosis of PC12 cells, with the effect of 20% serum containing GNC decoction better than 0.1 ?mol/L berberine and monomer mixture 3.Conclusion: Serum containing GNC decoction at 20% concentration has the potential neuroprotective effect on A?-induced cellular impairment. The serum containing GNC decoction was found to be stronger in action than the main monomers.

  9. Amyloid ? protein and Alzheimer disease

    OpenAIRE

    Square, D

    1997-01-01

    Amyloid beta protein is predominant in senile plaques, the neuropathologic hallmarks of Alzheimer disease. Researchers in Winnipeg have shown that this protein can overstimulate certain hydrolytic enzymes to break down the phospholipid building blocks of the brain-cell wall. They speculate that the abnormal destruction of phospholipids gradually drains the energy resources a neuron uses to rebuild its membrane. As neurons "burn out," the brain loses its ability to function normally. In view o...

  10. Human serum amyloid P is a multispecific adhesive protein whose ligands include 6-phosphorylated mannose and the 3-sulphated saccharides galactose, N-acetylgalactosamine and glucuronic acid.

    OpenAIRE

    Loveless, R. W.; Floyd-o Sullivan, G.; Raynes, J. G.; Yuen, C. T.; Feizi, T.

    1992-01-01

    Carbohydrate recognition by amyloid P component from human serum has been investigated by binding experiments using several glycosaminoglycans, polysaccharides and a series of structurally defined neoglycolipids and natural glycolipids. Two novel classes of carbohydrate ligands have been identified. The first is 6-phosphorylated mannose as found on lysosomal hydrolases, and the second is the 3-sulphated saccharides galactose, N-acetyl-galactosamine and glucuronic acid as found on sulphatide a...

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

    Energy Technology Data Exchange (ETDEWEB)

    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

    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.

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

    International Nuclear Information System (INIS)

    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 60Co ?-rays (0.55 Gy min-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 ?1 Gy and ?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 conventionalomplementary approach to conventional biodosimetry for early assessment of radiation exposures and may also contribute as diagnostic indices in the medical management of radiation accidents.

  13. Transthyretin Sequesters Amyloid ? Protein and Prevents Amyloid Formation

    Science.gov (United States)

    Schwarzman, Alexander L.; Gregori, Luisa; Vitek, Michael P.; Lyubski, Sergey; Strittmatter, Warren J.; Enghilde, Jan J.; Bhasin, Ramaninder; Silverman, Josh; Weisgraber, Karl H.; Coyle, Patricia K.; Zagorski, Michael G.; Talafous, Joseph; Eisenberg, Moises; Saunders, Ann M.; Roses, Allen D.; Goldgaber, Dmitry

    1994-08-01

    The cardinal pathological features of Alzheimer disease are depositions of aggregated amyloid ? protein (A?) in the brain and cerebrovasculature. However, the A? is found in a soluble form in cerebrospinal fluid in healthy individuals and patients with Alzheimer disease. We postulate that sequestration of A? precludes amyloid formation. Failure to sequester A? in Alzheimer disease may result in amyloidosis. When we added A? 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? in vitro, were not observed in cerebrospinal fluid. Additional in vitro studies showed that both purified transthyretin and apolipoprotein E prevent amyloid formation.

  14. Regulation of Amyloid ?-Protein Precursor by Phosphorylation and Protein Interactions*

    OpenAIRE

    Suzuki, Toshiharu; Nakaya, Tadashi

    2008-01-01

    Amyloid ?-protein precursor (APP), a type I membrane protein, is cleaved by primary ?-or ?-secretase and secondary ?-secretase. Cleavage of APP by ?- and ?-secretases generates amyloid ?-protein, the main constituent of the cerebrovascular amyloid that accompanies Alzheimer disease. The generation and aggregation of amyloid ?-protein in the brain are believed to be a primary cause of Alzheimer disease pathogenesis, and indeed, early onset Alzheimer disease is genetical...

  15. Transthyretin sequesters amyloid beta protein and prevents amyloid formation.

    OpenAIRE

    Schwarzman, A L; Gregori, L.; Vitek, M. P.; Lyubski, S; Strittmatter, W J; Enghilde, J J; Bhasin, R; Silverman, J.; Weisgraber, K H; Coyle, P. K.

    1994-01-01

    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 control...

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

    DEFF Research Database (Denmark)

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

    2007-01-01

    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.

  17. Avaliação da proteína amilóide A sérica na atividade clínica da artrite reumatóide / Evaluation of the serum amyloid A protein in the rheumatoid arthritis clinical activity

    Scientific Electronic Library Online (English)

    Carlos Roberto Machado, Gayer; Geraldo da Rocha Castelar, Pinheiro; Carlos Augusto Ferreira de, Andrade; Sérgio Miranda, Freire; Marsen Garcia Pinto, Coelho.

    2003-08-01

    Full Text Available A artrite reumatóide (AR) é uma doença auto-imune, crônica, caracterizada pelo comprometimento inflamatório das articulações sinoviais periféricas. A proteína amilóide A sérica (SAA) é uma das principais proteínas de fase aguda (PFA), porém seu uso na rotina do laboratório clínico ainda é pouco difu [...] ndido. OBJETIVO: O objetivo deste trabalho foi analisar a utilidade da SAA na avaliação da atividade clínica da AR. MÉTODOS: Foram estudados 113 pacientes com AR, diagnosticados segundo os critérios do Colégio Americano de Reumatologia. Para a caracterização da atividade de doença, foi utilizado o Índice de Atividade de Doença (IAD), proposto pela Liga Européia Contra o Reumatismo. RESULTADOS: A SAA apresentou correlação positiva, estatisticamente significativa, com a proteína C-Reativa (PCR), tanto como a ?-1-glicoproteína ácida (AGP), quanto com o IAD. Nossos resultados demonstraram que a SAA apresentou, particularmente, uma maior sensibilidade na determinação da atividade inflamatória da AR, em comparação às outras PFA. Apresentou, também, uma boa capacidade de discriminar os grupos de atividade moderada e alta do IAD. Como o IAD não mede unicamente o componente inflamatório da AR, a dosagem de uma PFA é de grande utilidade para a caracterização da atividade dessa enfermidade. CONCLUSÕES: Os resultados deste estudo sugerem que a SAA pode ser de grande valor na determinação da atividade inflamatória da AR. Abstract in english Rheumatoid arthritis (RA) is a systemic autoimmune disease, with chronic inflammation of synovial peripheral joints as the most prominent feature. The serum amyloid A (SAA) is one of the major acute phase proteins (APP), however its use in the clinical laboratory routine is uncommon. OBJECTIVE: The [...] aim of this work was to analyze the usefulness of SAA in the evaluation of RA clinical activity. METHODS: We studied 113 patients diagnosed with RA according to the criteria of the American College of Rheumatology. The disease activity was evaluated by the Disease Activity Score (DAS) according to the European League Against Rheumatism. RESULTS: SAA presented positive correlation, statistically significant, with C-reactive protein (CRP), as well as ?-1-acid glycoprotein (AGP) and DAS. The results demonstrated that SAA presented a higher sensibility in relation to other APP in the determination of the inflammatory activity on RA patients. SAA also shows a good capability to discriminate the groups of moderate and high activity of DAS. As DAS doesn't measure only the inflammatory behavior of RA, the determination of APP is of great usefulness for the activity characterization of this illness. CONCLUSIONS: Our data suggest that SAA can be of great value in the determination of the RA inflammatory activity.

  18. Binding of human proteins to amyloid-? protofibrils.

    Science.gov (United States)

    Rahman, M Mahafuzur; Zetterberg, Henrik; Lendel, Christofer; Härd, Torleif

    2015-03-20

    The progressive neurodegeneration in Alzheimer's disease is believed to be linked to the presence of prefibrillar aggregates of the amyloid-? (A?) peptide in the brain. The exact role of these aggregates in the disease pathology is, however, still an open question. Any mechanism by which oligomeric A? may cause damage to neuronal cells must, in one way or another, involve interactions with other molecules. Here, we identify proteins in human serum and cerebrospinal fluid that bind to stable protofibrils formed by an engineered variant of A?42 (A?42CC). We find that the protofibrils attract a substantial number of protein binding partners. Many of the 101 identified proteins are involved in lipid transport and metabolism, the complement system, or in hemostasis. Binding of representative proteins from all of these groups with micromolar affinity was confirmed using surface plasmon resonance. In addition, binding of apolipoprotein E to the protofibrils with nanomolar affinity was demonstrated. We also find that aggregation of A? enhances protein binding, as lower amounts of proteins bind monomeric A?. Proteins that bind to A? protofibrils might contribute to biological effects in which these aggregates are involved. Our results therefore suggest that an improved understanding of the mechanisms by which A? causes cytotoxicity and neurodegeneration might be gained from studies carried out in biologically relevant matrices in which A?-binding proteins are present. PMID:25469473

  19. Transthyretin sequesters amyloid beta protein and prevents amyloid formation

    DEFF Research Database (Denmark)

    Schwarzman, A L; Gregori, L

    1994-01-01

    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.

  20. Ligand-binding sites in human serum amyloid P component

    DEFF Research Database (Denmark)

    Heegaard, N.H.H.; Heegaard, Peter M. H.

    1996-01-01

    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.

  1. Neuronal overexpression of mutant amyloid precursor protein results in prominent deposition of cerebrovascular amyloid

    OpenAIRE

    Calhoun, Michael E.; Burgermeister, Patrick; Phinney, Amie L.; Stalder, Martina; Tolnay, Markus; Wiederhold, Karl-heinz; Abramowski, Dorothee; Sturchler-pierrat, Christine; Sommer, Bernd; Staufenbiel, Matthias; Jucker, Mathias

    1999-01-01

    Transgenic mice that overexpress mutant human amyloid precursor protein (APP) exhibit one hallmark of Alzheimer’s disease pathology, namely the extracellular deposition of amyloid plaques. Here, we describe significant deposition of amyloid ? (A?) in the cerebral vasculature [cerebral amyloid angiopathy (CAA)] in aging APP23 mice that had striking similarities to that observed in human aging and Alzheimer’s disease. Amyloid deposition occurred preferentially in arterioles and capillarie...

  2. Multiphoton absorption in amyloid protein fibres

    Science.gov (United States)

    Hanczyc, Piotr; Samoc, Marek; Norden, Bengt

    2013-12-01

    Fibrillization of peptides leads to the formation of amyloid fibres, which, when in large aggregates, are responsible for diseases such as Alzheimer's and Parkinson's. Here, we show that amyloids have strong nonlinear optical absorption, which is not present in native non-fibrillized protein. Z-scan and pump-probe experiments indicate that insulin and lysozyme ?-amyloids, as well as ?-synuclein fibres, exhibit either two-photon, three-photon or higher multiphoton absorption processes, depending on the wavelength of light. We propose that the enhanced multiphoton absorption is due to a cooperative mechanism involving through-space dipolar coupling between excited states of aromatic amino acids densely packed in the fibrous structures. This finding will provide the opportunity to develop nonlinear optical techniques to detect and study amyloid structures and also suggests that new protein-based materials with sizable multiphoton absorption could be designed for specific applications in nanotechnology, photonics and optoelectronics.

  3. Serum Amyloid P Aids Complement-Mediated Immunity to Streptococcus pneumoniae

    OpenAIRE

    Yuste, J.; Botto, M.; Bottoms, S. E.; Brown, J. S.

    2007-01-01

    The physiological functions of the acute phase protein serum amyloid P ( SAP) component are not well defined, although they are likely to be important, as no natural state of SAP deficiency has been reported. We have investigated the role of SAP for innate immunity to the important human pathogen Streptococcus pneumoniae. Using flow cytometry assays, we show that SAP binds to S. pneumoniae, increases classical pathway-dependent deposition of complement on the bacteria, and improves the effici...

  4. Biofilm Inhibitors that Target Amyloid Proteins

    OpenAIRE

    Romero, Diego; Sanabria-Valentín, Edgardo; Vlamakis, Hera; Kolter, Roberto

    2013-01-01

    Bacteria establish stable communities, known as biofilms, that are resistant to antimicrobials. Biofilm robustness is due to the presence of an extracellular matrix, which for several species - among them Bacillus subtilis - includes amyloid-like protein fibers. In this work, we show that B. subtilis biofilms can be a simple and reliable tool for screening of molecules with anti-amyloid activity. We identified two molecules, AA-861 and parthenolide, which efficiently inhibited biofilms by pre...

  5. Human serum amyloid A genes are expressed in monocyte/macrophage cell lines.

    OpenAIRE

    Urieli-Shoval, S; Meek, R L; Hanson, R. H.; Eriksen, N.; Benditt, E. P.

    1994-01-01

    Serum amyloid A (apoSAA) is a family of proteins found, mainly associated with high density lipoproteins, in the blood plasma of mammals and at least one avian species, the Pekin duck. These proteins are present in small amounts under normal circumstances, but their concentration is capable of rising 100- to 1,000-fold in situations involving tissue injury or infection. Like classic acute phase proteins they are produced in the liver; however, expression of one of the apoSAA genes is known to...

  6. Multiple isoforms of the human pentraxin serum amyloid P component.

    DEFF Research Database (Denmark)

    SØrensen, Inge Juul; Andersen, Ove

    1995-01-01

    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.

  7. Divalent cation tolerance protein binds to ?-secretase and inhibits the processing of amyloid precursor protein?

    OpenAIRE

    Liu, Runzhong; Hou, Haibo; Yi, Xuelian; Wu, Shanwen; Zeng, Huan

    2013-01-01

    The deposition of amyloid-beta is a pathological hallmark of Alzheimer's disease. Amyloid-beta is derived from amyloid precursor protein through sequential proteolytic cleavages by ?-secretase (beta-site amyloid precursor protein-cleaving enzyme 1) and ?-secretase. To further elucidate the roles of beta-site amyloid precursor protein-cleaving enzyme 1 in the development of Alzheimer's disease, a yeast two-hybrid system was used to screen a human embryonic brain cDNA library for proteins direc...

  8. Conformational dynamics of amyloid proteins at the aqueous interface

    Science.gov (United States)

    Armbruster, Matthew; Horst, Nathan; Aoki, Brendy; Malik, Saad; Soto, Patricia

    2013-03-01

    Amyloid proteins is a class of proteins that exhibit distinct monomeric and oligomeric conformational states hallmark of deleterious neurological diseases for which there are not yet cures. Our goal is to examine the extent of which the aqueous/membrane interface modulates the folding energy landscape of amyloid proteins. To this end, we probe the dynamic conformational ensemble of amyloids (monomer prion protein and Alzheimer's Ab protofilaments) interacting with model bilayers. We will present the results of our coarse grain molecular modeling study in terms of the existence of preferential binding spots of the amyloid to the bilayer and the response of the bilayer to the interaction with the amyloid. Amyloid proteins is a class of proteins that exhibit distinct monomeric and oligomeric conformational states hallmark of deleterious neurological diseases for which there are not yet cures. Our goal is to examine the extent of which the aqueous/membrane interface modulates the folding energy landscape of amyloid proteins. To this end, we probe the dynamic conformational ensemble of amyloids (monomer prion protein and Alzheimer's Ab protofilaments) interacting with model bilayers. We will present the results of our coarse grain molecular modeling study in terms of the existence of preferential binding spots of the amyloid to the bilayer and the response of the bilayer to the interaction with the amyloid. NSF Nebraska EPSCoR First Award

  9. A prospective study of the sensitivity, specificity and diagnostic performance of soluble intercellular adhesion molecule 1, highly sensitive C-reactive protein, soluble E-selectin and serum amyloid A in the diagnosis of neonatal infection

    Directory of Open Access Journals (Sweden)

    Gallimore J Ruth

    2010-04-01

    Full Text Available Abstract Background Diagnosis of neonatal infection is difficult, because of it's non-specific clinical presentation and the lack of reliable diagnostic tests. The purpose of this study was to examine the potential diagnostic value of serum soluble intercellular adhesion molecule-1 (sICAM-1, soluble E-selectin (sE-selectin, highly sensitive C-reactive protein (hsCRP and serum amyloid A (SAA measurements, both individually and in combination in the setting of a neonatal intensive care unit. Methods 219 consecutive serum samples were taken from 149 infants undergoing sepsis work up in a neonatal intensive care unit. Clinical diagnosis was established in a prospective manner, blind to the results of the study measurements. Infants were classified by an experienced paediatrician as infected or not-infected, one week after presentation. Classification was based on clinical presentation, routine laboratory and radiological investigations and response to therapy. The infected group were sub-classified as (a culture positive infection or (b culture negative infection. sICAM-1, sE-selectin, hsCRP and SAA levels were determined from stored serum samples after diagnosis was established. Further sub-group analysis of results was undertaken according to early or late onset of infection and preterm or term status. Statistical analysis utilised Mann Whitney U test and ROC curve analysis. Results There were significantly increased serum levels of sICAM-1, hsCRP, E selectin (p Conclusions All four study measurements demonstrated some diagnostic value for neonatal infection however sICAM-1, hsCRP and sE-selectin demonstrated the highest NPV individually. The optimum diagnostic cut off level for hsCRP measurement in this study was much lower than currently used in routine clinical practice. Use of a combination of measurements enhanced diagnostic performance, demonstrating sensitivity of 90.3% and NPV of 91.3%. This study suggests there may be value in use of several of these markers, individually and in combination to assist in excluding neonatal infection. Further work is needed to confirm a specific role in the exclusion of early onset infection.

  10. Apple Procyanidins Suppress Amyloid ?-Protein Aggregation

    OpenAIRE

    Toda, Toshihiko; Sunagawa, Tadahiro; Kanda, Tomomasa; Tagashira, Motoyuki; Shirasawa, Takuji; Shimizu, Takahiko

    2011-01-01

    Procyanidins (PCs) are major components of the apple polyphenols (APs). We previously reported that treatment with PC extended the mean lifespan of Caenorhabditis elegans (Sunagawa et al., 2011). In order to estimate the neuroprotective effects of PC, we investigated the antiaggregative activity of PC on amyloid ?-protein (A?) aggregation, which is a pathological hallmark of Alzheimer's disease. We herein report that PC significantly suppressed A?42 aggregation and dissociated A?42 aggreg...

  11. Amyloid ?-sheet mimics that antagonize protein aggregation and reduce amyloid toxicity

    Science.gov (United States)

    Cheng, Pin-Nan; Liu, Cong; Zhao, Minglei; Eisenberg, David; Nowick, James S.

    2012-11-01

    The amyloid protein aggregation associated with diseases such as Alzheimer's, Parkinson's and type II diabetes (among many others) features a bewildering variety of ?-sheet-rich structures in transition from native proteins to ordered oligomers and fibres. The variation in the amino-acid sequences of the ?-structures presents a challenge to developing a model system of ?-sheets for the study of various amyloid aggregates. Here, we introduce a family of robust ?-sheet macrocycles that can serve as a platform to display a variety of heptapeptide sequences from different amyloid proteins. We have tailored these amyloid ?-sheet mimics (ABSMs) to antagonize the aggregation of various amyloid proteins, thereby reducing the toxicity of amyloid aggregates. We describe the structures and inhibitory properties of ABSMs containing amyloidogenic peptides from the amyloid-? peptide associated with Alzheimer's disease, ?2-microglobulin associated with dialysis-related amyloidosis, ?-synuclein associated with Parkinson's disease, islet amyloid polypeptide associated with type II diabetes, human and yeast prion proteins, and Tau, which forms neurofibrillary tangles.

  12. Serum amyloid A overrides Treg anergy via monocyte-dependent and Treg-intrinsic, SOCS3-associated pathways

    OpenAIRE

    Nguyen, Khoa D.; Macaubas, Claudia; Nadeau, Kari C.; Truong, Phi; Yoon, Taejin; Lee, Tzielan; Park, Jane L.; Mellins, Elizabeth D.

    2011-01-01

    The acute phase protein serum amyloid A (SAA) has been well characterized as an indicator of inflammation. Nevertheless, its functions in pro versus anti-inflammatory processes remain obscure. Here we provide unexpected evidences that SAA induces the proliferation of the tolerogenic subset of regulatory T cells (Treg). Intriguingly, SAA reverses Treg anergy via its interaction with monocytes to activate distinct mitogenic pathways in Treg but not effector T cells. This selective responsivenes...

  13. Statistical Mechanical Treatments of Protein Amyloid Formation

    CERN Document Server

    Schreck, John S

    2013-01-01

    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 amo...

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

    Directory of Open Access Journals (Sweden)

    Gonzalez-Velasquez Francisco J

    2009-04-01

    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.

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

    Directory of Open Access Journals (Sweden)

    Langen Ralf

    2008-10-01

    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.

  16. Neuronal overexpression of mutant amyloid precursor protein results in prominent deposition of cerebrovascular amyloid

    Science.gov (United States)

    Calhoun, Michael E.; Burgermeister, Patrick; Phinney, Amie L.; Stalder, Martina; Tolnay, Markus; Wiederhold, Karl-Heinz; Abramowski, Dorothee; Sturchler-Pierrat, Christine; Sommer, Bernd; Staufenbiel, Matthias; Jucker, Mathias

    1999-01-01

    Transgenic mice that overexpress mutant human amyloid precursor protein (APP) exhibit one hallmark of Alzheimer’s disease pathology, namely the extracellular deposition of amyloid plaques. Here, we describe significant deposition of amyloid ? (A?) in the cerebral vasculature [cerebral amyloid angiopathy (CAA)] in aging APP23 mice that had striking similarities to that observed in human aging and Alzheimer’s disease. Amyloid deposition occurred preferentially in arterioles and capillaries and within individual vessels showed a wide heterogeneity (ranging from a thin ring of amyloid in the vessel wall to large plaque-like extrusions into the neuropil). CAA was associated with local neuron loss, synaptic abnormalities, microglial activation, and microhemorrhage. Although several factors may contribute to CAA in humans, the neuronal origin of transgenic APP, high levels of A? in cerebrospinal fluid, and regional localization of CAA in APP23 mice suggest transport and drainage pathways rather than local production or blood uptake of A? as a primary mechanism underlying cerebrovascular amyloid formation. APP23 mice on an App-null background developed a similar degree of both plaques and CAA, providing further evidence that a neuronal source of APP/A? is sufficient to induce cerebrovascular amyloid and associated neurodegeneration. PMID:10570203

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

    OpenAIRE

    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

    2010-01-01

    Accumulation of amyloid fibrils in the viscera and connective tissues causes systemic amyloidosis, which is responsible for about one per thousand deaths in developed countries1. Localised amyloid can also be very serious, 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 present1. There is therefore a major unmet medical need ...

  18. Reference intervals for acute phase protein and serum protein electrophoresis values in captive Asian elephants (Elephas maximus).

    Science.gov (United States)

    Isaza, Ramiro; Wiedner, Ellen; Hiser, Sarah; Cray, Carolyn

    2014-09-01

    Acute phase protein (APP) immunoassays and serum protein electrophoresis (SPEP) are assays for evaluating the inflammatory response and have use as diagnostic tools in a variety of species. Acute phase proteins are markers of inflammation that are highly conserved across different species while SPEP separates and quantifies serum protein fractions based on their physical properties. In the current study, serum samples from 35 clinically healthy Asian elephants (Elephas maximus) were analyzed using automated assays for C-reactive protein, serum amyloid A, and haptoglobin and SPEP. Robust methods were used to generate reference intervals for the APPs: C-reactive protein (1.3-12.8 mg/l), serum amyloid A (0-47.5 mg/l), and haptoglobin (0-1.10 mg/ml). In addition, SPEP was performed on these samples to establish reference intervals for each protein fraction. A combination of APPs and SPEP measurements are valuable adjunctive diagnostic tools in elephant health care. PMID:25057161

  19. SERF Protein Is a Direct Modifier of Amyloid Fiber Assembly

    Directory of Open Access Journals (Sweden)

    S. Fabio Falsone

    2012-08-01

    Full Text Available The inherent cytotoxicity of aberrantly folded protein aggregates contributes substantially to the pathogenesis of amyloid diseases. It was recently shown that a class of evolutionary conserved proteins, called MOAG-4/SERF, profoundly alter amyloid toxicity via an autonomous but yet unexplained mode. We show that the biological function of human SERF1a originates from its atypical ability to specifically distinguish between amyloid and nonamyloid aggregation. This inherently unstructured protein directly affected the aggregation kinetics of a broad range of amyloidogenic proteins in vitro, while being inactive against nonamyloid aggregation. A representative biophysical analysis of the SERF1a:?-synuclein (aSyn complex revealed that the amyloid-promoting activity resulted from an early and transient interaction, which was sufficient to provoke a massive increase of soluble aSyn amyloid nucleation templates. Therefore, the autonomous amyloid-modifying activity of SERF1a observed in living organisms relies on a direct and dedicated manipulation of the early stages in the amyloid aggregation pathway.

  20. Micropurification techniques in the analysis of amyloid proteins

    OpenAIRE

    Kaplan, B.; Shtrasburg, S.; Pras, M.

    2003-01-01

    This review describes the different microtechniques developed for the extraction and purification of amyloid proteins from small specimens of fresh and formalin fixed tissues. These procedures differ with respect to solvent type, extraction conditions, and protein purification strategy. The advantages and disadvantages of the different microtechniques are discussed by taking into consideration tissue type (fresh of fixed) and size, amyloid type, and its content in the tissue. The review demon...

  1. Protein corona composition of gold nanoparticles/nanorods affects amyloid beta fibrillation process

    Science.gov (United States)

    Mirsadeghi, Somayeh; Dinarvand, Rassoul; Ghahremani, Mohammad Hossein; Hormozi-Nezhad, Mohammad Reza; Mahmoudi, Zohreh; Hajipour, Mohammad Javad; Atyabi, Fatemeh; Ghavami, Mahdi; Mahmoudi, Morteza

    2015-03-01

    Protein fibrillation process (e.g., from amyloid beta (A?) and ?-synuclein) is the main cause of several catastrophic neurodegenerative diseases such as Alzheimer's and Parkinson diseases. During the past few decades, nanoparticles (NPs) were recognized as one of the most promising tools for inhibiting the progress of the disease by controlling the fibrillation kinetic process; for instance, gold NPs have a strong capability to inhibit A? fibrillations. It is now well understood that a layer of biomolecules would cover the surface of NPs (so called ``protein corona'') upon the interaction of NPs with protein sources. Due to the fact that the biological species (e.g., cells and amyloidal proteins) ``see'' the protein corona coated NPs rather than the pristine coated particles, one should monitor the fibrillation process of amyloidal proteins in the presence of corona coated NPs (and not pristine coated ones). Therefore, the previously obtained data on NPs effects on the fibrillation process should be modified to achieve a more reliable and predictable in vivo results. Herein, we probed the effects of various gold NPs (with different sizes and shapes) on the fibrillation process of A? in the presence and absence of protein sources (i.e., serum and plasma). We found that the protein corona formed a shell at the surface of gold NPs, regardless of their size and shape, reducing the access of A? to the gold inhibitory surface and, therefore, affecting the rate of A? fibril formation. More specifically, the anti-fibrillation potencies of various corona coated gold NPs were strongly dependent on the protein source and their concentrations (10% serum/plasma (simulation of an in vitro milieu) and 100% serum/plasma (simulation of an in vivo milieu)).Protein fibrillation process (e.g., from amyloid beta (A?) and ?-synuclein) is the main cause of several catastrophic neurodegenerative diseases such as Alzheimer's and Parkinson diseases. During the past few decades, nanoparticles (NPs) were recognized as one of the most promising tools for inhibiting the progress of the disease by controlling the fibrillation kinetic process; for instance, gold NPs have a strong capability to inhibit A? fibrillations. It is now well understood that a layer of biomolecules would cover the surface of NPs (so called ``protein corona'') upon the interaction of NPs with protein sources. Due to the fact that the biological species (e.g., cells and amyloidal proteins) ``see'' the protein corona coated NPs rather than the pristine coated particles, one should monitor the fibrillation process of amyloidal proteins in the presence of corona coated NPs (and not pristine coated ones). Therefore, the previously obtained data on NPs effects on the fibrillation process should be modified to achieve a more reliable and predictable in vivo results. Herein, we probed the effects of various gold NPs (with different sizes and shapes) on the fibrillation process of A? in the presence and absence of protein sources (i.e., serum and plasma). We found that the protein corona formed a shell at the surface of gold NPs, regardless of their size and shape, reducing the access of A? to the gold inhibitory surface and, therefore, affecting the rate of A? fibril formation. More specifically, the anti-fibrillation potencies of various corona coated gold NPs were strongly dependent on the protein source and their concentrations (10% serum/plasma (simulation of an in vitro milieu) and 100% serum/plasma (simulation of an in vivo milieu)). Electronic supplementary information (ESI) available: Full characterization results of the nanoparticles, protein corona, and fibrillation process. See DOI: 10.1039/c4nr06009a

  2. ?-amyloid oligomers and cellular prion protein in Alzheimer's disease

    OpenAIRE

    Gunther, Erik C.; Strittmatter, Stephen M

    2009-01-01

    Prefibrillar oligomers of the ?-amyloid peptide (A?) are recognized as potential mediators of Alzheimer's disease (AD) pathophysiology. Deficits in synaptic function, neurotoxicity, and the progression of AD have all been linked to the oligomeric A? assemblies rather than to A? monomers or to amyloid plaques. However, the molecular sites of A? oligomer action have remained largely unknown. Recently, the cellular prion protein (PrPC) has been shown to act as a functional receptor for A? ...

  3. Haptoglobin and serum amyloid a in subacute ruminal acidosis in goats

    Directory of Open Access Journals (Sweden)

    F.H.D. González

    2010-12-01

    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.

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

    DEFF Research Database (Denmark)

    Pihl, Tina Holberg; Andersen, Pia Haubro

    2013-01-01

    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 

  5. Serum protein capillary electrophoresis and measurement of acute phase proteins in a captive cheetah (Acinonyx jubatus) population

    DEFF Research Database (Denmark)

    Depauw, Sarah; Delanghe, Joris

    2014-01-01

    Renal and gastrointestinal pathologies are widespread in the captive cheetah (Acinonyx jubatus) population but are often diagnosed at a late stage, because diagnostic tools are limited to the evaluation of clinical signs or general blood examination. Presently, no data are available on serum proteins and acute-phase proteins in cheetahs during health or disease, although they might be important to improve health monitoring. This study aimed to quantify serum proteins by capillary electrophoresis in 80 serum samples from captive cheetahs, categorized according to health status and disease type. Moreover, serum amyloid A concentrations were measured via a turbidimetric immunoassay validated in domestic cats, whereas haptoglobin and C-reactive protein were determined by non-species-specific functional tests. Cheetahs classified as healthy had serum protein and acute phase protein concentrations within reference ranges for healthy domestic cats. In contrast, unhealthy cheetahs had higher (P < 0.001) serum amyloidA, alpha2-globulin, and haptoglobin concentrations compared with the healthy subgroup. Moreover, serum amyloid A (P = 0.020), alpha2-globulin (P < 0.001) and haptoglobin (P = 0.001) concentrations in cheetahs suffering from chronic kidney disease were significantly greater compared to the reportedly healthy cheetahs. Our study indicates that serum proteins in the cheetah can be analyzed by routine capillary electrophoresis, whereas acute-phase proteins can be measured using available immunoassays or non-species-specific techniques, which are also likely to be applicable in other exotic felids. Moreover, results suggest that serum amyloid A and haptoglobin are important acute-phase proteins in the diseased cheetah and highlight the need to evaluate their role as early-onset markers for disease.

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

    DEFF Research Database (Denmark)

    Nybo, Mads; Nielsen, E H

    1999-01-01

    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.

  7. The role of serum amyloid A and sphingosine-1-phosphate on high-density lipoprotein functionality.

    Science.gov (United States)

    Prüfer, Nicole; Kleuser, Burkhard; van der Giet, Markus

    2015-06-01

    The high-density lipoprotein (HDL) is one of the most important endogenous cardiovascular protective markers. HDL is an attractive target in the search for new pharmaceutical therapies and in the prevention of cardiovascular events. Some of HDL's anti-atherogenic properties are related to the signaling molecule sphingosine-1-phosphate (S1P), which plays an important role in vascular homeostasis. However, for different patient populations it seems more complicated. Significant changes in HDL's protective potency are reduced under pathologic conditions and HDL might even serve as a proatherogenic particle. Under uremic conditions especially there is a change in the compounds associated with HDL. S1P is reduced and acute phase proteins such as serum amyloid A (SAA) are found to be elevated in HDL. The conversion of HDL in inflammation changes the functional properties of HDL. High amounts of SAA are associated with the occurrence of cardiovascular diseases such as atherosclerosis. SAA has potent pro-atherogenic properties, which may have impact on HDL's biological functions, including cholesterol efflux capacity, antioxidative and anti-inflammatory activities. This review focuses on two molecules that affect the functionality of HDL. The balance between functional and dysfunctional HDL is disturbed after the loss of the protective sphingolipid molecule S1P and the accumulation of the acute-phase protein SAA. This review also summarizes the biological activities of lipid-free and lipid-bound SAA and its impact on HDL function. PMID:25252751

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

    DEFF Research Database (Denmark)

    Kania, Per Walter; Chettri, Jiwan Kumar

    2014-01-01

    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.

  9. Serum amyloid A-containing human high density lipoprotein 3. Density, size, and apolipoprotein composition.

    Science.gov (United States)

    Coetzee, G A; Strachan, A F; van der Westhuyzen, D R; Hoppe, H C; Jeenah, M S; de Beer, F C

    1986-07-25

    Serum amyloid A protein (apo-SAA), an acute phase reactant, is an apolipoprotein of high density lipoproteins (HDL), in particular the denser subpopulation HDL3. The structure of HDL3 isolated from humans affected by a variety of severe disease states was investigated with respect to density, size, and apolipoprotein composition, using density gradient ultracentrifugation, gradient gel electrophoresis, gel filtration, and solid phase immunoadsorption. Apo-SAA was present in HDL particles in increasing amounts as particle density increased. Apo-SAA-containing HDL3 had bigger radii than normal HDL3 of comparable density. Purified apo-SAA associated readily with normal HDL3 in vitro, giving rise to particles containing up to 80% of their apoproteins as apo-SAA. The addition of apo-SAA resulted in a displacement of apo-A-I and an increase in particle size. Acute phase HDL3 represented a mixture of particles, polydisperse with respect to apolipoprotein content; for example, some particles were isolated that contained apo-A-I, apo-A-II, and apo-SAA, whereas others contained apo-A-I and apo-SAA but no apo-A-II. We conclude that apo-SAA probably associates in the circulation of acute phase patients with existing HDL particles, causing the remodeling of the HDL shell to yield particles of bigger size and higher density that are relatively depleted of apo-A-I. PMID:3525531

  10. Serum amyloid P ameliorates radiation-induced oral mucositis and fibrosis

    Directory of Open Access Journals (Sweden)

    Murray Lynne A

    2010-07-01

    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.

  11. Heterocomplexes of Mannose-binding Lectin and the Pentraxins PTX3 or Serum Amyloid P Component Trigger Cross-activation of the Complement System*

    OpenAIRE

    Ma, Ying Jie; Doni, Andrea; Skjoedt, Mikkel-Ole; Honoré, Christian; Arendrup, Maiken; Mantovani, Alberto; Garred, Peter

    2010-01-01

    The long pentraxin 3 (PTX3), serum amyloid P component (SAP), and C-reactive protein belong to the pentraxin family of pattern recognition molecules involved in tissue homeostasis and innate immunity. They interact with C1q from the classical complement pathway. Whether this also occurs via the analogous mannose-binding lectin (MBL) from the lectin complement pathway is unknown. Thus, we investigated the possible interaction between MBL and the pentraxins. We report that MBL bound PTX3 and SA...

  12. Predictive values of serum amyloid-A and CRP for infection in febrile neutropenic cancer patients

    Directory of Open Access Journals (Sweden)

    Ay?e Bat?rel

    2014-12-01

    Full Text Available Objectives: To evaluate predictive values of serum amyloid A (SAA and C-reactive protein (CRP for infection and mor­tality in patients with febrile neutropenia (FEN. Methods: Daily measurement of serum SAA and CRP levels of patients during antibiotherapy for FEN. Results: Sixty-five FEN episodes of 52 patients were evaluated. Median CRP and SAA levels on 1st day of FEN were 137 mg/L (23-420 mg/L and 547 mg/L (11-1660 mg/L, respectively. For detection of infection of infection the sensitivity, positive predictive value (PPV, and negative predictive value (NPV of SAA at a level of >80 mg/L were as 100%, 48% and 100%. Whilethe sensitivity, PPV, and NPV of CRP at a level of >50mg/L were as 86%, 47% and 60%, respectively. Predictive values of initial SAA and CRP levels for infection didn’t differ significantly (CRP: p=0.24, SAA: p=0.39. SAA and CRP levels on the last day of FEN course were significant for infection and mortality (for infection: p=0.003 for CRP and p=0.026 for SAA; for mortality: p<0.001 for CRP and p=0.021 for SAA. Both initial and daily SAA and CRP levels correlated with each other positively and statistically significantly (p<0.001. The area under the curve (AUC on the re­ceiver operating character (ROC curve for CRP and SAA were 0.72 (p=0.003, 95% CI: 0.59-0.86 and 0.68 (p=0.19, 95% CI: 0.54-0.82, respectively. Conclusions: Despite low predictive values in decision of initial therapy, these parameters would be helpful in decision of modification and evaluation of response to therapy. J Microbiol Infect Dis 2014; 4(4: 128-135

  13. Monoclonal antibodies to amyloid subunit proteins for in vivo radioimmunodetection of amyloid diseases

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.; Meinken, G.E.; Gorevic, P.; Atkins, H.L.; Fand, I.; Marshall, J.; McNally, W.; Muller, D.; Wood, D.

    1984-01-01

    Amyloid fibrils of systemic amyloidosis are low molecular weight subunit proteins with poor immunogenicity and a tendency to polymerize. Antibodies to these proteins are useful for the detection of amyloid deposits in-situ. The extracellular location of amyloid deposits and proximity to congophilic angiopathy suggest the potential of labeled monocional antibodies (MAb) for in vivo radioimmunodetection. The authors tested feasibility of this approach using two rat MAbs to mouse AA protein in casein-induced amyloidosis, a model system for human secondary amyloidosis. The antibodies were labeled with I-125, I-123, and In-111 with good specificity retention. Amyloidotic mice were pretreated with 50 ..mu..g colchicine ip 3 hr before receiving radioiodinated MAb via the tall vein. Controls included injection of MAb to normal mice and of labeled polyclonal normal rat IgG (pIg) into amyloidotic and control mice. Blood clearance of MAb was faster in amyloidotic than control groups. Fractionation studies showed that both MAb and pIg were uncomplexed. Studies up to 96 hr showed specific and high uptake at sites of amyloid deposition (saline perfused liver, spleen, kidney. Specific localization was confirmed by whole body autoradiography (I-125, 20 ..mu..Ci/animal; 50 ..mu..g MAb) and by external imaging (I-123, 200 ..mu..Ci/animal, 10-15 ..mu..g MAb) of amyloidotic mice studied at 4-72 hr. Amyloidotic animals showed perifollicular localization in spleen, periportal in liver, and glomerular in kidney; scans of controls showed diffuse early washout. These results document the feasibility of using MAbs to fibril subunit proteins for the in vivo detection and therapy of amyloidosis.

  14. Monoclonal antibodies to amyloid subunit proteins for in vivo radioimmunodetection of amyloid diseases

    International Nuclear Information System (INIS)

    Amyloid fibrils of systemic amyloidosis are low molecular weight subunit proteins with poor immunogenicity and a tendency to polymerize. Antibodies to these proteins are useful for the detection of amyloid deposits in-situ. The extracellular location of amyloid deposits and proximity to congophilic angiopathy suggest the potential of labeled monocional antibodies (MAb) for in vivo radioimmunodetection. The authors tested feasibility of this approach using two rat MAbs to mouse AA protein in casein-induced amyloidosis, a model system for human secondary amyloidosis. The antibodies were labeled with I-125, I-123, and In-111 with good specificity retention. Amyloidotic mice were pretreated with 50 ?g colchicine ip 3 hr before receiving radioiodinated MAb via the tall vein. Controls included injection of MAb to normal mice and of labeled polyclonal normal rat IgG (pIg) into amyloidotic and control mice. Blood clearance of MAb was faster in amyloidotic than control groups. Fractionation studies showed that both MAb and pIg were uncomplexed. Studies up to 96 hr showed specific and high uptake at sites of amyloid deposition (saline perfused liver, spleen, kidney. Specific localization was confirmed by whole body autoradiography (I-125, 20 ?Ci/animal; 50 ?g MAb) and by external imaging (I-123, 200 ?Ci/animal, 10-15 ?g MAb) of amyloidotic mice studied at 4-72 hr. Amyloidotic animals showed perifollicular localization in spleen, periportal in liver, and glomerular in kidney; scans of controls showed diffuse early washout. These results document the feasibility of using MAbs to fibril subunit proteins for the in vivo detection and therapy of amyloidosis

  15. AMYPdb: A database dedicated to amyloid precursor proteins

    Directory of Open Access Journals (Sweden)

    Delamarche Christian

    2008-06-01

    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.

  16. Maldi-Tof /Tof-MS Reveals Elevated Serum Haptoglobin and Amyloid A in Behcet's Disease

    OpenAIRE

    Mao, L.; Dong, H; Yang, P.; Zhou, H.; Huang, X.; Lin, X.; Kijlstra, A

    2008-01-01

    Behcet¿s disease (BD) is a multisystemic autoimmune disease with unclear etiology and pathogenesis. To screen aberrant serum proteins in BD, serum samples were obtained from eight male BD patients with active uveitis and eight male healthy volunteers with informed consent. The serum samples from active BD patients and normal controls were pooled. Highly abundant serum proteins (albumin and IgG) were depleted from these two samples using an affinity capture based kit. The obtained samples wer...

  17. Amino Acid Position-specific Contributions to Amyloid ?-Protein Oligomerization*

    OpenAIRE

    Samir K. Maji; Rachel R. Ogorzalek Loo; Inayathullah, Mohammed; Spring, Sean M.; Vollers, Sabrina S.; Condron, Margaret M.; Bitan, Gal; Joseph A. Loo; David B. Teplow

    2009-01-01

    Understanding the structural and assembly dynamics of the amyloid ?-protein (A?) has direct relevance to the development of therapeutic agents for Alzheimer disease. To elucidate these dynamics, we combined scanning amino acid substitution with a method for quantitative determination of the A? oligomer frequency distribution, photo-induced cross-linking of unmodified proteins (PICUP), to perform “scanning PICUP.” Tyr, a reactive group in PICUP, was substituted at position 1, 10, 20, 30...

  18. Immune hyporesponsiveness to amyloid ?-peptide in amyloid precursor protein transgenic mice: Implications for the pathogenesis and treatment of Alzheimer's disease

    OpenAIRE

    Monsonego, Alon; Maron, Ruth; Zota, Victor; Selkoe, Dennis J.; Weiner, Howard L.

    2001-01-01

    Alzheimer's disease is a dementia that involves progressive deposition of amyloid ?-protein (A?) in brain regions important for memory and cognition, followed by secondary inflammation that contributes to the neuropathologic process. Immunization with A? can reduce cerebral A? burden and consequent neuropathologic changes in the brains of mice transgenic for the ?-amyloid precursor protein (APP). We found that transgenic expression of human APP in B6SJL mice, under the prion promoter, results...

  19. Increased plasma concentration of serum amyloid P component in centenarians with impaired cognitive performance.

    Science.gov (United States)

    Nybo, M; Olsen, H; Jeune, B; Andersen-Ranberg, K; Holm Nielsen, E; Svehag, S E

    1998-01-01

    Serum amyloid P component (SAP) binds to all amyloid fibrils including those in the plaques and tangles of Alzheimer patients. To investigate whether the plasma SAP concentration correlated to cognitive impairment, we measured SAP levels in blood samples from 41 centenarians and compared these to the cognitive performance evaluated by Mini Mental State Examination (MMSE). We observed a significantly (p 24) showed a normal SAP concentration (38.4+/-9.3 microg/ml). No dehydration or hepatic dysfunction was demonstrable in the centenarians. We conclude that the centenarians with impaired cognitive performance had significantly increased plasma concentrations of SAP, while the values for cognitive intact centenarians were within the normal range. PMID:9621998

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

    DEFF Research Database (Denmark)

    Busk, Peter; Jacobsen, Stine

    2010-01-01

    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.

  1. Pathogenetic mechanisms of amyloid A amyloidosis

    OpenAIRE

    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.

    2013-01-01

    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 depositi...

  2. Large Proteins Have a Great Tendency to Aggregate but a Low Propensity to Form Amyloid Fibrils

    OpenAIRE

    Ramshini, Hassan; Parrini, Claudia; Relini, Annalisa; Zampagni, Mariagioia; Mannini, Benedetta; Pesce, Alessandra; SABOURY, Ali Akbar; Nemat-Gorgani, Mohsen; Chiti, Fabrizio

    2011-01-01

    The assembly of soluble proteins into ordered fibrillar aggregates with cross-? structure is an essential event of many human diseases. The polypeptides undergoing aggregation are generally small in size. To explore if the small size is a primary determinant for the formation of amyloids under pathological conditions we have created two databases of proteins, forming amyloid-related and non-amyloid deposits in human diseases, respectively. The size distributions of the two protein populations...

  3. Development and evaluation of agents for targeting visceral amyloid

    OpenAIRE

    Wall, Jonathan S.; Solomon, Alan; Kennel, Stephen J.

    2011-01-01

    Wall, JS, Solomon A, Kennel, SJ. Development and evaluation of agents for targeting visceral amyloid. Visceral amyloidosis is a rare disease characterized by the deposition in organs and tissues of protein fibrils, heparan sulfate proteoglycan as well as serum amyloid P component and other serum proteins. Imaging these pathologic deposits aids in the clinical management of patients with amyloidosis. Whole body scintigraphic imaging of amyloid load as well as organ specific anatomic imaging pr...

  4. Acute-Phase Serum Amyloid A as a Marker of Insulin Resistance in Mice

    OpenAIRE

    Klaus Seedorf; Ulrike Beisiegel; Heike Pospisil; Siesky, Angela M.; Mervyn D. Michael; Heike Zitzer; Barbara Heese; Ludger Scheja

    2008-01-01

    Acute-phase serum amyloid A (A-SAA) was shown recently to correlate with obesity and insulin resistance in humans. However, the mechanisms linking obesity-associated inflammation and elevated plasma A-SAA to insulin resistance are poorly understood. Using high-fat diet- (HFD-) fed mice, we found that plasma A-SAA was increased early upon HFD feeding and was tightly associated with systemic insulin resistance. Plasma A-SAA elevation was due to induction of Saa1 and Saa2 expression in liver but...

  5. Diagnosis Value of the Serum Amyloid A Test in Neonatal Sepsis: A Meta-Analysis

    OpenAIRE

    Haining Yuan; Jie Huang; Bokun Lv; Wenying Yan; Guang Hu; Jian Wang; Bairong Shen

    2013-01-01

    Neonatal sepsis (NS), a common disorder for humans, is recognized as a leading global public health challenge. This meta-analysis was performed to assess the accuracy of the serum amyloid A (SAA) test for diagnosing NS. The studies that evaluated the SAA test as a diagnotic marker were searched in Pubmed, EMBASE, the Cochrane Library, and Google Network between January 1996 and June 2013. A total of nine studies including 823 neonates were included in our meta-analysis. Quality of each study ...

  6. 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)

    Christensen, Michelle BrØnniche; SØrensen, Jens Christian

    2013-01-01

    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.

  7. Protein Polymers and Amyloids : Focus on ?1-Antitrypsin

    DEFF Research Database (Denmark)

    RisØr, Michael Wulff

    2014-01-01

    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

  8. Serum protein capillary electrophoresis and measurement of acute phase proteins in a captive cheetah (Acinonyx jubatus) population

    DEFF Research Database (Denmark)

    Depauw, Sarah; Delanghe, Joris

    2014-01-01

    Renal and gastrointestinal pathologies are widespread in the captive cheetah (Acinonyx jubatus) population but are often diagnosed at a late stage, because diagnostic tools are limited to the evaluation of clinical signs or general blood examination. Presently, no data are available on serum proteins and acute-phase proteins in cheetahs during health or disease, although they might be important to improve health monitoring. This study aimed to quantify serum proteins by capillary electrophoresis in 80 serum samples from captive cheetahs, categorized according to health status and disease type. Moreover, serum amyloid A concentrations were measured via a turbidimetric immunoassay validated in domestic cats, whereas haptoglobin and C-reactive protein were determined by non-species-specific functional tests. Cheetahs classified as healthy had serum protein and acute phase protein concentrations within reference ranges for healthy domestic cats. In contrast, unhealthy cheetahs had higher (P < 0.001) serum amyloidA, alpha2-globulin, and haptoglobin concentrations compared with the healthy subgroup. Moreover, serum amyloid A (P = 0.020), alpha2-globulin (P < 0.001) and haptoglobin (P = 0.001) concentrations in cheetahs suffering from chronic kidney disease were significantly greater compared to the reportedly healthy cheetahs. Our study indicates that serum proteins in the cheetah can be analyzed by routine capillary electrophoresis, whereas acute-phase proteins can be measured using available immunoassays or non-species-specific techniques, which are also likely to be applicable in other exotic felids. Moreover, results suggest that serum amyloid A and haptoglobin are important acute-phase proteins in the diseased cheetah and highlight the need to evaluate their role as early-onset markers for disease.

  9. An antibody to the ?-secretase cleavage site on amyloid ?-protein precursor inhibits amyloid ? production

    Science.gov (United States)

    Thomas, Rhian S.; Liddell, J. Eryl; Murphy, Lynne S.; Pache, David M.; Kidd, Emma J.

    2015-01-01

    Proteolytic cleavage of amyloid ?-protein precursor (A?PP) by ?– and ?–secretases results in production of the amyloid ? peptide (A?) that accumulates in the brains of sufferers of Alzheimer’s Disease (AD). We have developed a monoclonal antibody, 2B12, which binds in the vicinity of the ?-secretase cleavage site on A?PP but does not bind within the A? region. We hypothesised that this antibody, directed against the substrate rather than the enzyme, could inhibit cleavage of A?PP by ?-secretase via steric hindrance and thus reduce downstream production of A?. The antibody would enter cells by binding to A?PP when it is at the cell surface and then be internalised with the protein. We subsequently demonstrated that, after addition of 2B12 to standard growth media, this antibody was indeed capable of inhibiting A?40 production in neuroblastoma and astrocytoma cells expressing native A?PP, as measured by an ELISA. This inhibition was both concentration- and time-dependent and was specific to 2B12. We were only able to inhibit approximately 50% of A?40 production suggesting that not all APP is trafficked to the cell surface. We propose that this antibody could be used as a novel, putative therapy for the treatment of AD. PMID:17183149

  10. Evaluation of Infective Property of Recombinant Prion Protein Amyloids in Cultured Cells Overexpressing Cellular Prion Protein

    OpenAIRE

    Kim, Dae-hwan; Lee, Hye-mi; Ryou, Chongsuk

    2014-01-01

    Misfolded isoform of prion protein (PrP), termed scrapie PrP (PrPSc), tends to aggregate into various fibril forms. Previously, we reported various conditions that affect aggregation of recombinant PrP into amyloids. Because amyloidogenesis of PrP is closely associated with transmissible spongiform encephalopathies such as Creutzfeldt-Jakob disease in humans, we investigated infectivity of recombinant PrP amyloids generated in vitro. Using cultured cell lines which overexpress cellular PrP of...

  11. A systematic exploration of the influence of the protein stability on amyloid fibril formation in vitro

    OpenAIRE

    Rami?rez-alvarado, Marina; Merkel, Jane S.; Regan, Lynne

    2000-01-01

    There are a number of diseases in which normally soluble proteins associate into regular, insoluble amyloid fibrils. The development of in vitro model systems in which detailed structural, kinetic, and thermodynamic characterization are feasible is of critical importance to our understanding of the amyloid fibril phenomenon. The formation of amyloid fibrils by proteins that are not associated with disease has been recently described, suggesting that this may be a common property of many prote...

  12. Membrane mediated aggregation of amyloid-? protein : a potential key event in Alzheimer's disease

    OpenAIRE

    Bokvist, Marcus

    2007-01-01

    The pathogenesis of Alzheimer’s disease (AD), the most common senile dementia, is a complex process. A crucial event in AD is the aggregation of amyloid-? protein (A?), a cleavage product from the Amyloid Precursor Protein (APP). A?40, a common component in amyloid plaques found in patients, aggregates in vitro at concentrations, much higher than the one found in vivo. But in the presence of charged lipid membranes, aggregations occurs at much lower concentration in vitro compared to the...

  13. Immunohistochemical investigation of amyloid ß-protein (Aß) in the brain of aged cats

    OpenAIRE

    Brellou, G.; Vlemmas, I.; Lekkas, S.; Papaioannou, N.

    2005-01-01

    To clarify the immunohistochemical features of amyloid deposits and cerebral amyloid angiopathy (CAA), the distribution of the amyloid ß-protein subtypes Aß40, Aß42, Aß43 and Aß precursor protein (APP) were examined in the brains of fourteen aged cats (7.5-21 year-old). Two types of plaques were detected. The first type was characterized by Aß positive antigenic material and detected in the cortical layers of the frontal and parietal lobes of all examined...

  14. 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)

    S., Martínez-Subiela; J. J., Cerón.

    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 (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.

  15. In vitro Generation of Amyloid A4 Peptide from Amyloid Protein Precursor Through Nonspecific Proteolysis

    Directory of Open Access Journals (Sweden)

    Golam Sadik

    2001-01-01

    Full Text Available Amyloid A4 peptide, the principal constituents of the senile plaques in Alzheimer`s disease (AD originates from proteolysis of a larger protein precursor (APP. Several lines of evidence suggest that this peptide may be generated from aggregated precursor through nonspecific proteolysis. In this work, we used a sensitive in vitro method of detection to investigate the role of nonspecific proteases in the processing of a 100-amino acid C-terminal fragment (C100 inclusive of A4 and cytoplasmic domain of APP. We demonstrate first that C100 forms high molecular weight aggregates in vitro as determined by size exclusion chromatography. Digestion of aggregated C100 with the nonspecific enzyme, proteinase K resulted in cleavage at the amyloidogenic -secretase sites. This occurred at Ala 42-Val 43 generating A4 12-42 & A4 16-42 amyloid peptides. The enzyme cleaved most of the peptide bonds of the cytoplasmic domain and the upstream of A4 domain of the substrate. The result suggests that both the N- and C-terminus A4 can be generated by nonspecific proteases, acting on a aggregated substrate and support the notion that the A4 can be formed in organelles containing proteases capable of cleaving most peptide bonds.

  16. Analysis of Amyloid beta (Ab) protein in Amyloid precursor protein (APP) transgenic mouse models of Alzheimer’s disease (AD) and in human brains

    OpenAIRE

    Rijal Upadhaya, Ajeet

    2012-01-01

    Soluble amyloid b-protein (Ab) aggregates have been identified in Alzheimer"s disease (AD) brain. To address the roles of soluble and dispersible Ab aggregates for neurodegeneration we analyzed two different amyloid precursor protein (APP)-transgenic mouse models. APP23 mice overexpress human mutant APP with the Swedish mutation. APP51/16 mice express high levels of human wild-type APP. Both mice develop Ab plaques. Dendritic degeneration, neuron loss, and loss of asymmetric synapses were see...

  17. The human ?-amyloid precursor protein: biomolecular and epigenetic aspects.

    Science.gov (United States)

    Nguyen, Khue Vu

    2015-03-01

    Beta-amyloid precursor protein (APP) is a membrane-spanning protein with a large extracellular domain and a much smaller intracellular domain. APP plays a central role in Alzheimer's disease (AD) pathogenesis: APP processing generates ?-amyloid (A?) peptides, which are deposited as amyloid plaques in the brains of AD individuals; point mutations and duplications of APP are causal for a subset of early-onset familial AD (FAD) (onset age 65 years old), and the pathophysiology of this disorder is not yet fully understood. APP is an extremely complex molecule that may be functionally important in its full-length configuration, as well as the source of numerous fragments with varying effects on neural function, yet the normal function of APP remains largely unknown. This article provides an overview of our current understanding of APP, including its structure, expression patterns, proteolytic processing and putative functions. Importantly, and for the first time, my recent data concerning its epigenetic regulation, especially in alternative APP pre-mRNA splicing and in the control of genomic rearrangements of the APP gene, are also reported. These findings may provide new directions for investigating the role of APP in neuropathology associated with a deficiency in the enzyme hypoxanthine-guanine phosphoribosyltransferase (HGprt) found in patients with Lesch-Nyhan syndrome (LNS) and its attenuated variants (LNVs). Also, these findings may be of significance for research in neurodevelopmental and neurodegenerative disorders in which the APP gene is involved in the pathogenesis of diseases such as autism, fragile X syndrome (FXS) and AD, with its diversity and complexity, SAD in particular. Accurate quantification of various APP-mRNA isoforms in brain tissues is needed, and antisense drugs are potential treatments. PMID:25719338

  18. beta -Amyloid peptide-induced apoptosis regulated by a novel protein containing a g protein activation module.

    Science.gov (United States)

    Kajkowski, E M; Lo, C F; Ning, X; Walker, S; Sofia, H J; Wang, W; Edris, W; Chanda, P; Wagner, E; Vile, S; Ryan, K; McHendry-Rinde, B; Smith, S C; Wood, A; Rhodes, K J; Kennedy, J D; Bard, J; Jacobsen, J S; Ozenberger, B A

    2001-06-01

    Degeneration of neurons in Alzheimer's disease is mediated by beta-amyloid peptide by diverse mechanisms, which include a putative apoptotic component stimulated by unidentified signaling events. This report describes a novel beta-amyloid peptide-binding protein (denoted BBP) containing a G protein-coupling module. BBP is one member of a family of three proteins containing this conserved structure. The BBP subtype bound human beta-amyloid peptide in vitro with high affinity and specificity. Expression of BBP in cell culture induced caspase-dependent vulnerability to beta-amyloid peptide toxicity. Expression of a signaling-deficient dominant negative BBP mutant suppressed sensitivity of human Ntera-2 neurons to beta-amyloid peptide mediated toxicity. These findings suggest that BBP is a target of neurotoxic beta-amyloid peptide and provide new insight into the molecular pathophysiology of Alzheimer's disease. PMID:11278849

  19. Altered processing of Alzheimer amyloid precursor protein in response to neuronal degeneration.

    OpenAIRE

    Iverfeldt, K; Walaas, S.I.; Greengard, P.

    1993-01-01

    In the brains of individuals with Alzheimer disease, senile plaques containing aggregates of beta-amyloid peptide, derived from the beta-amyloid precursor protein (APP), are seen in association with degenerating nerve terminals. It is not known whether the degenerating nerve terminals cause the formation of these aggregates or whether beta-amyloid peptide in the aggregates causes nerve-terminal degeneration. In the present study of rat brain, degeneration either of local neurons or of nerve t...

  20. Herpes simplex virus interferes with amyloid precursor protein processing

    Directory of Open Access Journals (Sweden)

    Itzhaki Ruth F

    2005-08-01

    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.

  1. The amyloid precursor protein and postnatal neurogenesis/neuroregeneration

    International Nuclear Information System (INIS)

    The amyloid precursor protein (APP) is the source of amyloid-beta (A?) peptide, produced via its sequential cleavage ?- and ?-secretases. Various biophysical forms of A? (and the mutations of APP which results in their elevated levels) have been implicated in the etiology and early onset of Alzheimer's disease. APP's evolutionary conservation and the existence of APP-like isoforms (APLP1 and APLP2) which lack the A? sequence, however, suggest that these might have important physiological functions that are unrelated to A? production. Soluble N-terminal fragments of APP have been known to be neuroprotective, and the interaction of its cytoplasmic C-terminus with a myriad of proteins associates it with diverse processes such as axonal transport and transcriptional regulation. The notion for an essential postnatal function of APP has been demonstrated genetically, as mice deficient in both APP and APLP2 or all three APP isoforms exhibit early postnatal lethality and neuroanatomical abnormalities. Recent findings have also brought to light two possible functions of the APP family in Brain-regulation of neural progenitor cell proliferation and axonal outgrowth after injury. Interestingly, these two apparently related neurogenic/neuroregenerative functions of APP involve two separate domains of the molecule

  2. Large Proteins Have a Great Tendency to Aggregate but a Low Propensity to Form Amyloid Fibrils

    Science.gov (United States)

    Ramshini, Hassan; Parrini, Claudia; Relini, Annalisa; Zampagni, Mariagioia; Mannini, Benedetta; Pesce, Alessandra; Saboury, Ali Akbar; Nemat-Gorgani, Mohsen; Chiti, Fabrizio

    2011-01-01

    The assembly of soluble proteins into ordered fibrillar aggregates with cross-? structure is an essential event of many human diseases. The polypeptides undergoing aggregation are generally small in size. To explore if the small size is a primary determinant for the formation of amyloids under pathological conditions we have created two databases of proteins, forming amyloid-related and non-amyloid deposits in human diseases, respectively. The size distributions of the two protein populations are well separated, with the systems forming non-amyloid deposits appearing significantly larger. We have then investigated the propensity of the 486-residue hexokinase-B from Saccharomyces cerevisiae (YHKB) to form amyloid-like fibrils in vitro. This size is intermediate between the size distributions of amyloid and non-amyloid forming proteins. Aggregation was induced under conditions known to be most effective for amyloid formation by normally globular proteins: (i) low pH with salts, (ii) pH 5.5 with trifluoroethanol. In both situations YHKB aggregated very rapidly into species with significant ?-sheet structure, as detected using circular dichroism and X-ray diffraction, but a weak Thioflavin T and Congo red binding. Moreover, atomic force microscopy indicated a morphology distinct from typical amyloid fibrils. Both types of aggregates were cytotoxic to human neuroblastoma cells, as indicated by the MTT assay. This analysis indicates that large proteins have a high tendency to form toxic aggregates, but low propensity to form regular amyloid in vivo and that such a behavior is intrinsically determined by the size of the protein, as suggested by the in vitro analysis of our sample protein. PMID:21249193

  3. Anti-amyloid precursor protein antibodies inhibit amyloid-? production by steric hindrance

    Science.gov (United States)

    Thomas, Rhian S.; Liddell, J. Eryl; Kidd, Emma J.

    2015-01-01

    Summary Cleavage of amyloid precursor protein (APP) by ?- and ?-secretases results in the production of amyloid-? (A?) in Alzheimer’s disease (AD). We raised two monoclonal antibodies, 2B3 and 2B12, that recognise the ?-secretase cleavage site on APP but not A?. We hypothesised that these antibodies would reduce A? levels via steric hindrance of ?-secretase. Both antibodies decreased extracellular A? levels from astrocytoma cells, but 2B3 was more potent than 2B12. Levels of soluble sAPP? from the non-amyloidogenic ?-secretase pathway and intracellular APP were not affected by either antibody nor were there any effects on cell viability. 2B3 exhibited a higher affinity for APP than 2B12 and its epitope appeared to span the cleavage site while 2B12 bound slightly upstream. Both of these factors probably contribute to its greater effect on A? levels. After 60 minutes incubation at pH 4.0, most 2B3 and 2B12 remained bound to their antigen, suggesting that the antibodies will remain bound to APP in the acidic endosomes where ?-secretase cleavage probably occurs. Only 2B3 and 2B12, but not control antibodies, inhibited the cleavage of sAPP? by ?-secretase in a cell-free assay where effects of antibody internalisation and intracellular degradation were excluded. 2B3 virtually abolished this cleavage. In addition, levels of C-terminal APP fragments, ?CTF, generated following ?-secretase cleavage, were significantly reduced in cells after incubation with 2B3. These results strongly suggest that anti-cleavage site antibodies can generically reduce A? levels via inhibition of ?-secretase by steric hindrance and may provide a novel alternative therapy for AD. PMID:21122073

  4. The coarse-grained OPEP force field for non-amyloid and amyloid proteins.

    Science.gov (United States)

    Chebaro, Yassmine; Pasquali, Samuela; Derreumaux, Philippe

    2012-08-01

    Coarse-grained protein models with various levels of granularity and degrees of freedom offer the possibility to explore many phenomena including folding, assembly, and recognition in terms of dynamics and thermodynamics that are inaccessible to all-atom representations in explicit aqueous solution. Here, we present a refined version of the coarse-grained optimized potential for efficient protein structure prediction (OPEP) based on a six-bead representation. The OPEP version 4.0 parameter set, which uses a new analytical formulation for the nonbonded interactions and adds specific side-chain-side-chain interactions for ?-helix, is subjected to three tests. First, we show that molecular dynamics simulations at 300 K preserve the experimental rigid conformations of 17 proteins with 37-152 amino acids within a root-mean-square deviation (RMSD) of 3.1 Å after 30 ns. Extending the simulation time to 100 ns for five proteins does not change the RMSDs. Second, replica exchange molecular dynamics (REMD) simulations recover the NMR structures of three prototypical ?-hairpin and ?-helix peptides and the NMR three-stranded ?-sheet topology of a 37-residue WW domain, starting from randomly chosen states. Third, REMD simulations on the cc? peptide show a temperature transition from a three-stranded coiled coil to amyloid-like aggregates consistent with experiments, while simulations on low molecular weight aggregates of the prion protein helix 1 do not. Overall, these studies indicate the effectiveness of our OPEP4 coarse-grained model for protein folding and aggregation, and report two future directions for improvement. PMID:22742737

  5. Serum proteins analysis by capillary electrophoresis

    OpenAIRE

    Uji, Yoshinori; Okabe, Hiroaki

    2001-01-01

    The purpose of this study was to evaluate the efficacy of multi-capillary electrophoresis instrument in clinical laboratory. An automated clinical capillary electrophoresis system was evaluated for performing serum proteins electrophoresis and immuno-fixation electrophoresis by subtraction. In this study the performance of capillary electrophoresis was compared with the cellulose acetate membrane electrophoresis and agarose gel immunofixation electrophoresis for serum proteins. The results of...

  6. Serum amyloid a gene expression and immunohistochemical localization in rainbow trout, Oncorhynchus mykiss, infected by Yersinia ruckeri

    DEFF Research Database (Denmark)

    Kania, Per Walter; Buchmann, Kurt

    Serum amyloid A (SAA) is an integral part of the innate immune response in general and in particular the acute phase response. SAA belongs to a highly conserved group of apolipoproteins reported from different groups of organisms such as mammals, birds, fish and even invertebrates. The present study was undertaken to elucidate the role of SAA protein in the innate immune response of rainbow trout. For this purpose a monoclonal antibody was raised against a recombinant peptide of rainbow trout SAA. The antibody was characterized using Western blot, immunohistochemistry and ELISA techniques. SAA was found to be associated with high density lipoprotein (HDL) which complicated band identification in Western blot, but delipidization of the SAA-HDL isolate, using a solvent extraction method, highly increased the quality of reaction in the Western blot. Inhibition ELISA indicated the presence of SAA in serum and tissues (head kidney, liver and spleen) of rainbow trout. 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 further increase at 96 h post infection. Non-significant up-regulations were seen at earlier time points i.e. 4 and 24 h. A weak staining with the monoclonal antibody was seen at 4 h post infection which increased in intensity during the course of infection i.e. 24, 72 and 96 h post infection. The expression pattern of SAA in the infected fry, analysed by qPCR, significantly correlated with the results obtained by immunohistochemical methods. From the present study it can be concluded that the SAA may act as an acute phase protein in rainbow trout and its expression increases significantly during the course of infection.

  7. FMRP Mediates mGluR5-Dependent Translation of Amyloid Precursor Protein

    OpenAIRE

    Westmark, Cara J; Malter, James S

    2007-01-01

    Amyloid precursor protein (APP) facilitates synapse formation in the developing brain, while beta-amyloid (A?) accumulation, which is associated with Alzheimer disease, results in synaptic loss and impaired neurotransmission. Fragile X mental retardation protein (FMRP) is a cytoplasmic mRNA binding protein whose expression is lost in fragile X syndrome. Here we show that FMRP binds to the coding region of APP mRNA at a guanine-rich, G-quartet–like sequence. Stimulation of cortical synaptoneur...

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

    OpenAIRE

    Raymond Chung; Cheng-I Lee; Chi-Fen Lin; Cheng-Ping Jheng; Kun-Hua Yu

    2013-01-01

    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 for...

  9. Proteomic evaluation of sheep serum proteins

    Directory of Open Access Journals (Sweden)

    Chiaradia Elisabetta

    2012-05-01

    Full Text Available Abstract Background The applications of proteomic strategies to ovine medicine remain limited. The definition of serum proteome may be a good tool to identify useful protein biomarkers for recognising sub-clinical conditions and overt disease in sheep. Findings from bovine species are often directly translated for use in ovine medicine. In order to characterize normal protein patterns and improve knowledge of molecular species-specific characteristics, we generated a two-dimensional reference map of sheep serum. The possible application of this approach was tested by analysing serum protein patterns in ewes with mild broncho-pulmonary disease, which is very common in sheep and in the peripartum period which is a stressful time, with a high incidence of infectious and parasitic diseases. Results This study generated the first reference 2-DE maps of sheep serum. Overall, 250 protein spots were analyzed, and 138 identified. Compared with healthy sheep, serum protein profiles of animals with rhino-tracheo-bronchitis showed a significant decrease in protein spots identified as transthyretin, apolipoprotein A1 and a significant increase in spots identified as haptoglobin, endopin 1b and alpha1B glycoprotein. In the peripartum period, haptoglobin, alpha-1-acid glycoprotein, apolipoprotein A1 levels rose, while transthyretin content dropped. Conclusions This study describes applications of proteomics in putative biomarker discovery for early diagnosis as well as for monitoring the physiological and metabolic situations critical for ovine welfare.

  10. Assessement of serum amyloid A levels in the rehabilitation setting in the Florida manatee (Trichechus manatus latirostris).

    Science.gov (United States)

    Cray, Carolyn; Dickey, Meranda; Brewer, Leah Brinson; Arheart, Kristopher L

    2013-12-01

    The acute phase protein serum amyloid A (SAA) has been previously shown to have value as a biomarker of inflammation and infection in many species, including manatees (Trichechus manatus latirostris). In the current study, results from an automated assay for SAA were used in a rehabilitation setting. Reference intervals were established from clinically normal manatees using the robust method: 0-46 mg/L. More than 30-fold higher mean SAA levels were observed in manatees suffering from cold stress and boat-related trauma. Poor correlations were observed between SAA and total white blood count, percentage of neutrophils, albumin, and albumin/globulin ratio. A moderate correlation was observed between SAA and the presence of nucleated red blood cells. The sensitivity of SAA testing was 93% and the specificity was 98%, representing the highest combined values of all the analytes. The results indicate that the automated method for SAA quantitation can provide important clinical data for manatees in a rehabilitation setting. PMID:24450049

  11. Role of serum amyloid P component in bacterial infection: Protection of the host or protection of the pathogen

    OpenAIRE

    Noursadeghi, Mahdad; Bickerstaff, Maria C. M.; Gallimore, J Ruth; Herbert, Jeff; Cohen, Jonathan; Pepys, Mark B.

    2000-01-01

    Serum amyloid P component (SAP) binds to Streptococcus pyogenes, and we show here that it also binds to Neisseria meningitidis, including a lipopolysaccharide (LPS)-negative mutant, and to rough variants of Escherichia coli. Surprisingly, this binding had a powerful antiopsonic effect both in vitro and in vivo, reducing phagocytosis and killing of bacteria. Furthermore, SAP knockout mice survived lethal infection with S. pyogenes and rough E. coli J5, organisms...

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

    OpenAIRE

    Lee, Ha Young; Kim, Sang Doo; Baek, Suk-hwan; Choi, Joon Hyuk; Cho, Kyung-hyun; Zabel, Brian A.; Bae, Yoe-sik

    2013-01-01

    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 re...

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

    Czech Academy of Sciences Publication Activity Database

    Krištofíková, Z.; ?ípová, D.; Hegnerová, Kate?ina; Šírová, J.; Homola, Ji?í

    2013-01-01

    Ro?. 38, ?. 9 (2013), s. 1949-1959. ISSN 0364-3190 Institutional support: RVO:67985882 Keywords : Tau protein * Amyloid ? peptide * Choline transporter Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 2.551, year: 2013

  14. B-Amyloid Precursor Protein Staining of the Brain in Sudden Infant and Early Childhood Death

    DEFF Research Database (Denmark)

    Jensen, Lisbeth Lund; Banner, Jytte

    2013-01-01

    To develop and validate a scoring method for assessing ?-amyloid precursor protein (APP) staining in cerebral white matter and to investigate the occurrence, amount and deposition pattern based on the cause of death in infants and young children.

  15. The role of protein hydrophobicity in conformation change and self-assembly into large amyloid fibers.

    Science.gov (United States)

    Ridgley, Devin M; Claunch, Elizabeth C; Lee, Parker W; Barone, Justin R

    2014-04-14

    It has been found that a short hydrophobic "template" peptide and a larger ?-helical "adder" protein cooperatively self-assemble into micrometer sized amyloid fibers. Here, a common template of trypsin hydrolyzed gliadin is combined with six adder proteins (?-casein, ?-lactalbumin, amylase, hemoglobin, insulin, and myoglobin) to determine what properties of the adder protein drive amyloid self-assembly. Utilizing Fourier Transform-Infrared (FT-IR) spectroscopy, the Amide I absorbance reveals that the observed decrease in ?-helix with time is approximately equal to the increase in high strand density ?-sheet, which is indicative of amyloid formation. The results show that the hydrophobic moment is a good predictor of conformation change but the fraction of aliphatic amino acids within the ?-helices is a better predictor. Upon drying, the protein mixtures form large amyloid fibers. The fiber twist is dependent on the aliphatic index and molecular weight of the adder protein. Here we demonstrate that it is possible to predict the propensity of an adder protein to unfold into an amyloid structure and to predict the fiber morphology, both from adder protein molecular features, which can be applied to the pragmatic engineering of large amyloid fibers. PMID:24601565

  16. Serum amyloid A mediates the inhibitory effect of Ganoderma lucidum polysaccharides on tumor cell adhesion to endothelial cells.

    Science.gov (United States)

    Li, Ying-Bo; Wang, Rui; Wu, Hong-Li; Li, Yu-Hua; Zhong, Li-Jun; Yu, He-Ming; Li, Xue-Jun

    2008-09-01

    Ganoderma ludicum polysaccharides (GlPS) are the major bioactive composition of Ganoderma lucidum, a well-recognized oriental medical fungus. The published data have shown a complementary effect of GlPS in cancer therapy. The present study was designed to determine the anti-tumor efficacy of GlPS and the possible mechanism covering this effect. Murine Sarcoma 180 (S180) model was established, and GlPS administered orally for 10 days. On the 10th day, tumors were weighed to assess the inhibitory effect of GlPS and sera were collected for proteomic analysis and in vitro study. The in vivo results demonstrated that 25, 50 and 100 mg/kg GlPS inhibited S180 growth by 32.67, 44.80 and 45.24%, respectively (P<0.01). Proteomic study revealed marked protein changes after the process of treatment. Three significantly changed proteins were identified by ESI-Q-TOF-MS and database search indicated that they were haptoblobin, apolipoprotein A-II and serum amyloid A (SAA), respectively. Additionally, the expression change of SAA was confirmed by both Western blot and RT-PCR. The adhesion assay showed that GlPS-treated sera dramatically inhibited the adhesion ability of human prostate carcinoma (PC-3M) cells to human umbilical cord vascular endothelial cells (HUVECs), and this effect partially recovered after immunodepletion by the antibody against SAA. Collectively, these results suggest that GlPS inhibited the tumor growth and tumor cell adhesion to HUVECs via up-regulation of SAA protein expression. PMID:18695905

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

    DEFF Research Database (Denmark)

    Olsen, Helle G; Skovgaard, Kerstin

    2013-01-01

    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.

  18. Nanomechanical Properties of Human Prion Protein Amyloid as Probed by Force Spectroscopy

    OpenAIRE

    Ganchev, Dragomir N.; Cobb, Nathan J.; Surewicz, Krystyna; Surewicz, Witold K.

    2008-01-01

    Amyloids are associated with a number of protein misfolding disorders, including prion diseases. In this study, we used single-molecule force spectroscopy to characterize the nanomechanical properties and molecular structure of amyloid fibrils formed by human prion protein PrP90-231. Force-extension curves obtained by specific attachment of a gold-covered atomic force microscope tip to engineered Cys residues could be described by the worm-like chain model for entropic elasticity of a polymer...

  19. Do Amyloid Oligomers Act As Traps for Misfolded Proteins? A Hypothesis

    OpenAIRE

    Gruschus, James M

    2008-01-01

    Mounting evidence points to soluble peptide oligomers as the primary agents in various amyloid and prion diseases. Multiple mechanisms appear to contribute to the cytotoxic effects of these oligomers. Here, an additional, general mechanism is proposed. Hypothesis: soluble amyloid peptide oligomers serve as “all-purpose” beta strands that can interact with transiently unfolded or nascent proteins where interior beta sheet edges are exposed. The proteins, trapped in misfolded states through...

  20. HIV-1 Tat Interacts with and Regulates the Localization and Processing of Amyloid Precursor Protein

    OpenAIRE

    Kim, Jiyoung; Yoon, Jee-Hyun; Kim, Yeon-Soo

    2013-01-01

    HIV-1 Tat protein plays various roles in virus proliferation and in the regulation of numerous host cell functions. Accumulating evidence suggests that HIV-1 Tat also plays an important role in HIV-associated neurocognitive disorders (HAND) by disrupting intracellular communication. Amyloid beta (A?) is generated from amyloid precursor protein (APP) and accumulates in the senile plaques of Alzheimer's disease patients. This study demonstrates that Tat interacts with APP both in vitro and in ...

  1. The Role of Phospholipase D in Amyloid Precursor Protein Processing

    Directory of Open Access Journals (Sweden)

    Mark McLaughlin

    2005-01-01

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-05-29

    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.

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

    OpenAIRE

    Gonzalez-Velasquez Francisco J; Reyes Barcelo Adriana A; Moss Melissa A

    2009-01-01

    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-asse...

  4. Protein Folding and Aggregation into Amyloid: The Interference by Natural Phenolic Compounds

    Directory of Open Access Journals (Sweden)

    Massimo Stefani

    2013-06-01

    Full Text Available Amyloid aggregation is a hallmark of several degenerative diseases affecting the brain or peripheral tissues, whose intermediates (oligomers, protofibrils and final mature fibrils display different toxicity. Consequently, compounds counteracting amyloid aggregation have been investigated for their ability (i to stabilize toxic amyloid precursors; (ii to prevent the growth of toxic oligomers or speed that of fibrils; (iii to inhibit fibril growth and deposition; (iv to disassemble preformed fibrils; and (v to favor amyloid clearance. Natural phenols, a wide panel of plant molecules, are one of the most actively investigated categories of potential amyloid inhibitors. They are considered responsible for the beneficial effects of several traditional diets being present in green tea, extra virgin olive oil, red wine, spices, berries and aromatic herbs. Accordingly, it has been proposed that some natural phenols could be exploited to prevent and to treat amyloid diseases, and recent studies have provided significant information on their ability to inhibit peptide/protein aggregation in various ways and to stimulate cell defenses, leading to identify shared or specific mechanisms. In the first part of this review, we will overview the significance and mechanisms of amyloid aggregation and aggregate toxicity; then, we will summarize the recent achievements on protection against amyloid diseases by many natural phenols.

  5. Protein folding and aggregation into amyloid: the interference by natural phenolic compounds.

    Science.gov (United States)

    Stefani, Massimo; Rigacci, Stefania

    2013-01-01

    Amyloid aggregation is a hallmark of several degenerative diseases affecting the brain or peripheral tissues, whose intermediates (oligomers, protofibrils) and final mature fibrils display different toxicity. Consequently, compounds counteracting amyloid aggregation have been investigated for their ability (i) to stabilize toxic amyloid precursors; (ii) to prevent the growth of toxic oligomers or speed that of fibrils; (iii) to inhibit fibril growth and deposition; (iv) to disassemble preformed fibrils; and (v) to favor amyloid clearance. Natural phenols, a wide panel of plant molecules, are one of the most actively investigated categories of potential amyloid inhibitors. They are considered responsible for the beneficial effects of several traditional diets being present in green tea, extra virgin olive oil, red wine, spices, berries and aromatic herbs. Accordingly, it has been proposed that some natural phenols could be exploited to prevent and to treat amyloid diseases, and recent studies have provided significant information on their ability to inhibit peptide/protein aggregation in various ways and to stimulate cell defenses, leading to identify shared or specific mechanisms. In the first part of this review, we will overview the significance and mechanisms of amyloid aggregation and aggregate toxicity; then, we will summarize the recent achievements on protection against amyloid diseases by many natural phenols. PMID:23765219

  6. The E1 copper binding domain of full-length amyloid precursor protein mitigates copper-induced growth inhibition in brain metastatic prostate cancer DU145 cells

    OpenAIRE

    Gough, Mallory; Blanthorn-Hazell, Sophee; Delury, Craig; Parkin, Edward

    2014-01-01

    Copper plays an important role in the aetiology and growth of tumours and levels of the metal are increased in the serum and tumour tissue of patients affected by a range of cancers including prostate cancer (PCa). The molecular mechanisms that enable cancer cells to proliferate in the presence of elevated copper levels are, therefore, of key importance in our understanding of tumour growth progression. In the current study, we have examined the role played by the amyloid precursor protein (A...

  7. Methylated BSA Mimics Amyloid-Related Proteins and Triggers Inflammation

    OpenAIRE

    Di Domizio, Jeremy; Dorta-estremera, Stephanie; Cao, Wei

    2013-01-01

    The mechanistic study of inflammatory or autoimmune diseases requires the generation of mouse models that reproduce the alterations in immune responses observed in patients. Methylated bovine serum albumin (mBSA) has been widely used to induce antigen-specific inflammation in targeted organs or in combination with single stranded DNA (ssDNA) to generate anti-nucleic acids antibodies in vivo. However, the mechanism by which this modified protein triggers inflammation is poorly understood. By a...

  8. Acute-Phase Serum Amyloid A as a Marker of Insulin Resistance in Mice

    Directory of Open Access Journals (Sweden)

    Klaus Seedorf

    2008-06-01

    Full Text Available Acute-phase serum amyloid A (A-SAA was shown recently to correlate with obesity and insulin resistance in humans. However, the mechanisms linking obesity-associated inflammation and elevated plasma A-SAA to insulin resistance are poorly understood. Using high-fat diet- (HFD- fed mice, we found that plasma A-SAA was increased early upon HFD feeding and was tightly associated with systemic insulin resistance. Plasma A-SAA elevation was due to induction of Saa1 and Saa2 expression in liver but not in adipose tissue. In adipose tissue Saa3 was the predominant isoform and the earliest inflammatory marker induced, suggesting it is important for initiation of adipose tissue inflammation. To assess the potential impact of A-SAA on adipose tissue insulin resistance, we treated 3T3-L1 adipocytes with recombinant A-SAA. Intriguingly, physiological levels of A-SAA caused alterations in gene expression closely resembling those observed in HFD-fed mice. Proinflammatory genes (Ccl2, Saa3 were induced while genes critical for insulin sensitivity (Irs1, Adipoq, Glut4 were down-regulated. Our data identify HFD-fed mice as a suitable model to study A-SAA as a biomarker and a novel possible mediator of insulin resistance.

  9. Diagnosis Value of the Serum Amyloid A Test in Neonatal Sepsis: A Meta-Analysis

    Science.gov (United States)

    Yuan, Haining; Huang, Jie; Lv, Bokun; Yan, Wenying; Hu, Guang; Wang, Jian; Shen, Bairong

    2013-01-01

    Neonatal sepsis (NS), a common disorder for humans, is recognized as a leading global public health challenge. This meta-analysis was performed to assess the accuracy of the serum amyloid A (SAA) test for diagnosing NS. The studies that evaluated the SAA test as a diagnotic marker were searched in Pubmed, EMBASE, the Cochrane Library, and Google Network between January 1996 and June 2013. A total of nine studies including 823 neonates were included in our meta-analysis. Quality of each study was evaluated by the quality assessment of diagnostic accuracy studies tool (QUADAS). The SAA test showed moderate accuracy in the diagnosis of NS both at the first suspicion of sepsis and 8–96?h after the sepsis onset, both with Q* = 0.91, which is similar to the PCT and CRP tests for the diagnosis of NS in the same period. Heterogeneity between studies was also explained by cut-off point, SAA assay, and age of included neonates. On the basis of our meta-analysis, therefore, SAA could be promising and meaningful in the diagnosis of NS. PMID:23984377

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

    DEFF Research Database (Denmark)

    Horvath, A; Andersen, I

    2001-01-01

    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.

  11. Intestinal Epithelial Serum Amyloid A Modulates Bacterial Growth In Vitro and Pro-Inflammatory Responses in Mouse Experimental Colitis

    Directory of Open Access Journals (Sweden)

    Wang Yu

    2010-11-01

    Full Text Available Abstract Background Serum Amyloid A (SAA is a major acute phase protein of unknown function. SAA is mostly expressed in the liver, but also in other tissues including the intestinal epithelium. SAA reportedly has anti-bacterial effects, and because inflammatory bowel diseases (IBD result from a breakdown in homeostatic interactions between intestinal epithelia and bacteria, we hypothesized that SAA is protective during experimental colitis. Methods Intestinal SAA expression was measured in mouse and human samples. Dextran sodium sulfate (DSS colitis was induced in SAA 1/2 double knockout (DKO mice and in wildtype controls. Anti-bacterial effects of SAA1/2 were tested in intestinal epithelial cell lines transduced with adenoviral vectors encoding the CE/J SAA isoform or control vectors prior to exposure to live Escherichia coli. Results Significant levels of SAA1/SAA2 RNA and SAA protein were detected by in situ hybridization and immunohistochemistry in mouse colonic epithelium. SAA3 expression was weaker, but similarly distributed. SAA1/2 RNA was present in the ileum and colon of conventional mice and in the colon of germfree mice. Expression of SAA3 was strongly regulated by bacterial lipopolysaccharides in cultured epithelial cell lines, whereas SAA1/2 expression was constitutive and not LPS inducible. Overexpression of SAA1/2 in cultured epithelial cell lines reduced the viability of co-cultured E. coli. This might partially explain the observed increase in susceptibility of DKO mice to DSS colitis. SAA1/2 expression was increased in colon samples obtained from Crohn's Disease patients compared to controls. Conclusions Intestinal epithelial SAA displays bactericidal properties in vitro and could play a protective role in experimental mouse colitis. Altered expression of SAA in intestinal biopsies from Crohn's Disease patients suggests that SAA is involved in the disease process..

  12. 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)

    Stenfeldt, Carolina; Heegaard, Peter M. H.

    2011-01-01

    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.

  13. Serum Amyloid A and Clusterin as Potential Predictive Biomarkers for Severe Hand, Foot and Mouth Disease by 2D-DIGE Proteomics Analysis

    Science.gov (United States)

    Hong, Wen-Xu; Ren, Xiaohu; Yang, Xifei; He, Yanxia; Wang, Wenjian; Zhang, Renli; Yang, Hong; Zhao, Zhiguang; Huang, Haiyan; Chen, Long; Zhao, Dejian; Xian, Huixia; Yang, Fang; Ma, Dongli; Yang, Linqing; Yin, Yundong; Zhou, Li; Chen, Xiaozhen; Cheng, Jinquan

    2014-01-01

    Hand, foot, and mouth disease (HFMD) affects more than one million children, is responsible for several hundred child deaths every year in China and is the cause of widespread concerns in society. Only a small fraction of HFMD cases will develop further into severe HFMD with neurologic complications. A timely and accurate diagnosis of severe HFMD is essential for assessing the risk of progression and planning the appropriate treatment. Human serum can reflect the physiological or pathological states, which is expected to be an excellent source of disease-specific biomarkers. In the present study, a comparative serological proteome analysis between severe HFMD patients and healthy controls was performed via a two-dimensional difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) strategy. Fifteen proteins were identified as differentially expressed in the sera of the severe HFMD patients compared with the controls. The identified proteins were classified into different groups according to their molecular functions, biological processes, protein classes and physiological pathways by bioinformatics analysis. The up-regulations of two identified proteins, serum amyloid A (SAA) and clusterin (CLU), were confirmed in the sera of the HFMD patients by ELISA assay. This study not only increases our background knowledge about and scientific insight into the mechanisms of HFMD, but also reveals novel potential biomarkers for the clinical diagnosis of severe HFMD. PMID:25268271

  14. Membrane-protein interactions hold the key to understanding amyloid formation

    CERN Document Server

    Straub, John E

    2014-01-01

    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.

  15. Predicted alpha-helical regions of the prion protein when synthesized as peptides form amyloid.

    OpenAIRE

    Gasset, M.; Baldwin, M. A.; Lloyd, D. H.; Gabriel, J. M.; Holtzman, D. M.; Cohen, F.; Fletterick, R.; Prusiner, S. B.

    1992-01-01

    By comparing the amino acid sequences of 11 mammalian and 1 avian prion proteins (PrP), structural analyses predicted four alpha-helical regions. Peptides corresponding to these regions of Syrian hamster PrP were synthesized, and, contrary to predictions, three of the four spontaneously formed amyloids as shown by electron microscopy and Congo red staining. By IR spectroscopy, these amyloid peptides exhibited secondary structures composed largely of beta-sheets. The first of the predicted hel...

  16. A study of b-secretase cleaved Alzheimer amyloid precursor protein

    OpenAIRE

    Sennvik, Kristina

    2002-01-01

    Alzheimer's disease (AD) is characterized by the degeneration and loss of neurons, intracellular neurofibrillary tangles and the accumulation of extracellular senile plaques consisting mainly of beta- amyloid (A-beta). A-beta is generated from the amyloid precursor protein (APP) through sequential cleavage by proteases P- and gamma-secretase. Alternatively, APP may be cleaved within the A-beta region by alpha-secretase, preventing intact A-beta formation. Both the alpha- and...

  17. Amyloid-? protein oligomerization and the importance of tetramers and dodecamers in the aetiology of Alzheimer's disease.

    OpenAIRE

    Bernstein, SL; Dupuis, NF; Lazo, ND; Wyttenbach, T; Condron, MM; Bitan, G; Teplow, DB; Shea, JE; Ruotolo, BT; Robinson, CV; Bowers, MT

    2009-01-01

    In recent years, small protein oligomers have been implicated in the aetiology of a number of important amyloid diseases, such as type 2 diabetes, Parkinson’s disease and Alzheimer’s disease. As a consequence, research efforts are being directed away from traditional targets, such as amyloid plaques, and towards characterization of early oligomer states. Here we present a new analysis method, ion mobility coupled with mass spectrometry, for this challenging problem, which allows determina...

  18. Amyloid precursor protein secretases as therapeutic targets for traumatic brain injury

    OpenAIRE

    Loane, David J.; Pocivavsek, Ana; Moussa, Charbel E-h; Thompson, Rachel; Matsuoka, Yasuji; Faden, Alan I.; Rebeck, G. William; Burns, Mark P.

    2009-01-01

    Amyloid-? (A?) peptides, found in Alzheimer’s disease brain, accumulate rapidly after traumatic brain injury (TBI) in both humans and animals. Here we show that blocking either ?- or ?-secretase, enzymes required for production of A? from amyloid precursor protein (APP), can ameliorate motor and cognitive deficits and reduce cell loss after experimental TBI in mice. Thus, APP secretases are promising targets for treatment of TBI.

  19. Two amyloid precursor protein transgenic mouse models with Alzheimer?disease-like?pathology

    OpenAIRE

    Sturchler-pierrat, Christine; Abramowski, Dorothee; Duke, Mairead; Wiederhold, Karl-heinz; Mistl, Claudia; Rothacher, Sabin; Ledermann, Birgit; Bu?rki, Kurt; Frey, Peter; Paganetti, Paolo A.; Waridel, Caroline; Calhoun, Michael E.; Jucker, Mathias; Probst, Alphonse; Staufenbiel, Matthias

    1997-01-01

    Mutations in the amyloid precursor protein (APP) gene cause early-onset familial Alzheimer disease (AD) by affecting the formation of the amyloid ? (A?) peptide, the major constituent of AD plaques. We expressed human APP751 containing these mutations in the brains of transgenic mice. Two transgenic mouse lines develop pathological features reminiscent of AD. The degree of pathology depends on expression levels and specific mutations. A 2-fold overexpression of human APP with the Swedish do...

  20. Oxysterol-binding protein-1 (OSBP1 modulates processing and trafficking of the amyloid precursor protein

    Directory of Open Access Journals (Sweden)

    Seabrook Guy R

    2008-03-01

    Full Text Available Background Evidence from biochemical, epidemiological and genetic findings indicates that cholesterol levels are linked to amyloid-? (A? production and Alzheimer's disease (AD. Oxysterols, which are cholesterol-derived ligands of the liver X receptors (LXRs and oxysterol binding proteins, strongly regulate the processing of amyloid precursor protein (APP. Although LXRs have been studied extensively, little is known about the biology of oxysterol binding proteins. Oxysterol-binding protein 1 (OSBP1 is a member of a family of sterol-binding proteins with roles in lipid metabolism, regulation of secretory vesicle generation and signal transduction, and it is thought that these proteins may act as sterol sensors to control a variety of sterol-dependent cellular processes. Results We investigated whether OSBP1 was involved in regulating APP processing and found that overexpression of OSBP1 downregulated the amyloidogenic processing of APP, while OSBP1 knockdown had the opposite effect. In addition, we found that OSBP1 altered the trafficking of APP-Notch2 dimers by causing their accumulation in the Golgi, an effect that could be reversed by treating cells with OSBP1 ligand, 25-hydroxycholesterol. Conclusion These results suggest that OSBP1 could play a role in linking cholesterol metabolism with intracellular APP trafficking and A? production, and more importantly indicate that OSBP1 could provide an alternative target for A?-directed therapeutic.

  1. Protein A-gold immunoelectron microscopic study of amyloid fibrils, granular deposits, and fibrillar luminal aggregates in renal amyloidosis.

    OpenAIRE

    G. C. Yang; Gallo, G. R.

    1990-01-01

    Glomeruli of archival renal biopsies, stored frozen at -70 degrees C, from three patients with amyloid were examined by protein A-gold immunoelectron microscopy. In one with both fibrillar and granular deposits from a 'skin popper' drug abuser, the granular deposits were labeled with anti-IgG, while the fibrillar deposits were labeled with anti-amyloid-A (AA) protein and amyloid P component (AP), suggesting coexisting immune complex disease and AA due to different, but possibly related, patho...

  2. Evidence that the 42- and 40-amino acid forms of amyloid ? protein are generated from the ?-amyloid precursor protein by different?protease?activities

    Science.gov (United States)

    Citron, Martin; Diehl, Thekla?S.; Gordon, Grace; Biere, Anja?Leona; Seubert, Peter; Selkoe, Dennis?J.

    1996-01-01

    Cerebral deposition of the amyloid ? protein (A?) is an early and invariant feature of Alzheimer disease (AD). Whereas the 40-amino acid form of A? (A?40) accounts for ?90% of all A? normally released from cells, it appears to contribute only to later phases of the pathology. In contrast, the longer more amyloidogenic 42-residue form (A?42), accounting for only ?10% of secreted A?, is deposited in the earliest phase of AD and remains the major constituent of most amyloid plaques throughout the disease. Moreover, its levels have been shown to be increased in all known forms of early-onset familial AD. Thus, inhibition of A?42 production is a prime therapeutic goal. The same protease, ?-secretase, is assumed to generate the C termini of both A?40 and A?42. Herein, we analyze the effect of the compound MDL 28170, previously suggested to inhibit ?-secretase, on ?-amyloid precursor protein processing. By immunoprecipitating conditioned medium of different cell lines with various A?40- and A?42-specific antibodies, we demonstrate a much stronger inhibition of the ?-secretase cleavage at residue 40 than of that at residue 42. These data suggest that different proteases generate the A?40 and A?42 C termini. Further, they raise the possibility of identifying compounds that do not interfere with general ?-amyloid precursor protein metabolism, including A?40 production, but specifically block the generation of the pathogenic A?42 peptide. PMID:8917563

  3. In silico study of amyloid -protein folding and oligomerization

    Science.gov (United States)

    Urbanc, B.; Cruz, L.; Yun, S.; Buldyrev, S. V.; Bitan, G.; Teplow, D. B.; Stanley, H. E.

    2004-12-01

    Experimental findings suggest that oligomeric forms of the amyloid protein (A) play a critical role in Alzheimer's disease. Thus, elucidating their structure and the mechanisms of their formation is critical for developing therapeutic agents. We use discrete molecular dynamics simulations and a four-bead protein model to study oligomerization of two predominant alloforms, A40 and A42, at the atomic level. The four-bead model incorporates backbone hydrogen-bond interactions and amino acid-specific interactions mediated through hydrophobic and hydrophilic elements of the side chains. During the simulations we observe monomer folding and aggregation of monomers into oligomers of variable sizes. A40 forms significantly more dimers than A42, whereas pentamers are significantly more abundant in A42 relative to A40. Structure analysis reveals a turn centered at Gly-37-Gly-38 that is present in a folded A42 monomer but not in a folded A40 monomer and is associated with the first contacts that form during monomer folding. Our results suggest that this turn plays an important role in A42 pentamer formation. A pentamers have a globular structure comprising hydrophobic residues within the pentamer's core and hydrophilic N-terminal residues at the surface of the pentamer. The N termini of A40 pentamers are more spatially restricted than A42 pentamers. A40 pentamers form a -strand structure involving Ala-2-Phe-4, which is absent in A42 pentamers. These structural differences imply a different degree of hydrophobic core exposure between pentamers of the two alloforms, with the hydrophobic core of the A?42 pentamer being more exposed and thus more prone to form larger oligomers.

  4. Serum concentrations of selected acute phase proteins and enzyme activities after injection of a combined mineral preparation in calves

    Directory of Open Access Journals (Sweden)

    Tóthová Csilla

    2009-01-01

    Full Text Available The objective of this study was to evaluate the possible effects of various forms of injections of a combined mineral preparation in calves on changes in serum concentrations of selected acute phase proteins - haptoglobin (Hp and serum amyloid A (SAA, as well as on changes in the activity of selected enzymes - creatine kinase (CK and aspartate amino-transferase (AST. The changes in serum concentrations and activities of the aforementioned variables were evaluated after administration of the recommended therapeutic dose of the mentioned preparation and its repeated administration, and compared with changes recorded after a double therapeutic dose and its repeated administration. Analyses showed a more marked increase of mean SAA concentrations after intramuscular and subcutaneous injections of the higher preparation dose. A gradual insignificant increase in Hp serum concentrations was observed. The most significant changes were recorded in increasing serum activities of CK and AST after intramuscular injections of higher dose of the remedy (P<0.001 and P<0.01, respectively. Presented results indicate that tissue injury and muscle damage caused by repeated injections, predominantly at higher doses of drugs, may result not only in increased activities of some enzymes, but also in more marked changes in serum concentrations of certain acute phase proteins, particularly serum amyloid A.

  5. Tolfenamic acid interrupts the de novo synthesis of the ?-amyloid precursor protein and lowers amyloid beta via a transcriptional pathway.

    Science.gov (United States)

    Adwan, L I; Basha, R; Abdelrahim, M; Subaiea, G M; Zawia, N H

    2011-06-01

    Amyloid beta (A?) peptides are related to the pathogenesis of Alzheimer's disease (AD). The search for therapeutic strategies that lower these peptides has mainly focused on the proteolytic processing of the ?-amyloid precursor protein (APP), and other post-transcriptional pathways. The transcription factor specificity protein 1 (Sp1) is vital for the regulation of several genes involved in AD including APP and the beta site APP cleaving enzyme 1 (BACE1). We have previously reported that tolfenamic acid promotes the degradation of Sp1 protein (SP1) in pancreatic human cancer cells and mice tumors. This study examines the ability of tolfenamic acid to reduce SP1 levels, and thereby decrease APP transcription and A? levels in rodent brains. Tolfenamic acid was administered by oral gavage to C57BL/6 mice at variable dosages and for different time periods. Results have shown that tolfenamic acid was able to down regulate brain protein levels of SP1, APP, and A?. These findings demonstrate that interference with upstream transcriptional pathways can lower pathogenic intermediates associated with AD, and thus tolfenamic acid represents a novel approach for the development of a therapeutic intervention for AD. PMID:21557719

  6. Amyloid-related biomarkers and axonal damage proteins in parkinsonian syndromes

    DEFF Research Database (Denmark)

    Bech, Sara; Hjermind, Lena E

    2012-01-01

    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-?(1-42), and the soluble ?- and ?-cleaved fragments of amyloid precursor proteins in a cohort of patients with various PS.

  7. Serum protein concentrations in Plasmodium falciparum malaria.

    Science.gov (United States)

    Graninger, W; Thalhammer, F; Hollenstein, U; Zotter, G M; Kremsner, P G

    1992-12-01

    In patients with uncomplicated Plasmodium falciparum infection cytokine-mediated serum protein levels of C-reactive protein (CRP), coeruloplasmin (COE), beta 2-microglobulin (B2M), alpha 1-acid glycoprotein (AAG), alpha 1-antitrypsin (AAT), haptoglobin (HPT), prealbumin (PRE), retinol binding protein (RBP), albumin (ALB) and transferrin (TRF) were measured in an endemic area of the Amazonian rain forest. Semi-immune (SI) and nonimmune (NI) patients were investigated. In both patient groups the serum concentrations of CRP, COE and B2M were elevated on admission. In addition AAG and AAT concentrations were increased in NI patients compared to control subjects. Significantly lower serum concentrations of HPT, PRE, RBP, ALB and TRF were seen in both patient groups during the acute phase of the disease, and were more pronounced in NI patients. After a 28-day follow-up, AAT and B2M were normal in SI patients but HPT, AAT and B2M were still significantly altered in NI patients. PMID:1283805

  8. Self-assembling of amyloid-like proteins

    Energy Technology Data Exchange (ETDEWEB)

    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

    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)

  9. The E693? Mutation in Amyloid Precursor Protein Increases Intracellular Accumulation of Amyloid ? Oligomers and Causes Endoplasmic Reticulum Stress-Induced Apoptosis in Cultured Cells

    OpenAIRE

    Nishitsuji, Kazuchika; Tomiyama, Takami; Ishibashi, Kenichi; Ito, Kazuhiro; Teraoka, Rie; Lambert, Mary P.; Klein, William L.; Mori, Hiroshi

    2009-01-01

    The E693? mutation within the amyloid precursor protein (APP) has been suggested to cause dementia via the enhanced formation of synaptotoxic amyloid ? (A?) oligomers. However, this mutation markedly decreases A? secretion, implying the existence of an additional mechanism of neuronal dysfunction that is independent of extracellular A?. We therefore examined the effects of this mutation on both APP processing to produce A? as well as subcellular localization and accumulation of A? in t...

  10. Amyloid fibrils in human insulinoma and islets of Langerhans of the diabetic cat are derived from a neuropeptide-like protein also present in normal islet cells.

    OpenAIRE

    Westermark, P.; Wernstedt, C.; Wilander, E.; Hayden, D. W.; O Brien, T. D.; Johnson, K. H.

    1987-01-01

    Amyloid deposits localized to the islets of Langerhans are typical of non-insulin-dependent human diabetes mellitus and of diabetes mellitus in adult cats. Amyloid deposits also commonly occur in insulin-producing pancreatic tumors. We have purified a major protein--insulinoma or islet amyloid polypeptide (IAPP)--from human and cat islet amyloid and from amyloid of a human insulinoma. IAPP from human insulinoma contained 37 amino acid residues and had a theoretical molecular mass of 3850 Da. ...

  11. 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)

    Sheng-Wei Ren

    2014-04-01

    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.

  12. Immune hyporesponsiveness to amyloid ?-peptide in amyloid precursor protein transgenic mice: Implications for the pathogenesis and treatment of Alzheimer's disease

    Science.gov (United States)

    Monsonego, Alon; Maron, Ruth; Zota, Victor; Selkoe, Dennis J.; Weiner, Howard L.

    2001-01-01

    Alzheimer's disease is a dementia that involves progressive deposition of amyloid ?-protein (A?) in brain regions important for memory and cognition, followed by secondary inflammation that contributes to the neuropathologic process. Immunization with A? can reduce cerebral A? burden and consequent neuropathologic changes in the brains of mice transgenic for the ?-amyloid precursor protein (APP). We found that transgenic expression of human APP in B6SJL mice, under the prion promoter, results in immune hyporesponsiveness to human A?, in terms of both antibody and cellular immune responses. The decreased antibody responses were related not to B cell tolerance but rather to the inability of A?-specific T cells to provide help for antibody production. The immune hyporesponsiveness could be overcome if T cell help was provided by coupling an A? B cell epitope to BSA. Our results suggest that expression of APP in transgenic mice is associated with an A?-specific impaired adaptive immune response that may contribute to the neuropathology. Moreover, humans with life-long elevation of brain and peripheral A? (e.g., patients with presenilin mutations or Down syndrome) could have reduced immune responses to A? vaccination. PMID:11517335

  13. Structure of a Functional Amyloid Protein Subunit Computed Using Sequence Variation

    DEFF Research Database (Denmark)

    Tian, Pengfei; Boomsma, Wouter

    2014-01-01

    Functional amyloid fibers, called curli, play a critical role in adhesion and invasion of many bacteria. Unlike pathological amyloids, curli structures are formed by polypeptide sequences whose amyloid structure has been selected for during evolution. This important distinction provides us with an opportunity to obtain structural insights from an unexpected source: the covariation of amino acids in sequences of different curli proteins. We used recently developed methods to extract amino acid contacts from a multiple sequence alignment of homologues of the curli subunit protein, CsgA. Together with an efficient force field, these contacts allow us to determine structural models of CsgA. We find that CsgA forms a ?-helical structure, where each turn corresponds to previously identified repeat sequences in CsgA. The proposed structure is validated by previously measured solid-state NMR, electron microscopy and X-ray diffraction data, and agrees with an earlier proposed model derived by complementary means.

  14. Nanomechanical properties of human prion protein amyloid as probed by force spectroscopy.

    Science.gov (United States)

    Ganchev, Dragomir N; Cobb, Nathan J; Surewicz, Krystyna; Surewicz, Witold K

    2008-09-15

    Amyloids are associated with a number of protein misfolding disorders, including prion diseases. In this study, we used single-molecule force spectroscopy to characterize the nanomechanical properties and molecular structure of amyloid fibrils formed by human prion protein PrP90-231. Force-extension curves obtained by specific attachment of a gold-covered atomic force microscope tip to engineered Cys residues could be described by the worm-like chain model for entropic elasticity of a polymer chain, with the size of the N-terminal segment that could be stretched entropically depending on the tip attachment site. The data presented here provide direct information about the forces required to extract an individual monomer from the core of the PrP90-231 amyloid, and indicate that the beta-sheet core of this amyloid starts at residue approximately 164-169. The latter finding has important implications for the ongoing debate regarding the structure of PrP amyloid. PMID:18539633

  15. Evidence that the 42- and 40-amino acid forms of amyloid beta protein are generated from the beta-amyloid precursor protein by different protease activities.

    Science.gov (United States)

    Citron, M; Diehl, T S; Gordon, G; Biere, A L; Seubert, P; Selkoe, D J

    1996-11-12

    Cerebral deposition of the amyloid beta protein (A beta) is an early and invariant feature of Alzheimer disease (AD). Whereas the 40-amino acid form of A beta (A beta 40) accounts for approximately 90% of all A beta normally released from cells, it appears to contribute only to later phases of the pathology. In contrast, the longer more amyloidogenic 42-residue form (A beta 42), accounting for only approximately 10% of secreted A beta, is deposited in the earliest phase of AD and remains the major constituent of most amyloid plaques throughout the disease. Moreover, its levels have been shown to be increased in all known forms of early-onset familial AD. Thus, inhibition of A beta 42 production is a prime therapeutic goal. The same protease, gamma-secretase, is assumed to generate the C termini of both A beta 40 and A beta 42. Herein, we analyze the effect of the compound MDL 28170, previously suggested to inhibit gamma-secretase, on beta-amyloid precursor protein processing. By immunoprecipitating conditioned medium of different cell lines with various A beta 40- and A beta 42-specific antibodies, we demonstrate a much stronger inhibition of the gamma-secretase cleavage at residue 40 than of that at residue 42. These data suggest that different proteases generate the A beta 40 and A beta 42 C termini. Further, they raise the possibility of identifying compounds that do not interfere with general beta-amyloid precursor protein metabolism, including A beta 40 production, but specifically block the generation of the pathogenic A beta 42 peptide. PMID:8917563

  16. Preparation of amyloid-like fibrils containing magnetic iron oxide nanoparticles: Effect of protein aggregation on proton relaxivity

    International Nuclear Information System (INIS)

    Highlights: ? Preparation of amyloid materials labeled with magnetic iron oxide nanoparticles. ? Characterization of amyloid materials by electron tomography. ? Influence of protein aggregation on the magnetic nanoparticle properties. -- Abstract: A method to prepare amyloid-like fibrils functionalized with magnetic nanoparticles has been developed. The amyloid-like fibrils are prepared in a two step procedure, where insulin and magnetic nanoparticles are mixed simply by grinding in the solid state, resulting in a water soluble hybrid material. When the hybrid material is heated in aqueous acid, the insulin/nanoparticle hybrid material self assembles to form amyloid-like fibrils incorporating the magnetic nanoparticles. This results in magnetically labeled amyloid-like fibrils which has been characterized by Transmission Electron Microscopy (TEM) and electron tomography. The influence of the aggregation process on proton relaxivity is investigated. The prepared materials have potential uses in a range of bio-imaging applications.

  17. Biochemical indicators of vitamin A deficiency: Serum retinol and serum retinol binding protein

    OpenAIRE

    Pee, S.; Dary, O.

    2002-01-01

    Two biochemical indicators are currently recommended for determining whether vitamin A deficiency (VAD) is a public health problem: serum retinol and serum retinol-binding protein (RBP). After consideration of 40 data sets and the original rationale for previously proposed cut-offs, a cut-off for serum retinol concentration was proposed at

  18. 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)

    Darusman, Huda Shalahudin; Sajuthi, D

    2013-01-01

    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.

  19. 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 migh 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.)

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

    OpenAIRE

    Heber, S.; Herms, J; Gajic, V; J.A. Hainfellner; Aguzzi, A.; Rülicke, T; von Kretzschmar, H; von Koch, C; Sisodia, S; Tremml, P; Lipp, H P; Wolfer, D. P.; U Müller

    2000-01-01

    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...

  1. Synthetic peptide homologous to ? protein from Alzheimer's disease forms amyloid-like fibrils in vitro

    International Nuclear Information System (INIS)

    Progressive amyloid deposition in senile plaques and cortical blood vessels may play a central role in the pathogenesis of Alzheimer's disease. The authors have used x-ray diffraction and electron microscopy to study the molecular organization and morphology of macromolecular assemblies formed by three synthetic peptides homologous to ? protein of brain amyloid: ?-(1-28), residues 1-28 of the ? protein; [Ala1-?-(1-28), ?-(1-28) with alanine substituted for lysine at position 16; and ?-(18-28), residues 18-28 of the ? protein. ?-(1-28) readily formed fibrils in vitro that were similar in ultrastructure to the in vivo amyloid and aggregated into large bundles resembling those of senile plaque cores. X-ray patterns from partially dried, oriented pellets showed a cross-?-conformation. [Ala16]?-(1-28) formed ?-pleated sheet assemblies that were dissimilar to in vivo fibrils. The width of the 10-A spacing indicated stacks of about six sheets. Thus, substitution of the uncharged alanine for the positively charged lysine in the ?-strand region enhances the packing of the sheets and dramatically alters the type of macromolecular aggregate formed. ?-(18-28) formed assemblies that had even a greater number of stacked sheets. The findings on these homologous synthetic assemblies help to define the specific sequence that is required to form Alzheimer's-type amyloid fibrils, thus providing an in vitro model of age-related cerebral amyloidogenesisted cerebral amyloidogenesis

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

    Science.gov (United States)

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

    2013-01-01

    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

  3. Soluble Amyloid Precursor Protein Induces Rapid Neural Differentiation of Human Embryonic Stem Cells

    OpenAIRE

    Freude, Kristine K.; Penjwini, Mahmud; Davis, Joy L.; Laferla, Frank M.; Blurton-jones, Mathew

    2011-01-01

    Human embryonic stem cells (hESCs) offer tremendous potential for not only treating neurological disorders but also for their ability to serve as vital reagents to model and investigate human disease. To further our understanding of a key protein involved in Alzheimer disease pathogenesis, we stably overexpressed amyloid precursor protein (APP) in hESCs. Remarkably, we found that APP overexpression in hESCs caused a rapid and robust differentiation of pluripotent stem cells toward a neural fa...

  4. A link between inflammation and metastasis: serum amyloid A1 and A3 induce metastasis, and are targets of metastasis-inducing S100A4.

    Science.gov (United States)

    Hansen, M T; Forst, B; Cremers, N; Quagliata, L; Ambartsumian, N; Grum-Schwensen, B; Klingelhöfer, J; Abdul-Al, A; Herrmann, P; Osterland, M; Stein, U; Nielsen, G H; Scherer, P E; Lukanidin, E; Sleeman, J P; Grigorian, M

    2015-01-22

    S100A4 is implicated in metastasis and chronic inflammation, but its function remains uncertain. Here we establish an S100A4-dependent link between inflammation and metastatic tumor progression. We found that the acute-phase response proteins serum amyloid A (SAA) 1 and SAA3 are transcriptional targets of S100A4 via Toll-like receptor 4 (TLR4)/nuclear factor-?B signaling. SAA proteins stimulated the transcription of RANTES (regulated upon activation normal T-cell expressed and presumably secreted), G-CSF (granulocyte-colony-stimulating factor) and MMP2 (matrix metalloproteinase 2), MMP3, MMP9 and MMP13. We have also shown for the first time that SAA stimulate their own transcription as well as that of proinflammatory S100A8 and S100A9 proteins. Moreover, they strongly enhanced tumor cell adhesion to fibronectin, and stimulated migration and invasion of human and mouse tumor cells. Intravenously injected S100A4 protein induced expression of SAA proteins and cytokines in an organ-specific manner. In a breast cancer animal model, ectopic expression of SAA1 or SAA3 in tumor cells potently promoted widespread metastasis formation accompanied by a massive infiltration of immune cells. Furthermore, coordinate expression of S100A4 and SAA in tumor samples from colorectal carcinoma patients significantly correlated with reduced overall survival. These data show that SAA proteins are effectors for the metastasis-promoting functions of S100A4, and serve as a link between inflammation and tumor progression. PMID:24469032

  5. Preliminary proteomic-based identification of a novel protein for Down's syndrome in maternal serum.

    Science.gov (United States)

    Yu, Bin; Zhang, Bin; Wang, Jing; Wang, Qiu-wei; Huang, Rui-ping; Yang, Yu-qi; Shao, Shi-he

    2012-05-01

    Prenatal screening for Down's syndrome (DS) is in need of improvement. As a powerful platform, proteomics techniques could also be used for identification of new biomarkers for DS screening. In this case-control proteome study, pregnant women were diagnosed prenatally by karyotype analysis from amniotic fluid (AF). Maternal serum samples were collected from six pregnancies with fetuses affected by DS and six pregnancies with normal fetuses. First, we used two-dimensional electrophoresis and mass spectrometry to identify the different levels of expression of proteins in maternal serum between the DS and control groups in the second trimester. Second, we used bioinformatics to analyze the proteins by DAVID. Then, the interesting candidates were further tested by enzyme-linked immunosorbent assay (ELISA). Twenty-nine proteins were successfully identified in maternal serum obtained from pregnancies with fetuses affected by DS. The top five proteins up-regulated were serotransferrin (TF), alpha-1b-glycoprotein (A1BG), desmin (DES), alpha-1-antitrypsin (SERPINA1) and ceruloplasmin (CP), while serum amyloid P-component (APCS) was the most down-regulated protein. These 29 proteins were categorized based on binding, catalytic activity and enzyme regulator activity. The biological roles were involved in biological regulation, metabolic processes, cellular processes and response to a stimulus. Based on ELISA, the median concentrations of CP and complement factor B (CFB) were 332.3 and 412.3 ng/mL, respectively. The concentrations of CP and CFB were significantly higher in the DS group than in the control group (P < 0.05). In conclusion, proteomic approaches offer the possibility of further improving the performance of DS screening and our identification of up- and down-regulated proteins may lead to new candidates for DS screening. PMID:22678011

  6. Heterocomplexes of mannose-binding lectin and the pentraxins PTX3 or serum amyloid P component trigger cross-activation of the complement system

    DEFF Research Database (Denmark)

    Ma, Ying Jie; Doni, Andrea

    2011-01-01

    The long pentraxin 3 (PTX3), serum amyloid P component (SAP), and C-reactive protein belong to the pentraxin family of pattern recognition molecules involved in tissue homeostasis and innate immunity. They interact with C1q from the classical complement pathway. Whether this also occurs via the analogous mannose-binding lectin (MBL) from the lectin complement pathway is unknown. Thus, we investigated the possible interaction between MBL and the pentraxins. We report that MBL bound PTX3 and SAP partly via its collagen-like domain but not C-reactive protein. MBL-PTX3 complex formation resulted in recruitment of C1q, but this was not seen for the MBL-SAP complex. However, both MBL-PTX3 and MBL-SAP complexes enhanced C4 and C3 deposition and opsonophagocytosis of Candida albicans by polymorphonuclear leukocytes. Interaction between MBL and PTX3 led to communication between the lectin and classical complement pathways via recruitment of C1q, whereas SAP-enhanced complement activation occurs via a hitherto unknown mechanism. Taken together, MBL-pentraxin heterocomplexes trigger cross-activation of the complement system.

  7. Isolation and purification of two major serum amyloid A isotypes SAA1 and SAA2 from the acute phase plasma of mice.

    Science.gov (United States)

    Kaplan, B; Yakar, S; Balta, Y; Pras, M; Martin, B

    1997-12-19

    A new procedure was developed for isolation of two major serum amyloid A (SAA) isotypes SAA1 and SAA2 from acute-phase plasma of mice. The procedure included preparation of high-density lipoproteins (HDLs) and their separation by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The SAA proteins (Mr 12,000) were electroeluted and afterwards purified from SDS by gel permeation chromatography on a Fractogel TSK-40F column in aqueous 50% acetonitrile-0.1% TFA. Finally, the SAA proteins free from SDS were fractionated by high-performance liquid chromatography on a Vydac 214TP54 column (250 x 4.6 mm I.D., particle size 5 microm), yielding two major fractions with k=5.2 and k=5.5. The N- and C-terminal sequence analyses and mass spectrometry demonstrated the purity of these two major fractions and their identity with apo SAA1 (k=5.2) and apo SAA2 (k=5.5). The developed procedure is applicable to small amounts of pooled murine plasma (6-7 ml) and could be readily modified from small to large scale preparations. PMID:9518179

  8. Posttranscriptional regulation of acute phase serum amyloid A2 expression by the 5'- and 3'-untranslated regions of its mRNA.

    Science.gov (United States)

    Longley, D B; Steel, D M; Whitehead, A S

    1999-10-15

    Human acute-phase serum amyloid A protein (A-SAA) is a major acute phase reactant, the concentration of which increases dramatically as part of the body's early response to inflammation. A-SAA is the product of two almost identical genes, SAA1 and SAA2, which are induced by the pro-inflammatory cytokines, IL-1 and IL-6. In this study, we examine the roles played by the 5'- and 3'-untranslated regions (UTRs) of the SAA2 mRNA in regulating A-SAA2 expression. SAA2 promoter-driven luciferase reporter gene constructs carrying the SAA2 5'-UTR and/or 3'-UTR were transiently transfected into the HepG2 human hepatoma cell line. After induction of chimeric mRNA with IL-1beta and IL-6, the SAA2 5'- and 3'-UTRs were both able to posttranscriptionally modify the expression of the luciferase reporter. The SAA2 5'-UTR promotes efficient translation of the chimeric luciferase transcripts, whereas the SAA2 3'-UTR shares this property and also significantly accelerates the rate of reporter mRNA degradation. Our data strongly suggest that the SAA2 5'- and 3'-UTRs each play significant independent roles in the posttranscriptional regulation of A-SAA2 protein synthesis. PMID:10510397

  9. Estrogen protects neuronal cells from amyloid beta-induced apoptosis via regulation of mitochondrial proteins and function

    Directory of Open Access Journals (Sweden)

    Iwamoto Sean

    2006-11-01

    Full Text Available Abstract Background Neurodegeneration in Alzheimer's disease is associated with increased apoptosis and parallels increased levels of amyloid beta, which can induce neuronal apoptosis. Estrogen exposure prior to neurotoxic insult of hippocampal neurons promotes neuronal defence and survival against neurodegenerative insults including amyloid beta. Although all underlying molecular mechanisms of amyloid beta neurotoxicity remain undetermined, mitochondrial dysfunction, including altered calcium homeostasis and Bcl-2 expression, are involved in neurodegenerative vulnerability. Results In this study, we investigated the mechanism of 17?-estradiol-induced prevention of amyloid beta-induced apoptosis of rat hippocampal neuronal cultures. Estradiol treatment prior to amyloid beta exposure significantly reduced the number of apoptotic neurons and the associated rise in resting intracellular calcium levels. Amyloid beta exposure provoked down regulation of a key antiapoptotic protein, Bcl-2, and resulted in mitochondrial translocation of Bax, a protein known to promote cell death, and subsequent release of cytochrome c. E2 pretreatment inhibited the amyloid beta-induced decrease in Bcl-2 expression, translocation of Bax to the mitochondria and subsequent release of cytochrome c. Further implicating the mitochondria as a target of estradiol action, in vivo estradiol treatment enhanced the respiratory function of whole brain mitochondria. In addition, estradiol pretreatment protected isolated mitochondria against calcium-induced loss of respiratory function. Conclusion Therefore, we propose that estradiol pretreatment protects against amyloid beta neurotoxicity by limiting mitochondrial dysfunction via activation of antiapoptotic mechanisms.

  10. Identification and characterisation of the novel amyloid-beta peptide-induced protein p17.

    Science.gov (United States)

    Nehar, Saheen; Mishra, Manisha; Heese, Klaus

    2009-10-01

    Amyloid-beta peptide (Abeta) achieves neurodegeneration through unknown mechanisms. To elucidate some of these mechanisms, we conducted a cDNA subtraction analysis of Abeta-mediated neurotoxicity in neuronal cells and observed an up-regulation of the novel gene p17. The p17 protein was also found elevated in Alzheimer's disease (AD) mouse model. Here, we characterised p17 primarily in cell lines with respect to its localisation, function and physiological expression. We discovered that p17 acts downstream of protein kinase C and inhibits the tyrosine receptor kinase B-brain-derived neurotrophic factor (TrkB-BDNF) pathway. It impedes survival factors and enhances amyloid precursor protein expression thus suggesting its involvement in the Abeta-mediated pro-apoptotic pathways in AD. PMID:19755123

  11. Hydration, cavities and volume in protein folding, aggregation and amyloid assembly

    International Nuclear Information System (INIS)

    Differential hydration dictates various biological processes, including protein folding, ligand binding, macromolecular assembly, enzyme kinetics and signal transduction. If water is partially or totally removed (experimentally or in silico), the outcome of these processes can be significantly affected. The aggregation of proteins into amyloids or other aggregate forms also results in profound changes in hydration. High hydrostatic pressure is a unique tool to study hydration, as increases in water binding usually lead to decreases in volume. Pressure changes can favor the formation or disassembly of amyloids depending on the volume changes associated with protein folding and misfolding/aggregation. The packing and formation of cavities will also contribute to changes in volume, and therefore, to sensitivity to pressure. Therefore, the formation of water-excluding cavities is predicted to be an important event in folding and aggregation landscapes

  12. Differential regulation of amyloid-?-protein mRNA expression within hippocampal neuronal subpopulations in Alzheimer disease

    International Nuclear Information System (INIS)

    The authors have mapped the neuroanatomical distribution of amyloid-?-protein mRNA within neuronal subpopulations of the hippocampal formation in the cynomolgus monkey (Macaca fascicularis), normal aged human, and patients with Alzheimer disease. Amyloid-?-protein mRNA appears to be expressed in all hippocampal neurons, but at different levels of abundance. In the central nervous system of monkey and normal aged human, image analysis shows that neurons of the dentate gyrus and cornu Ammonis fields contain a 2.5-times-greater hybridization signal than is present in neurons of the subiculum and entorhinal cortex. In contrast, in the Alzheimer disease hippocampal formation, the levels of amyloid-?-protein mRNA in the cornu Ammonis field 3 and parasubiculum are equivalent. These findings suggest that within certain neuronal subpopulations cell type-specific regulation of amyloid-?-protein gene expression may be altered in Alzheimer disease

  13. Feasibility of Predicting MCI/AD Using Neuropsychological Tests and Serum ?-Amyloid

    OpenAIRE

    Luis, Cheryl A.; Abdullah, Laila; Ait-ghezala, Ghania; Mouzon, Benoit; Keegan, Andrew P.; Crawford, Fiona; Mullan, Michael

    2011-01-01

    We examined the usefulness of brief neuropsychological tests and serum A? as a predictive test for detecting MCI/AD in older adults. Serum A? levels were measured from 208 subjects who were cognitively normal at enrollment and blood draw. Twenty-eight of the subjects subsequently developed MCI (n = 18) or AD (n = 10) over the follow-up period. Baseline measures of global cognition, memory, language fluency, and serum A?1–42 and the ratio of serum A?1–42/A?1–40 were significant pred...

  14. Human neurons derived from a teratocarcinoma cell line express solely the 695-amino acid amyloid precursor protein and produce intracellular beta-amyloid or A4 peptides.

    OpenAIRE

    Wertkin, A M; Turner, R. S.; Pleasure, S J; Golde, T. E.; Younkin, S. G.; Trojanowski, J. Q.; Lee, V M

    1993-01-01

    The beta-amyloid or beta/A4 peptides that accumulate as filamentous aggregates in the extracellular space of Alzheimer disease (AD) brains are derived from one or more alternatively spliced amyloid precursor proteins (APPs). The more abundant APPs in the central nervous system are the 695-(APP695), 751- (APP751), and 770- (APP770) amino acid isoforms, and each could be the source of beta/A4 peptide that accumulates in the AD brain. It is plausible that altered metabolism of these APPs by cent...

  15. Molecular simulations of beta-amyloid protein near hydrated lipids (PECASE).

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, Aidan Patrick; Han, Kunwoo (Texas A& M University, College Station, TX); Ford, David M. (Texas A& M University, College Station, TX)

    2005-12-01

    We performed molecular dynamics simulations of beta-amyloid (A{beta}) protein and A{beta} fragment(31-42) in bulk water and near hydrated lipids to study the mechanism of neurotoxicity associated with the aggregation of the protein. We constructed full atomistic models using Cerius2 and ran simulations using LAMMPS. MD simulations with different conformations and positions of the protein fragment were performed. Thermodynamic properties were compared with previous literature and the results were analyzed. Longer simulations and data analyses based on the free energy profiles along the distance between the protein and the interface are ongoing.

  16. Determination of the Oligomer Size of Amyloidogenic Protein ?-Amyloid(1–40) by Single-Molecule Spectroscopy

    OpenAIRE

    Ding, Hao; Wong, Pamela T.; Lee, Edgar L.; Gafni, Ari; Steel, Duncan G.

    2009-01-01

    Amyloid diseases are traditionally characterized by the appearance of inter- and intracellular fibrillar protein deposits, termed amyloid. Historically, these deposits have been thought to be the etiology of the disease. However, recent evidence suggests that small oligomers of the amyloidogenic protein/peptide are the origin of neurotoxicity. Although the importance of identifying the toxic oligomeric species is widely recognized, such identification is challenging because these oligomers ar...

  17. Multiplexed microfluidic quantification of proteins in serum

    Science.gov (United States)

    Rajan, Nitin; Rajauria, Sukumar; Cleland, Andrew

    2015-03-01

    Rapid and low cost immunoassays targeting proteins in blood or other bodily fluids are highly sought after for point-of-care devices and early screening of patients. Immunoturbidimetric assays utilize latex particles functionalized with antibodies, with particle aggregation in the presence of the analyte detected by a change in absorbance. Using a high throughput micro-fluidic particle analyzer based solely on electrical signals (resistive pulse sensing), we are able to accurately quantify the degree of aggregation by analyzing the changes in the particle size distribution. Thus we study the aggregation of streptavidin (SAv) coated beads in the presence of biotinylated bovine serum albumin as a proof-of-principle assay and extract the binding capacity of the SAv beads from the dose-response curve. We also use our aggregation measurement platform to characterize a commercial C-reactive protein (CRP) immunoturbidimetric assay (hsCRP, Diazyme Inc.). We obtain a linear calibration curve as well as a better limit of detection of CRP than that obtained by absorbance measurements. By using different bead sizes functionalized with different antibodies, multiplexed analyte detection is also possible. We demonstrate this by combining the commercial anti-CRP functionalized beads (0.4 microns) with biotin coated beads (1.0 microns), and carry out the simultaneous detection of SAv and CRP in a single sample.

  18. 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.)

  19. Proteolytic processing of Alzheimer's disease beta A4 amyloid precursor protein in human platelets.

    Science.gov (United States)

    Li, Q X; Evin, G; Small, D H; Multhaup, G; Beyreuther, K; Masters, C L

    1995-06-01

    The processing of amyloid precursor protein (APP) and production of beta A4 amyloid are events likely to influence the development and progression of Alzheimer's disease, since beta A4 is the major constituent of amyloid deposited in this disorder. Our previous studies showed that human platelets contain full-length APP (APPFL) and are a suitable substrate to study normal APP processing. In the present study, we show that a 22-kDa beta A4-containing carboxyl-terminal fragment (22-CTF) of APP is present in unstimulated platelets. Both APPFL and 22-CTF are proteolytically degraded when platelets are activated with thrombin, collagen, or calcium ionophore A23187. Complete cleavage of APPFL and 22-CTF require the presence of extracellular calcium. Following stimulation in the presence of calcium, a new CTF of 17 kDa is generated, and the NH2-terminal epitope of beta A4 amyloid is lost. Preincubation of platelets with the cell-permeable cysteine protease inhibitors calpeptin, (2S,3S)-trans-epoxysuccinyl-L-leucyl-amido-3-methylbutane ethyl ester (E64d), Na alpha-p-tosyl-L-lysine chloromethyl ketone, or calcium chelator EGTA before platelet stimulation inhibits the degradation of both APPFL and 22-CTF. Divalent metal ions including zinc, copper, and cobalt inhibit the degradation of APPFL and 22-CTF. This study suggests that a calcium-dependent neutral cysteine protease is involved in the proteolytic processing of an amyloidogenic species of APP in human platelets. PMID:7775475

  20. Ginsenoside Rh2 promotes nonamyloidgenic cleavage of amyloid precursor protein via a cholesterol-dependent pathway.

    Science.gov (United States)

    Qiu, J; Li, W; Feng, S H; Wang, M; He, Z Y

    2014-01-01

    Ginsenoside Rh2 (Rh2) is a ginseng derivative used in Chinese traditional medicine. We investigated whether Rh2 can help prevent Alzheimer's disease symptoms and examined underlying mechanisms. We injected Rh2 into tg2576 Alzheimer's disease model mice and looked for behavioral improvement and senile plaque reduction in brain slices. We measured amyloid precursor protein (APP) metabolism species changes, amyloid beta40 and 42 levels and ?, ? secretase activity in primary hippocampal neurons. By living cell staining, we detected surface and endocytosed APP. We also measured cholesterol and lipid rafts in primary neurons. Rh2 treatment significantly improved learning and memory performance at 14 months of age; it also reduced brain senile plaques at this age. Based on in vitro experiments, we found that Rh2 treatment increased soluble APP? (sAPP?) levels, increased CTF?/? ratios, and reduced amyloid beta 40 and 42 concentrations. Surface APP levels dramatically increased. Based on living cell staining, we found that Rh2 inhibited APP endocytosis. Based on lipid removal and reload experiments, we found that Rh2 can modulate APP by reducing cholesterol and lipid raft levels. We concluded that Rh2 improves learning and memory function in Alzheimer's disease model mice, and that this improvement is accomplished by reducing amyloid beta secretion and APP endocytosis, which in turn is achieved by reducing cholesterol and lipid raft concentrations. PMID:24854439

  1. Brain amyloid ? protein and memory disruption in Alzheimer’s disease

    Directory of Open Access Journals (Sweden)

    Weiming Xia

    2010-09-01

    Full Text Available 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 severity remains to be discovered. Oligomeric A? has been detected in cultured cells, rodent and human brains, as well as human cerebrospinal fluid. Synthetic, cell, and brain derived A? oligomers have been found to inhibit hippocampal long-term potentiation (LTP and this effect can be suppressed by the blockage of A? oligomer formation. A large body of evidence suggests that A? oligomers inhibit N-methyl-D-aspartate receptor dependent LTP; additional receptors have also been found to elicit downstream pathways upon binding to A? oligomers. Amyloid antibodies and small molecular compounds that reduce brain A? levels and block A? oligomer formation are capable of reversing synaptic dysfunction and these approaches hold a promising therapeutic potential to rescue memory disruption.Keywords: Alzheimer, amyloid, oligomer, long-term potentiation, NMDA

  2. Inhibition of glycosaminoglycan synthesis and protein glycosylation with WAS-406 and azaserine result in reduced islet amyloid formation in vitro.

    Science.gov (United States)

    Hull, Rebecca L; Zraika, Sakeneh; Udayasankar, Jayalakshmi; Kisilevsky, Robert; Szarek, Walter A; Wight, Thomas N; Kahn, Steven E

    2007-11-01

    Deposition of islet amyloid polypeptide (IAPP) as amyloid in the pancreatic islet occurs in approximately 90% of individuals with Type 2 diabetes and is associated with decreased islet beta-cell mass and function. Human IAPP (hIAPP), but not rodent IAPP, is amyloidogenic and toxic to islet beta-cells. In addition to IAPP, islet amyloid deposits contain other components, including heparan sulfate proteoglycans (HSPGs). The small molecule 2-acetamido-1,3,6-tri-O-acetyl-2,4-dideoxy-alpha-D-xylo-hexopyranose (WAS-406) inhibits HSPG synthesis in hepatocytes and blocks systemic amyloid A deposition in vivo. To determine whether WAS-406 inhibits localized amyloid formation in the islet, we incubated hIAPP transgenic mouse islets for up to 7 days in 16.7 mM glucose (conditions that result in amyloid deposition) plus increasing concentrations of the inhibitor. WAS-406 at doses of 0, 10, 100, and 1,000 microM resulted in a dose-dependent decrease in amyloid deposition (% islet area occupied by amyloid: 0.66 +/- 0.14%, 0.10 +/- 0.06%, 0.09 +/- 0.07%, and 0.004 +/- 0.003%, P Azaserine, an inhibitor of the rate-limiting step of the hexosamine biosynthesis pathway, replicated the effects of WAS-406, resulting in reduction of O-linked protein glycosylation and glycosaminoglycan synthesis and inhibition of islet amyloid formation. In summary, interventions that decrease both glycosaminoglycan synthesis and O-linked protein glycosylation are effective in reducing islet amyloid formation, but their utility as pharmacological agents may be limited due to adverse effects on the islet. PMID:17804609

  3. Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

    International Nuclear Information System (INIS)

    Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218–289) and ?-synuclein yielded 88–97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77–90 % correctness if also assignments classified as tentative by the algorithm are included

  4. Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, Elena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Gath, Julia [ETH Zurich, Physical Chemistry (Switzerland); Habenstein, Birgit [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Ravotti, Francesco; Szekely, Kathrin; Huber, Matthias [ETH Zurich, Physical Chemistry (Switzerland); Buchner, Lena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Boeckmann, Anja, E-mail: a.bockmann@ibcp.fr [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Meier, Beat H., E-mail: beme@ethz.ch [ETH Zurich, Physical Chemistry (Switzerland); Guentert, Peter, E-mail: guentert@em.uni-frankfurt.de [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany)

    2013-07-15

    Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218-289) and {alpha}-synuclein yielded 88-97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77-90 % correctness if also assignments classified as tentative by the algorithm are included.

  5. Investigation of amyloid deposition in uterine leiomyoma patients

    Directory of Open Access Journals (Sweden)

    Jinping Liu

    2012-08-01

    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.

  6. Soluble amyloid precursor protein ? as blood-based biomarker of Alzheimer's disease

    OpenAIRE

    Perneczky, R.; Guo, L-H; Kagerbauer, S M; Werle, L; Kurz, A.; Martin, J.; Alexopoulos, P.

    2013-01-01

    The aim of this study was to explore concentrations differences of soluble amyloid precursor protein (sAPP) ? and ? in blood plasma in patients with probable Alzheimer's disease (AD) and cognitively healthy age-matched control subjects, as well as patients with behavioural variant frontotemporal dementia (bvFTD). Concentrations of sAPP? and ? were measured using enzyme-linked immunosorbent assay technology in 80 patients with probable AD, 37 age-matched control subjects and 14 patients with b...

  7. Thyroid hormone negatively regulates the transcriptional activity of the ?-amyloid precursor protein gene

    OpenAIRE

    Belandia, Borja; Latasa, María Jesús; Villa, Ana; Pascual, Ángel

    1998-01-01

    The expression of the ?-amyloid precursor protein (APP), which plays a key role in the development of Alzheimer's disease, is regulated by a variety of cellular mediators in a cell-dependent manner. In the present study, we present evidence that thyroid hormones negatively regulate the expression of the APP gene in neuroblastoma cells. Transient transfection studies using plasmids that contain progressive deletions of the 5' region of the gene demonstrate that triiodothyronine (T3), the more ...

  8. A peptide zipcode sufficient for anterograde transport within amyloid precursor protein

    OpenAIRE

    Satpute-krishnan, Prasanna; Degiorgis, Joseph A.; Conley, Michael P.; Jang, Marcus; Bearer, Elaine L.

    2006-01-01

    Fast anterograde transport of membrane-bound organelles delivers molecules synthesized in the neuronal cell body outward to distant synapses. Identification of the molecular “zipcodes” on organelles that mediate attachment and activation of microtubule-based motors for this directed transport is a major area of inquiry. Here we identify a short peptide sequence (15 aa) from the cytoplasmic C terminus of amyloid precursor protein (APP-C) sufficient to mediate the anterograde transport of p...

  9. The Drosophila Homologue of the Amyloid Precursor Protein Is a Conserved Modulator of Wnt PCP Signaling

    OpenAIRE

    Soldano, Alessia; Okray, Zeynep; Janovska, Pavlina; Tmejova?, Kater?ina; Reynaud, Elodie; Claeys, Annelies; Yan, Jiekun; Atak, Zeynep Kalender; Strooper, Bart; Dura, Jean-maurice; Bryja, Vi?te?zslav; Hassan, Bassem A.

    2013-01-01

    Wnt Planar Cell Polarity (PCP) signaling is a universal regulator of polarity in epithelial cells, but it regulates axon outgrowth in neurons, suggesting the existence of axonal modulators of Wnt-PCP activity. The Amyloid precursor proteins (APPs) are intensely investigated because of their link to Alzheimer's disease (AD). APP's in vivo function in the brain and the mechanisms underlying it remain unclear and controversial. Drosophila possesses a single APP homologue called APP Like, or APPL...

  10. Structural dynamics of the ?E22 (Osaka) familial Alzheimer's disease-linked amyloid ?-protein

    OpenAIRE

    Inayathullah, Mohammed; Teplow, David B.

    2011-01-01

    A familial form of Alzheimer disease (AD) recently was described in a kindred in Osaka, Japan. This kindred possesses an amyloid ?-protein (A?) precursor mutation within the A? coding region that results in the deletion of Glu22 (?E22). We report here results of studies of [?E22]A?40 and ?E22A?42 that sought to elucidate the conformational dynamics, oligomerization behavior, fibril formation kinetics, fibril morphology, and fibril stability of these mutant peptides. Both [?E22]A? pe...

  11. Familial Alzheimer's disease mutations alter the stability of the amyloid ?-protein monomer folding nucleus

    OpenAIRE

    Grant, Marianne A.; Lazo, Noel D.; Lomakin, Aleksey; Condron, Margaret M.; Arai, Hiromi; Yamin, Ghiam; Rigby, Alan C.; Teplow, David B.

    2007-01-01

    Amyloid ?-protein (A?) oligomers may be the proximate neurotoxins in Alzheimer's disease (AD). Recently, to elucidate the oligomerization pathway, we studied A? monomer folding and identified a decapeptide segment of A?, 21Ala–22Glu–23Asp–24Val–25Gly–26Ser–27Asn–28Lys–29Gly–30Ala, within which turn formation appears to nucleate monomer folding. The turn is stabilized by hydrophobic interactions between Val-24 and Lys-28 and by long-range electrostatic interactions betwee...

  12. Discrete Molecular Dynamics Study of Oligomer Formation by N-Terminally Truncated Amyloid ?-Protein

    OpenAIRE

    Meral, Derya; Urbanc, Brigita

    2013-01-01

    In Alzheimer’s disease (AD), amyloid ?-protein (A?) self–assembles into toxic oligomers. Of the two predominant A? alloforms, A?1–40 and A?1–42, the latter is particularly strongly linked to AD. N-terminally truncated and pyroglutamated A? peptides were recently shown to seed A? aggregation and contribute significantly to A?–mediated toxicity, yet their folding and assembly were not explored computationally. Discrete molecular dynamics (DMD) approach previously captured in v...

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

    OpenAIRE

    Zhao Hong; Zhu Jinmin; Cui Kemi; Xu Xiaoyin; Brien Megan, O.; Wong Kelvin K; Kesari Santosh; Xia Weiming; Tc, Wong Stephen

    2009-01-01

    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-MB...

  14. Amyloid beta-protein activates tachykinin receptors and inositol trisphosphate accumulation by synergy with glutamate.

    OpenAIRE

    Kimura, H.; Schubert, D.

    1993-01-01

    The biological function of the soluble form of the amyloid beta-protein (ABP) was examined by assaying its interaction with neuronal receptors expressed in Xenopus oocytes. ABP weakly activated tachykinin receptors, but in the presence of N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methylisoxazole-4- propionate-type glutamate receptors ABP-induced responses were greatly enhanced. Glutamate and ABP together also induced accumulation of inositol trisphosphate and increases in intracellular...

  15. Increased T cell reactivity to amyloid ? protein in older humans and patients with Alzheimer disease

    OpenAIRE

    Monsonego, Alon; Zota, Victor; Karni, Arnon; Krieger, Jeffery I.; Bar-Or, Amit; Bitan, Gal; Budson, Andrew E.; Sperling, Reisa; Selkoe, Dennis J.; Weiner, Howard L.

    2003-01-01

    Alzheimer disease (AD) is characterized by the progressive deposition of the 42-residue amyloid ? protein (A?) in brain regions serving memory and cognition. In animal models of AD, immunization with A? results in the clearance of A? deposits from the brain. However, a trial of vaccination with synthetic human A?1–42 in AD resulted in the development of meningoencephalitis in some patients. We measured cellular immune responses to A? in middle-aged and elderly healthy subjects and in patients...

  16. 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)

    Glen D. Armstrong

    2011-08-01

    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.

  17. Insights into the physiological function of the ?-amyloid precursor protein: beyond Alzheimer's disease

    Science.gov (United States)

    Dawkins, Edgar; Small, David H

    2014-01-01

    The ?-amyloid precursor protein (APP) has been extensively studied for its role as the precursor of the ?-amyloid protein (A?) of Alzheimer's disease. However, the normal function of APP remains largely unknown. This article reviews studies on the structure, expression and post-translational processing of APP, as well as studies on the effects of APP in vitro and in vivo. We conclude that the published data provide strong evidence that APP has a trophic function. APP is likely to be involved in neural stem cell development, neuronal survival, neurite outgrowth and neurorepair. However, the mechanisms by which APP exerts its actions remain to be elucidated. The available evidence suggests that APP interacts both intracellularly and extracellularly to regulate various signal transduction mechanisms. This article reviews studies on the structure, expression and post-translational processing of ?-amyloid precursor protein (APP), as well as studies on the effects of APP in vitro and in vivo. We conclude that the published data provide strong evidence that APP has a trophic function. APP is likely to be involved in neural stem cell development, neuronal survival, neurite outgrowth and neurorepair. However, the mechanisms by which APP exerts its actions remain to be elucidated. The available evidence suggests that APP interacts both intracellularly and extracellularly to regulate various signal transduction mechanisms. PMID:24517464

  18. Amyloid precursor protein (APP) affects global protein synthesis in dividing human cells.

    Science.gov (United States)

    Sobol, Anna; Galluzzo, Paola; Liang, Shuang; Rambo, Brittany; Skucha, Sylvia; Weber, Megan J; Alani, Sara; Bocchetta, Maurizio

    2015-05-01

    Hypoxic non-small cell lung cancer (NSCLC) is dependent on Notch-1 signaling for survival. Targeting Notch-1 by means of ?-secretase inhibitors (GSI) proved effective in killing hypoxic NSCLC. Post-mortem analysis of GSI-treated, NSCLC-burdened mice suggested enhanced phosphorylation of 4E-BP1 at threonines 37/46 in hypoxic tumor tissues. In vitro dissection of this phenomenon revealed that Amyloid Precursor Protein (APP) inhibition was responsible for a non-canonical 4E-BP1 phosphorylation pattern rearrangement-a process, in part, mediated by APP regulation of the pseudophosphatase Styx. Upon APP depletion we observed modifications of eIF-4F composition indicating increased recruitment of eIF-4A to the mRNA cap. This phenomenon was supported by the observation that cells with depleted APP were partially resistant to silvestrol, an antibiotic that interferes with eIF-4A assembly into eIF-4F complexes. APP downregulation in dividing human cells increased the rate of global protein synthesis, both cap- and IRES-dependent. Such an increase seemed independent of mTOR inhibition. After administration of Torin-1, APP downregulation and Mechanistic Target of Rapamycin Complex 1 (mTORC-1) inhibition affected 4E-BP1 phosphorylation and global protein synthesis in opposite fashions. Additional investigations indicated that APP operates independently of mTORC-1. Key phenomena described in this study were reversed by overexpression of the APP C-terminal domain. The presented data suggest that APP may be a novel regulator of protein synthesis in dividing human cells, both cancerous and primary. Furthermore, APP appears to affect translation initiation using mechanisms seemingly dissimilar to mTORC-1 regulation of cap-dependent protein synthesis. PMID:25283437

  19. Powerful beneficial effects of benfotiamine on cognitive impairment and beta-amyloid deposition in amyloid precursor protein/presenilin-1 transgenic mice.

    Science.gov (United States)

    Pan, Xiaoli; Gong, Neng; Zhao, Jing; Yu, Zhe; Gu, Fenghua; Chen, Jia; Sun, Xiaojing; Zhao, Lei; Yu, Meijing; Xu, Zhiru; Dong, Wenxin; Qin, Yan; Fei, Guoqiang; Zhong, Chunjiu; Xu, Tian-Le

    2010-05-01

    Reduction of glucose metabolism in brain is one of the main features of Alzheimer's disease. Thiamine (vitamin B1)-dependent processes are critical in glucose metabolism and have been found to be impaired in brains from patients with Alzheimer's disease. However, thiamine treatment exerts little beneficial effect in these patients. Here, we tested the effect of benfotiamine, a thiamine derivative with better bioavailability than thiamine, on cognitive impairment and pathology alterations in a mouse model of Alzheimer's disease, the amyloid precursor protein/presenilin-1 transgenic mouse. We show that after a chronic 8 week treatment, benfotiamine dose-dependently enhanced the spatial memory of amyloid precursor protein/presenilin-1 mice in the Morris water maze test. Furthermore, benfotiamine effectively reduced both amyloid plaque numbers and phosphorylated tau levels in cortical areas of the transgenic mice brains. Unexpectedly, these effects were not mimicked by another lipophilic thiamine derivative, fursultiamine, although both benfotiamine and fursultiamine were effective in increasing the levels of free thiamine in the brain. Most notably, benfotiamine, but not fursultiamine, significantly elevated the phosphorylation level of glycogen synthase kinase-3alpha and -3beta, and reduced their enzymatic activities in the amyloid precursor protein/presenilin-1 transgenic brain. Therefore, in the animal Alzheimer's disease model, benfotiamine appears to improve the cognitive function and reduce amyloid deposition via thiamine-independent mechanisms, which are likely to include the suppression of glycogen synthase kinase-3 activities. These results suggest that, unlike many other thiamine-related drugs, benfotiamine may be beneficial for clinical Alzheimer's disease treatment. PMID:20385653

  20. Presenile dementia and cerebral haemorrhage linked to a mutation at codon 692 of the beta-amyloid precursor protein gene.

    Science.gov (United States)

    Hendriks, L; van Duijn, C M; Cras, P; Cruts, M; Van Hul, W; van Harskamp, F; Warren, A; McInnis, M G; Antonarakis, S E; Martin, J J

    1992-06-01

    Several families with an early-onset form of familial Alzheimer's disease have been found to harbour mutations at a specific codon (717) of the gene for the beta-amyloid precursor protein (APP) on chromosome 21. We now report, a novel base mutation in the same exon of the APP gene which co-segregates in one family with presenile dementia and cerebral haemorrhage due to cerebral amyloid angiopathy. The mutation results in the substitution of alanine into glycine at codon 692. These results suggest that the clinically distinct entities, presenile dementia and cerebral amyloid angiopathy, can be caused by the same mutation in the APP gene. PMID:1303239

  1. In equine grass sickness, serum amyloid A and fibrinogen are elevated, and can aid differential diagnosis from non-inflammatory causes of colic.

    Science.gov (United States)

    Copas, V E N; Durham, A E; Stratford, C H; McGorum, B C; Waggett, B; Pirie, R S

    2013-04-13

    Equine grass sickness (EGS) is a debilitating and often fatal neurodegenerative disease. A presumptive diagnosis of EGS may be made on the basis of clinical signs and subjective ancillary tests, but a definitive antemortem diagnosis can only be made following histopathological examination of intestinal biopsies. It has previously been reported that horses with EGS may show clinical and clinicopathological signs of systemic inflammation. The objective of this study was to (a) quantify acute inflammatory markers in blood samples collected from acute, subacute and chronic EGS cases, and (b) compare them with (i) clinically normal horses co-grazing with acute EGS cases (co-grazers), (ii) horses with other causes of colic and (iii) healthy horses. Serum amyloid A (SAA), serum activin A and plasma fibrinogen were quantified. There were marked increases in SAA and fibrinogen in EGS cases compared with healthy horses, co-grazers and non-inflammatory colic cases. The concentrations of SAA and fibrinogen in EGS cases were not significantly different from inflammatory colic cases. When concentrations of SAA, fibrinogen and activin A in each EGS subgroup were compared, no significant differences were detected. Activin A concentrations were significantly elevated in EGS cases and co-grazing horses; this could reflect the presence of subclinical disease in some horses that do not develop clinical signs of EGS, and suggests widespread exposure to the aetiological agent. When faced with sparse antemortem diagnostic techniques, identification of marked increases in acute phase protein concentrations may help to differentiate EGS from other causes of abdominal pain, such as intestinal obstructions; however, there could be diagnostic difficulty in differentiating other inflammatory abdominal conditions, such as peritonitis or enteritis. PMID:23428423

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

    Directory of Open Access Journals (Sweden)

    N. Elizabeth Pryor

    2012-03-01

    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.

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

    OpenAIRE

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

    1991-01-01

    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 ...

  4. ?-Amyloid Impairs AMPA Receptor Trafficking and Function by Reducing Ca2+/Calmodulin-dependent Protein Kinase II Synaptic Distribution*

    OpenAIRE

    Gu, Zhenglin; Liu, Wenhua; Yan, Zhen

    2009-01-01

    A fundamental feature of Alzheimer disease (AD) is the accumulation of ?-amyloid (A?), a peptide generated from the amyloid precursor protein (APP). Emerging evidence suggests that soluble A? oligomers adversely affect synaptic function, which leads to cognitive failure associated with AD. The A?-induced synaptic dysfunction has been attributed to the synaptic removal of ?-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors (AMPARs); however, i...

  5. Beta-amyloid protein-containing inclusions in skeletal muscle of apolipoprotein-E-deficient mice.

    Science.gov (United States)

    Robertson, T A; Dutton, N S; Martins, R N; Roses, A D; Kakulas, B A; Papadimitriou, J M

    1997-02-01

    The tibialis anterior muscle and soleus muscle of apolipoprotein-E-deficient mice were examined by light and electron microscopy. By light microscopy, sarcoplasmic inclusions were seen in tibialis anterior muscle and 40% of type 2 myofibers were affected in all animals over 8 months of age. These inclusions reacted for nonspecific esterase, cytochrome oxidase, and myoadenylate deaminase and were also periodic acid Schiff positive and stained basophilic with hematoxylin. Moreover, they reacted immunocytochemically with an antibody specific to fragment 17 to 24 of the published sequence of Alzheimer's cerebrovascular amyloid peptide. Immunoreactivity was lost when the antibody was adsorbed with the appropriate synthetic peptide. Ultrastructurally, the inclusions consisted of tubular arrays and were similar to those observed in human muscle in several pathological conditions. In type 1 myofibers of both tibialis anterior and soleus muscle, however, mitochondrial abnormalities including an increase in their number and size were detected, but tubular aggregates were not seen. These large mitochondria possessed an electron-dense inner chamber with an increased number of tightly packed cristae. The results obtained suggest that in these mice there is a disturbed lipid metabolism in skeletal muscle fibers that manifests itself with an accumulation of phospholipid in the form of sarcoplasmic reticulum tubules in the type 2 fibers and enlarged mitochondria with tightly packed cristae in the type 1 fibers. In addition, beta-amyloid protein was closely associated with the accumulated tubules and vesicles of sarcoplasmic reticulum and may represent dysregulation of amyloid precursor protein metabolism. PMID:9033257

  6. Gastrodia elata modulates amyloid precursor protein cleavage and cognitive functions in mice.

    Science.gov (United States)

    Mishra, Manisha; Huang, Junjie; Lee, Yin Yeng; Chua, Doreen See Kin; Lin, Xiaoyan; Hu, Jiang-Miao; Heese, Klaus

    2011-01-01

    Gastrodia elata (Tianma) is a traditional Chinese medicine often used for the treatment of headache, convulsions, hypertension, and cardiovascular diseases. The vasodilatory actions of Tianma led us to investigate its specific effects on memory and learning as well as on Alzheimer's disease (AD)-related signaling. We conducted a radial arm water maze analysis and the novel object recognition test to assess the cognitive functions of Tianma-treated mice. Our data show that Tianma enhances cognitive functions in mice. Further investigations revealed that Tianma enhances the ?-secretase-mediated proteolytic processing of the amyloid precursor protein (App) that precludes the amyloid-? peptide production and supports the non-amyloidogenic processing of App which is favorable in AD treatment. We hypothesize that Tianma promotes cognitive functions and neuronal survival by inhibiting ?-site App-cleaving enzyme 1 activity and promoting the neuroprotective ?-secretase activity. PMID:21788698

  7. Focally elevated creatine detected in amyloid precursor protein (APP) transgenic mice and Alzheimer disease brain tissue.

    Science.gov (United States)

    Gallant, Meghan; Rak, Margaret; Szeghalmi, Adriana; Del Bigio, Marc R; Westaway, David; Yang, Jin; Julian, Robert; Gough, Kathleen M

    2006-01-01

    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. PMID:16267054

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

    Science.gov (United States)

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

    2012-06-12

    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

  9. Tuberous Sclerosis Protein 2 (TSC2) Modulates CCN4 Cytoprotection During Apoptotic Amyloid Toxicity in Microglia

    OpenAIRE

    Shang, Yan Chen; Chong, Zhao Zhong; Wang, Shaohui; Maiese, Kenneth

    2013-01-01

    More than 110 million individuals will suffer from cognitive loss worldwide by the year 2050 with a majority of individuals presenting with Alzheimer’s disease (AD). Yet, successful treatments for etiologies that involve ?-amyloid (A?) toxicity in AD remain elusive and await novel avenues for drug development. Here we show that Wnt1 inducible signaling pathway protein 1 (WISP1/CCN4) controls the post-translational phosphorylation of Akt1, p70S6K, and AMP activated protein kinase (AMPK) to the...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2011-12-31

    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.

  11. Correlation between some hematological parameters, acute phase proteins and serum immunoglobulins in experimental caprine besnoitiosis.

    Science.gov (United States)

    Hosseini, A; Namazi, F; Oryan, A; Nazifi, S

    2015-06-01

    The present study was undertaken to investigate correlation between some hematological parameters, acute phase proteins and immunoglobulins in the experimentally infected goats with Besnoitia caprae from the time of infection till 360 days post infection (DPI). Six male goats, approximately 12-16 months old, were inoculated subcutaneously with approximately 1.3 × 10(8) bradyzoites of B. caprae and blood samples were collected at weekly intervals from the jugular vein of the goats. Total leukocyte count and differential leukocyte counts were determined. Acute phase proteins (APPs) including serum amyloid A (SAA), haptoglobin (Hp), fibrinogen and ceruloplasmin were undertaken at weekly intervals. We evaluated an enzyme-linked immunosorbent assay (ELISA) (using a somatic antigen of bradyzoite) to detect anti-B. caprae antibodies in caprine sera. Cysts were present in the skin biopsies of the distal parts of the leg of the infected goats from 28 DPI. From 30 to 360 DPI, results showed that the APPs concentrations including SAA, Hp, fibrinogen and ceruloplasmin were enhanced in the serum of infected goats. However, there were some variation in hematological parameters; the differences were not significant with those of the normal values. Some variations were seen in the levels of specific antibodies against this parasite and they had correlation with some hematological parameters and acute-phase proteins. PMID:26063991

  12. Analyse struktureller Determinanten der toxischen Wirkung amyloider Proteine

    OpenAIRE

    Olzscha, Heidi

    2010-01-01

    Ein lebender Organismus ist unter anderem durch seine Fähigkeit zum präzisen Auf- und Zusammenbau höherer molekularer Strukturen charakterisiert, wobei die Faltung und Assemblierung von Proteinen eine bedeutende Rolle spielt. Die Proteinfaltung wird durch molekulare Chaperone unterstützt und optimiert, bis ein Protein seine native, biologisch funktionelle Struktur eingenommen hat. Durch exogene Einflüsse oder endogene Veränderungen eines Proteins, z.B. bei neurodegenerativen Erkrankunge...

  13. Influence of the protein context on the polyglutamine length-dependent elongation of amyloid fibrils.

    Science.gov (United States)

    Huynen, Céline; Willet, Nicolas; Buell, Alexander K; Duwez, Anne-Sophie; Jerôme, Christine; Dumoulin, Mireille

    2015-03-01

    Polyglutamine (polyQ) diseases, including Huntington's disease, are neurodegenerative disorders associated with the abnormal expansion of a polyQ tract within nine proteins. The polyQ expansion is thought to be a major determinant in the development of neurotoxicity, triggering protein aggregation into amyloid fibrils, although non-polyQ regions play a modulating role. In this work, we investigate the relative importance of the polyQ length, its location within a host protein, and the conformational state of the latter in the amyloid fibril elongation. Model polyQ proteins made of the ?-lactamase BlaP containing up to 79Q inserted at two different positions, and quartz crystal microbalance and atomic force microscopy were used for this purpose. We demonstrate that, independently of the polyQ tract location and the conformational state of the host protein, the relative elongation rate of fibrils increases linearly with the polyQ length. The slope of the linear fit is similar for both sets of chimeras (i.e., the elongation rate increases by ~1.9% for each additional glutamine), and is also similar to that previously observed for polyQ peptides. The elongation rate is, however, strongly influenced by the location of the polyQ tract within BlaP and the conformational state of BlaP. Moreover, comparison of our results with those previously reported for aggregation in solution indicates that these two parameters also modulate the ability of BlaP-polyQ chimeras to form the aggregation nucleus. Altogether our results suggest that non-polyQ regions are valuable targets in order to interfere with the process of amyloid fibril formation associated with polyQ diseases. PMID:25489872

  14. 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

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

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Ha Young, E-mail: hayoung@skku.edu [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Mitochondria Hub Regulation Center, Dong-A University, Busan 602-714 (Korea, Republic of); Kim, Sang Doo [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Baek, Suk-Hwan [Department of Biochemistry and Molecular Biology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Choi, Joon Hyuk [Department of Pathology, College of Medicine, Yeungnam University, Daegu 705-717 (Korea, Republic of); Cho, Kyung-Hyun [School of Biotechnology, Yeungnam University, Gyeongsan 712-749 (Korea, Republic of); Zabel, Brian A. [Palo Alto Institute for Research and Education, Veterans Affairs Hospital, Palo Alto, CA 94304 (United States); Bae, Yoe-Sik, E-mail: yoesik@skku.edu [Department of Biological Science, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Mitochondria Hub Regulation Center, Dong-A University, Busan 602-714 (Korea, Republic of); Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul 135-710 (Korea, Republic of)

    2013-03-29

    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.

  16. Elucidation of O-glycosylation structures of the ß-amyloid precursor protein by liquid chromatography - mass spectrometry using electron transfer dissociation and collision induced dissociation

    OpenAIRE

    Perdivara, Irina; Petrovich, Robert; Alliquant, Bernadette; Deterding, Leesa J.; Tomer, Kenneth B.; Przybylski, Michael

    2009-01-01

    Accumulation and deposition of ß-amyloid peptide, a major constituent in neuritic plaques are hallmarks of Alzheimer’s disease (AD) and AD-related neurodegenerative diseases. ß-Amyloid (Aß) is derived from the proteolytic cleavage of amyloid precursor protein (APP), a transmembrane protein present in three major isoforms in brain comprising 695, 751 and 770 amino acids, respectively. Among other post-translational modifications, APP is modified during maturation by N- and O- glycosylation, wh...

  17. Specific spatial learning deficits become severe with age in ?-amyloid precursor protein transgenic mice that harbor diffuse ?-amyloid deposits but do not form plaques

    OpenAIRE

    Koistinaho, Milla; Ort, Michael; Cimadevilla, Jose M.; Vondrous, Roman; Cordell, Barbara; Koistinaho, Jari; Bures, Jan; Higgins, Linda S.

    2001-01-01

    Memory impairment progressing to dementia is the main clinical symptom of Alzheimer's disease (AD). AD is characterized histologically by the presence of ?-amyloid (A?) plaques and neurofibrillary tangles in specific brain regions. Although A? derived from the A? precursor protein (?-APP) is believed to play a central etiological role in AD, it is not clear whether soluble and/or fibrillar forms are responsible for the memory deficit. We have generated and pre...

  18. 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)

    Kevin Hartman

    2013-02-01

    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.

  19. Spectroscopic imaging of serum proteins using quantum cascade lasers.

    Science.gov (United States)

    Mukherjee, Anadi; Bylund, Quentin; Prasanna, Manu; Margalit, Yotam; Tihan, Tarik

    2013-03-01

    First measurements of biomedical imaging using quantum cascade lasers (QCL) are presented. We report spectroscopic imaging of serum proteins using QCLs as an example for monitoring surface biocontamination. We found that dry smears of human serum can be spectroscopically imaged, identified, and quantified with high sensitivity and specificity. The core parts of the imaging platform consist of optically multiplexing three QCLs and an uncooled microbolometer camera. We show imaging of human serum proteins at 6.1, 9.25, and 9.5 ?m QCLs with high sensitivity and specificity. The sensitivity limit of 3???g/cm² of the human serum spot was measured at an S/N=3.The specificity of human serum detection was measured at 99% probability at a threshold of 77???g/cm². We anticipate our imaging technique to be a starting point for more sophisticated biomolecular diagnostic applications. PMID:23515866

  20. The first molluscan acute phase serum amyloid A (A-SAA) identified from oyster Crassostrea hongkongensis: molecular cloning and functional characterization.

    Science.gov (United States)

    Qu, Fufa; Xiang, Zhiming; Yu, Ziniu

    2014-08-01

    Serum amyloid A (SAA), a major evolutionarily conserved acute-phase protein, participates in many biological processes in eukaryotic cells, including innate immunity. However, little information regarding the relationship between SAA and innate immunity in mollusks is currently available. In this report, the first bivalve SAA (referred to as ChSAA) gene was identified and characterized from the Hong Kong oyster Crassostrea hongkongensis. Its full-length cDNA is 623 bp, including a 5'-UTR of 147 bp, a 3'-UTR of 56 bp containing a poly(A) tail and an open reading frame (ORF) of 420 bp that encodes a polypeptide of 139 amino acids. The predicted amino acid sequence of ChSAA comprises characteristic motifs of the SAA family, including a typical signal peptide and a conserved SAA domain. Comparison and phylogenetic analyses suggested that ChSAA shares a high identity to known acute-phase SAA proteins (A-SAAs). In addition, quantitative real-time PCR analysis revealed that ChSAA is constitutively expressed in all tissues examined, with the highest expression level in the mantle, and that its expression was acutely and significantly up-regulated in hemocytes following challenge by Vibrio alginolyticus (G(-)), Staphylococcus haemolyticus (G(+)) or Saccharomyces cerevisiae (fungus). Furthermore, over-expression of ChSAA via transfection with a ChSAA expression vector led to significantly increased NF-?B activity in HEK293T cells. These results suggest that ChSAA is likely to constitute a member of the A-SAA family involved in anti-pathogen responses in C. hongkongensis. PMID:24859593

  1. Assessing novel prognostic serum biomarkers in advanced pancreatic cancer: the role of CYFRA 21-1, serum amyloid A, haptoglobin, and 25-OH vitamin D3.

    Science.gov (United States)

    Haas, Michael; Kern, Christoph; Kruger, Stephan; Michl, Marlies; Modest, Dominik P; Giessen, Clemens; Schulz, Christoph; von Einem, Jobst C; Ormanns, Steffen; Laubender, Rüdiger P; Holdenrieder, Stefan; Heinemann, Volker; Boeck, Stefan

    2015-04-01

    The present prospective single-center study investigated the prognostic role of novel serum biomarkers in advanced pancreatic cancer (PC). Patients (pts) with locally advanced or metastatic PC treated with first-line palliative chemotherapy were included. Among others, the serum markers CYFRA 21-1, haptoglobin, serum-amyloid A (SAA), and 25-OH vitamin D3 were determined at baseline and categorized by pre-defined cut-offs [median values (MV), upper limits of normal (ULN), lower limits of normal (LLN), or the natural logarithm (ln)] and correlated with overall survival (OS). Among the 59 pts included, pre-treatment CYFRA 21-1 levels showed a strong correlation with OS independent of the applied cut-off (MV 4.9 ng/ml-14.2 vs. 4.2 months, HR 0.18, p?=?0.001; ULN 3.3 ng/ml-14.2 vs. 4.4 months, HR 0.28, p?=?0.003; [ln] CYFRA 21-1-HR 0.77, p?=?0.013). Lower values of haptoglobin were additionally associated with an improvement in OS (categorized by LLN of 2.05 g/l-10.4 vs. 5.5 months, HR 0.46, p?=?0.023; [ln] haptoglobin-HR 0.51, p?=?0.036). Pts with baseline SAA values below the MV of 22 mg/l also had a prolonged OS (10.4 vs. 5.0 months, HR 0.47, p?=?0.036). For 25-OH vitamin D3 levels, no significant correlation with OS was found. In multivariate analyses, pre-treatment CYFRA 21-1 levels (categorized by MV-HR 0.15, p?=?0.032) as well as [ln] haptoglobin (HR 0.30, p?=?0.006) retained their independent prognostic significance for OS. CYFRA 21-1, haptoglobin, and SAA might provide useful prognostic information in advanced PC. An external multicenter validation of these results is necessary. PMID:25472579

  2. Studies of the aggregation of mutant proteins in vitro provide insights into the genetics of amyloid diseases.

    Science.gov (United States)

    Chiti, Fabrizio; Calamai, Martino; Taddei, Niccolo; Stefani, Massimo; Ramponi, Giampietro; Dobson, Christopher M

    2002-12-10

    Protein aggregation and the formation of highly insoluble amyloid structures is associated with a range of debilitating human conditions, which include Alzheimer's disease, Parkinson's disease, and the Creutzfeldt-Jakob disease. Muscle acylphosphatase (AcP) has already provided significant insights into mutational changes that modulate amyloid formation. In the present paper, we have used this system to investigate the effects of mutations that modify the charge state of a protein without affecting significantly the hydrophobicity or secondary structural propensities of the polypeptide chain. A highly significant inverse correlation was found to exist between the rates of aggregation of the protein variants under denaturing conditions and their overall net charge. This result indicates that aggregation is generally favored by mutations that bring the net charge of the protein closer to neutrality. In light of this finding, we have analyzed natural mutations associated with familial forms of amyloid diseases that involve alteration of the net charge of the proteins or protein fragments associated with the diseases. Sixteen mutations have been identified for which the mechanism of action that causes the pathological condition is not yet known or fully understood. Remarkably, 14 of these 16 mutations cause the net charge of the corresponding peptide or protein that converts into amyloid deposits to be reduced. This result suggests that charge has been a key parameter in molecular evolution to ensure the avoidance of protein aggregation and identifies reduction of the net charge as an important determinant in at least some forms of protein deposition diseases. PMID:12374855

  3. Interaction of Serum Proteins with Surface of Hemodialysis Fiber Membranes

    Science.gov (United States)

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

    2012-08-01

    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.

  4. Amyloid beta binding proteins in vitro and in normal human cerebrospinal fluid.

    Science.gov (United States)

    Golabek, A; Marques, M A; Lalowski, M; Wisniewski, T

    1995-05-19

    A major neuropathological feature of Alzheimer's disease (AD) is the deposition of amyloid beta (A beta) in the form of senile plaques. The A beta peptide exists both in a beta-pleated sheet fibrillar form in amyloid deposits and as a normal soluble protein in biological fluids. Numerous proteins have been identified immunohistochemically to be associated with senile plaques, where A beta is the major constituent. Some of the latter have also been suggested to be carriers of the normal soluble A beta (sA beta) including apolipoprotein J (apoJ), apolipoprotein E (apoE) and transthyretin (TTR). We have found, using several different methods, that numerous proteins can bind synthetic A beta peptides when high concentrations are used; however, using an affinity anti-sA beta column we confirm that apoJ is the major binding protein in pooled human cerebrospinal fluid. On the other hand it is known that apoE co-purifies with A beta biochemically extracted from senile plaques. In AD tissue there may be a change in the major apolipoprotein binding A beta from apoJ to apoE. PMID:7659297

  5. A Comparative Interaction between Copper Ions with Alzheimer's ? Amyloid Peptide and Human Serum Albumin

    OpenAIRE

    G. Rezaei Behbehani; L. Barzegar; M. Mohebbian; A.A.Saboury

    2012-01-01

    The interaction of Cu2+ with the first 16 residues of the Alzheimer's amyliod ? peptide, A?(1–16), and human serum albumin (HSA) were studied in vitro by isothermal titration calorimetry at pH 7.2 and 310?K in aqueous solution. The solvation parameters recovered from the extended solvation model indicate that HSA is involved in the transport of copper ion. Complexes between A?(1–16) and copper ions have been proposed to be an aberrant interaction in the development of Alzheimer's disease, whe...

  6. Manipulations of amyloid precursor protein cleavage disrupt the circadian clock in aging Drosophila.

    Science.gov (United States)

    Blake, Matthew R; Holbrook, Scott D; Kotwica-Rolinska, Joanna; Chow, Eileen S; Kretzschmar, Doris; Giebultowicz, Jadwiga M

    2015-05-01

    Alzheimer's disease (AD) is a neurodegenerative disease characterized by severe cognitive deterioration. While causes of AD pathology are debated, a large body of evidence suggests that increased cleavage of Amyloid Precursor Protein (APP) producing the neurotoxic Amyloid-? (A?) peptide plays a fundamental role in AD pathogenesis. One of the detrimental behavioral symptoms commonly associated with AD is the fragmentation of sleep-activity cycles with increased nighttime activity and daytime naps in humans. Sleep-activity cycles, as well as physiological and cellular rhythms, which may be important for neuronal homeostasis, are generated by a molecular system known as the circadian clock. Links between AD and the circadian system are increasingly evident but not well understood. Here we examined whether genetic manipulations of APP-like (APPL) protein cleavage in Drosophila melanogaster affect rest-activity rhythms and core circadian clock function in this model organism. We show that the increased ?-cleavage of endogenous APPL by the ?-secretase (dBACE) severely disrupts circadian behavior and leads to reduced expression of clock protein PER in central clock neurons of aging flies. Our data suggest that behavioral rhythm disruption is not a product of APPL-derived A? production but rather may be caused by a mechanism common to both ? and ?-cleavage pathways. Specifically, we show that increased production of the endogenous Drosophila Amyloid Intracellular Domain (dAICD) caused disruption of circadian rest-activity rhythms, while flies overexpressing endogenous APPL maintained stronger circadian rhythms during aging. In summary, our study offers a novel entry point toward understanding the mechanism of circadian rhythm disruption in Alzheimer's disease. PMID:25766673

  7. Formaldehyde at Low Concentration Induces Protein Tau into Globular Amyloid-Like Aggregates In Vitro and In Vivo

    Science.gov (United States)

    Nie, Chun Lai; Wei, Yan; Chen, Xinyong; Liu, Yan Ying; Dui, Wen; Liu, Ying; Davies, Martyn C.; Tendler, Saul J.B.; He, Rong Giao

    2007-01-01

    Recent studies have shown that neurodegeneration is closely related to misfolding and aggregation of neuronal tau. Our previous results show that neuronal tau aggregates in formaldehyde solution and that aggregated tau induces apoptosis of SH-SY5Y and hippocampal cells. In the present study, based on atomic force microscopy (AFM) observation, we have found that formaldehyde at low concentrations induces tau polymerization whilst acetaldehyde does not. Neuronal tau misfolds and aggregates into globular-like polymers in 0.01–0.1% formaldehyde solutions. Apart from globular-like aggregation, no fibril-like polymerization was observed when the protein was incubated with formaldehyde for 15 days. SDS-PAGE results also exhibit tau polymerizing in the presence of formaldehyde. Under the same experimental conditions, polymerization of bovine serum albumin (BSA) or ?-synuclein was not markedly detected. Kinetic study shows that tau significantly misfolds and polymerizes in 60 minutes in 0.1% formaldehyde solution. However, presence of 10% methanol prevents protein tau from polymerization. This suggests that formaldehyde polymerization is involved in tau aggregation. Such aggregation process is probably linked to the tau's special “worm-like” structure, which leaves the ?-amino groups of Lys and thiol groups of Cys exposed to the exterior. Such a structure can easily bond to formaldehyde molecules in vitro and in vivo. Polymerizing of formaldehyde itself results in aggregation of protein tau. Immunocytochemistry and thioflavin S staining of both endogenous and exogenous tau in the presence of formaldehyde at low concentrations in the cell culture have shown that formaldehyde can induce tau into amyloid-like aggregates in vivo during apoptosis. The significant protein tau aggregation induced by formaldehyde and the severe toxicity of the aggregated tau to neural cells may suggest that toxicity of methanol and formaldehyde ingestion is related to tau misfolding and aggregation. PMID:17637844

  8. Appearance of Sodium Dodecyl Sulfate-Stable Amyloid ?-Protein (A?) Dimer in the Cortex During Aging

    OpenAIRE

    Enya, Miho; Morishima-kawashima, Maho; Yoshimura, Masahiro; Shinkai, Yasuhisa; Kusui, Kaoru; Khan, Karen; Games, Dora; Schenk, Dale; Sugihara, Shiro; Yamaguchi, Haruyasu; Ihara, Yasuo

    1999-01-01

    We previously noted that some aged human cortical specimens containing very low or negligible levels of amyloid ?-protein (A?) by enzyme immunoassay (EIA) provided prominent signals at 6?8 kd on the Western blot, probably representing sodium dodecyl sulfate (SDS)-stable A? dimer. Re-examination of the specificity of the EIA revealed that BAN50- and BNT77-based EIA, most commonly used for the quantitation of A?, capture SDS-dissociable A? but not SDS-stable A? dimer. Thus, all cortical...

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

    OpenAIRE

    Kudo, Wataru; Lee, Hyun-Pil; Zou, Wen-Quan; WANG, XINGLONG; Perry, George; Zhu, Xiongwei; Smith, Mark A; Petersen, Robert B.; Lee, Hyoung-Gon

    2011-01-01

    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 ...

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

    OpenAIRE

    Liu, Qiang; Zerbinatti, Celina V; ZHANG, JUAN; Hoe, Hyang-Sook; Wang, Baiping; Cole, Sarah L; Herz, Joachim; Muglia, Louis; Bu, Guojun

    2007-01-01

    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 th...

  11. The interaction of beta-amyloid protein fragment (12-28) with lipid environments.

    OpenAIRE

    Fletcher, T. G.; Keire, D. A.

    1997-01-01

    The neurotoxicity of beta-amyloid protein (beta AP) fragments may be a result of their solution conformation, which is very sensitive to solution conditions. In this work we describe NMR and CD studies of the conformation of beta AP(12-28) in lipid (micelle) environments as a function of pH and lipid type. The interaction of beta AP(12-28) with zwitterionic dodecylphosphocholine (DPC) micelles is weak and alters the conformation when compared to water solution alone. By contrast, the interact...

  12. Amyloid-?-induced Synapse Damage Is Mediated via Cross-linkage of Cellular Prion Proteins

    OpenAIRE

    Bate, Clive; Williams, Alun

    2011-01-01

    The cellular prion protein (PrPC), which is highly expressed at synapses, was identified as a receptor for the amyloid-? (A?) oligomers that are associated with dementia in Alzheimer disease. Here, we report that A? oligomers secreted by 7PA2 cells caused synapse damage in cultured neurons via a PrPC-dependent process. Exogenous PrPC added to Prnp knock-out(0/0) neurons was targeted to synapses and significantly increased A?-induced synapse damage. In contrast, the synapse damage induced by a...

  13. Functional Amyloids Signal Their Arrival

    OpenAIRE

    Badtke, Matthew P.; Hammer, Neal D; Chapman, Matthew R

    2009-01-01

    Amyloids have traditionally been associated with misfolded protein aggregates and debilitating neurodegenerative diseases. However, a growing number of functional amyloids have now been described that demonstrate that amyloid formation can be an integral part of normal cellular physiology. Functional amyloid production is highly regulated, and the resulting fibers serve a variety of roles for the cells that produce them. A new role for amyloid as storage reservoirs for peptide hormones within...

  14. A Comparative Interaction between Copper Ions with Alzheimer's ? Amyloid Peptide and Human Serum Albumin.

    Science.gov (United States)

    Behbehani, G Rezaei; Barzegar, L; Mohebbian, M; Saboury, A A

    2012-01-01

    The interaction of Cu(2+) with the first 16 residues of the Alzheimer's amyliod ? peptide, A?(1-16), and human serum albumin (HSA) were studied in vitro by isothermal titration calorimetry at pH 7.2 and 310?K in aqueous solution. The solvation parameters recovered from the extended solvation model indicate that HSA is involved in the transport of copper ion. Complexes between A?(1-16) and copper ions have been proposed to be an aberrant interaction in the development of Alzheimer's disease, where Cu(2+) is involved in A?(1-16) aggregation. The indexes of stability indicate that HSA removed Cu(2+) from A?(1-16), rapidly, decreased Cu-induced aggregation of A?(1-16), and reduced the toxicity of A?(1-16) + Cu(2+) significantly. PMID:22844264

  15. 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)

  16. Targeting Vascular Amyloid in Arterioles of Alzheimer Disease Transgenic Mice with Amyloid Beta Protein Antibody-Coated Nanoparticles

    Science.gov (United States)

    Poduslo, Joseph F.; Hultman, Kristi L.; Curran, Geoffry L.; Preboske, Gregory M.; Chamberlain, Ryan; Marja?ska, Ma?gorzata; Garwood, Michael; Jack, Clifford R.; Wengenack, Thomas M.

    2015-01-01

    The relevance of cerebral amyloid angiopathy (CAA) to the pathogenesis of Alzheimer disease (AD) and dementia in general emphasizes the importance of developing novel targeting approaches for detecting and treating cerebrovascular amyloid (CVA) deposits. We developed a nanoparticle-based technology that utilizes a monoclonal antibody against fibrillar human amyloid-?42 that is surface-coated onto a functionalized phospholipid monolayer. We demonstrate that this conjugated nanoparticle binds to CVA deposits in arterioles of AD transgenic mice (Tg2576) following infusion into the external carotid artery using 3 different approaches. The first 2 approaches utilize a blood vessel enrichment of homogenized brain and a leptomeningeal vessel preparation from thin tangential brain slices from the surface of the cerebral cortex. Targeting of CVA by the antibody-coated nanoparticle was visualized using fluorescent lissamine rhodamine-labeled phospholipids in the nanoparticles, which were compared with fluorescent staining of the endothelial cells and amyloid deposits utilizing confocal laser scanning microscopy. The third approach utilized high field strength magnetic resonance imaging of antibody-coated iron oxide nanoparticles (MIONs) following infusion into the external carotid artery. Dark foci of contrast enhancement in cortical arterioles were observed in T2*-weighted images of ex vivo AD mouse brains that correlated histologically with CVA deposits. The targeting ability of these nanoparticles to CVA provides opportunities for the prevention and treatment of CAA. PMID:21760540

  17. Influence of apolipoprotein E and its receptors on cerebral amyloid precursor protein metabolism following traumatic brain injury.

    Science.gov (United States)

    Zhou, Shuai; Sun, Xiao-Chuan

    2012-01-01

    Traumatic brain injury (TBI) is the leading cause of mortality and disability among young individuals in our society, and globally the incidence of TBI is rising sharply. Mounting evidence has indicated that apolipoprotein E (apoE: protein; APOE: gene) genotype influences the outcome after TBI. The proposed mechanism by which APOE affects the clinicopathological consequences of TBI is multifactorial and includes amyloid deposition, disruption of lipid distribution, dysfunction of mitochondrial energy production, oxidative stress and increases intracellular calcium in response to injury. This paper reviews the current state of knowledge regarding the influence of apoE and its receptors on cerebral amyloid beta-protein precursor metabolism following TBI. PMID:22663916

  18. 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)

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

    2011-10-01

    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. PMID:22102678

  19. Serum Monocyte Chemoattractant Protein-1 in Pancreatic Cancer

    OpenAIRE

    Yeo, Charles J.; Galina Chipitsyna; Harish Lavu; Jennifer Sullivan; Terry Hyslop; Qiaoke Gong; Arafat, Hwyda A.

    2011-01-01

    Background/Aims. Pancreatic ductal adenocarcinoma (PDA) has etiological association with chronic inflammation. Elevated circulating levels of inflammatory mediators, such as monocyte chemoattractant protein-1 (MCP-1), are found in obese individuals. We hypothesized that serum MCP-1 levels are elevated in obese PDA patients. Methods. ELISA was used to analyze MCP-1 serum levels in PDA (n = 62) and intraductal papillary mucinous neoplasms (IPMN) (n = 27). Recursive partitioning statistical anal...

  20. Serum Protein Electrophoresis in Dogs With Intestinal Parasites

    OpenAIRE

    KAYMAZ, Alev AKDO?AN; BAKIREL, Utku; GÖNÜL, Remzi; TAN, Hüseyin

    1999-01-01

    The serum of 66 dogs with intestinal parasites (showing gastrointestinal problems caused by taeniosis, coccidiosis, ancylostomosis, trichuriosis and ascarididosis) was examined by electrophoresis. There were 6 dogs with coccidiosis, 6 dogs with ancylostomosis, 6 dogs with trichuriosis, 24 dogs with taeniosis and 24 dogs with ascarididosis. After agar gel protein electorphoresis of the serum samples, ?1 globulin levels were significantly lower in the coccidiosis group than in the other grou...

  1. Serum Copper and Plasma Protein Status in Normal Pregnancy

    Directory of Open Access Journals (Sweden)

    Nushrat Noor, Nasim Jahan, Nayma Sultana

    2012-12-01

    Full Text Available AbstractBackground: Gradual alteration of serum copper and some plasma protein levels may occur with advancement of pregnancy, which is associated with increased maternal and infant morbidity and mortality.Objective: To observe serum copper and plasma protein levels in normal pregnant women of different trimesters in order to find out their nutritional status.Methods: This cross sectional study was carried out in the Department of Physiology, Sir Salimullah Medical College (SSMC, Dhaka, between 1st January 2010 and December 2010. Ninety normal pregnant women of different trimesters with age 20-30 years were included in the study group. They were selected from Out Patient Department of Obstetrics and Gynaecology, SSMC. Age matched 30 non-pregnant women were taken as control. Serum copper level was measured by Spectrophotometric method, serum total protein and albumin levels were estimated by standard method. Statistical analysis was done by one way ANOVA, Bonferroni and Pearson’s correlation coefficient test as applicable.Results: Serum Cu levels were significantly higher in all trimesters of pregnant women compared to control. Again, this value was significantly higher in 3rd trimester than that of in 1st and 2nd trimester and also in 2nd trimester than that of in 1st trimester. In addition, mean serum total protein level was significantly lower in 3rd trimester than control but no statistically significant difference was observed among different trimesters. Again, mean serum albumin level was significantly lower in 2nd and 3rd trimester than 1st trimester and control. In addition, serum Cu concentration showed significant positive correlation with different trimesters of gestation.Conclusion: This study reveals that hypercupremia along with hypoproteinemia occur in pregnant women from 1st to 3rd trimester of gestation. This gradual alteration of micro and macronutrients become more profound with advancement of pregnancy.

  2. Localization and Trafficking of Amyloid-? Protein Precursor and Secretases: Impact on Alzheimer's Disease.

    Science.gov (United States)

    Agostinho, Paula; Pliássova, Anna; Oliveira, Catarina R; Cunha, Rodrigo A

    2015-01-01

    Alzheimer's disease (AD) affects almost 35 million people worldwide. One of the neuropathological features of AD is the presence of extracellular amyloid plaques, which are mainly composed of amyloid-? (A?) peptides. These peptides derive from the amyloidogenic proteolytic processing of the amyloid-? protein precursor (A?PP), through the sequential action of ?- and ?-secretases. However, A?PP can also be cleaved by a non-amyloidogenic pathway, involving an ?-secretase, and in this case the A? formation is precluded. The production of A? and of other A?PP catabolites depends on the spatial and temporal co-localization of A?PP with ?- or ?-secretases and ?-secretase, which traffic through the secretory pathway in a highly regulated manner. Disturbances on A?PP and secretases intracellular trafficking and, consequently, in their localization may affect dynamic interactions between these proteins with consequences in the AD pathogenesis. In this article, we critically review the recent knowledge about the trafficking and co-localization of A?PP and related secretases in the brain under physiological and AD conditions. A particular focus is given to data concerning the distribution of A?PP and secretases in different types of synapses relatively to other neuronal or glial localizations. Furthermore, we discuss some possible signals that govern the dynamic encounter of A?PP with each group of secretases, such as A?PP mutations, estrogen deprivation, chronic stress, metabolic impairment, and alterations in sleep pattern-associated with aging. The knowledge of key signals that are responsible for the shifting of A?PP processing away from ?-secretases and toward the ?-secretases might be useful to develop AD therapeutic strategies. PMID:25589722

  3. Ceruloplasmin and ?-amyloid precursor protein confer neuroprotection in traumatic brain injury and lower neuronal iron.

    Science.gov (United States)

    Ayton, Scott; Zhang, Moses; Roberts, Blaine R; Lam, Linh Q; Lind, Monica; McLean, Catriona; Bush, Ashley I; Frugier, Tony; Crack, Peter J; Duce, James A

    2014-04-01

    Traumatic brain injury (TBI) is in part complicated by pro-oxidant iron elevation independent of brain hemorrhage. Ceruloplasmin (CP) and ?-amyloid protein precursor (APP) are known neuroprotective proteins that reduce oxidative damage through iron regulation. We surveyed iron, CP, and APP in brain tissue from control and TBI-affected patients who were stratified according to time of death following injury. We observed CP and APP induction after TBI accompanying iron accumulation. Elevated APP and CP expression was also observed in a mouse model of focal cortical contusion injury concomitant with iron elevation. To determine if changes in APP or CP were neuroprotective we employed the same TBI model on APP(-/-) and CP(-/-) mice and found that both exhibited exaggerated infarct volume and iron accumulation postinjury. Evidence supports a regulatory role of both proteins in defence against iron-induced oxidative damage after TBI, which presents as a tractable therapeutic target. PMID:24509156

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

    Directory of Open Access Journals (Sweden)

    Anthony J Turner

    2014-09-01

    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.

  5. Analysis of glycans on serum proteins using antibody microarrays

    OpenAIRE

    CHEN, SONGMING; Haab, Brian B

    2009-01-01

    Antibody arrays can be employed for the profiling glycan structures on proteins. Antibody arrays capture multiple, specific proteins directly from biological samples (such as serum), and lectin and glycan-binding antibodies probe the levels of specific glycans on the captured proteins. We use a practical method of partitioning microscope slides to enable the convenient processing of many detection reagents or samples. A critical first step in the procedure is the chemical derivatization of th...

  6. Amyloid plaques, neurofibrillary tangles and neuronal loss in brains of transgenic mice overexpressing a C-terminal fragment of human amyloid precursor protein.

    Science.gov (United States)

    Kawabata, S; Higgins, G A; Gordon, J W

    1991-12-12

    Alzheimer's disease (AD) affects more than 30% of people over 80 years of age. The aetiology and pathogenesis of this progressive dementia is poorly understood, but symptomatic disease is associated histopathologically with amyloid plaques, neurofibrillary tangles and neuronal loss primarily in the temporal lobe and neocortex of the brain. The core of the extracellular plaque is a derivative of the amyloid precursor protein (APP), referred to as beta/A4, and contains the amino-acid residues 29-42 that are normally embedded in the membrane-spanning region of the precursor. The cellular source of APP and the relationship of its deposition to the neuropathology of AD is unknown. To investigate the relationship between APP overexpression and amyloidogenesis, we have developed a vector to drive expression specifically in neurons of a C-terminal fragment of APP that contains the beta/A4 region, and have used a transgenic mouse system to insert and express this construct. We report here that overexpression of this APP transgene in neurons is sufficient to produce extracellular dense-core amyloid plaques, neurofibrillary tangles and neuronal degeneration similar to that in the AD brain. PMID:1793460

  7. 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)

  8. Modulation of ?-amyloid precursor protein trafficking and processing by the low density lipoprotein receptor family

    Directory of Open Access Journals (Sweden)

    Cam Judy A

    2006-08-01

    Full Text Available Abstract Amyloid-? peptide (A? accumulation in the brain is an early, toxic event in the pathogenesis of Alzheimer's disease (AD. A? is produced by proteolytic processing of a transmembrane protein, ?-amyloid precursor protein (APP, by ?- and ?-secretases. Mounting evidence has demonstrated that alterations in APP cellular trafficking and localization directly impact its processing to A?. Recent studies have shown that members of the low-density lipoprotein receptor family, including LRP, LRP1B, SorLA/LR11, and apolipoprotein E (apoE receptor 2, interact with APP and regulate its endocytic trafficking. Another common feature of these receptors is their ability to bind apoE, which exists in three isoforms in humans and the presence of the ?4 allele represents a genetic risk factor for AD. In this review, we summarize the current understanding of the function of these apoE receptors with a focus on their role in APP trafficking and processing. Knowledge of the interactions between these distinct low-density lipoprotein receptor family members and APP may ultimately influence future therapies for AD.

  9. Structure of Alzheimer’s disease amyloid precursor protein copper-binding domain at atomic resolution

    International Nuclear Information System (INIS)

    An atomic resolution structure of the copper-binding domain of the Alzheimer’s disease amyloid precursor protein is presented. Amyloid precursor protein (APP) plays a central role in the pathogenesis of Alzheimer’s disease, as its cleavage generates the A? peptide that is toxic to cells. APP is able to bind Cu2+ and reduce it to Cu+ through its copper-binding domain (CuBD). The interaction between Cu2+ and APP leads to a decrease in A? production and to alleviation of the symptoms of the disease in mouse models. Structural studies of CuBD have been undertaken in order to better understand the mechanism behind the process. Here, the crystal structure of CuBD in the metal-free form determined to ultrahigh resolution (0.85 Å) is reported. The structure shows that the copper-binding residues of CuBD are rather rigid but that Met170, which is thought to be the electron source for Cu2+ reduction, adopts two different side-chain conformations. These observations shed light on the copper-binding and redox mechanisms of CuBD. The structure of CuBD at atomic resolution provides an accurate framework for structure-based design of molecules that will deplete A? production

  10. Mechanism of amyloid ?-protein dimerization determined using single-molecule AFM force spectroscopy

    Science.gov (United States)

    Lv, Zhengjian; Roychaudhuri, Robin; Condron, Margaret M.; Teplow, David B.; Lyubchenko, Yuri L.

    2013-10-01

    A?42 and A?40 are the two primary alloforms of human amyloid ?-protein (A?). The two additional C-terminal residues of A?42 result in elevated neurotoxicity compared with A?40, but the molecular mechanism underlying this effect remains unclear. Here, we used single-molecule force microscopy to characterize interpeptide interactions for A?42 and A?40 and corresponding mutants. We discovered a dramatic difference in the interaction patterns of A?42 and A?40 monomers within dimers. Although the sequence difference between the two peptides is at the C-termini, the N-terminal segment plays a key role in the peptide interaction in the dimers. This is an unexpected finding as N-terminal was considered as disordered segment with no effect on the A? peptide aggregation. These novel properties of A? proteins suggests that the stabilization of N-terminal interactions is a switch in redirecting of amyloids form the neurotoxic aggregation pathway, opening a novel avenue for the disease preventions and treatments.

  11. Genetic and environmental influences of surfactant protein D serum levels

    DEFF Research Database (Denmark)

    Sorensen, G.L.; Hjelmborg, J.V.

    2006-01-01

    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).

  12. Recombinant immunoglobulin variable domains generated from synthetic genes provide a system for in vitro characterization of light-chain amyloid proteins.

    OpenAIRE

    Stevens, P. W.; Raffen, R.; Hanson, D. K.; Deng, Y L; Berrios-Hammond, M.; Westholm, F A; Murphy, C.; Eulitz, M.; Wetzel, R.; Solomon, A.

    1995-01-01

    The primary structural features that render human monoclonal light chains amyloidogenic are presently unknown. To gain further insight into the physical and biochemical factors that result in the pathologic deposition of these proteins as amyloid fibrils, we have selected for detailed study three closely homologous protein products of the light-chain variable-region single-gene family VkIV. Two of these proteins, REC and SMA, formed amyloid fibrils in vivo. The third protein, LEN, was excrete...

  13. Unraveling the mysteries of serum albumin—more than just a serum protein

    OpenAIRE

    Merlot, Angelica M.; Kalinowski, Danuta S.; Richardson, Des R.

    2014-01-01

    Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are re...

  14. Unraveling the mysteries of serum albumin – more than just a serum protein.

    OpenAIRE

    AngelicaM.Merlot

    2014-01-01

    Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are re...

  15. Doped diamond-like carbon coatings for surgical instruments reduce protein and prion-amyloid biofouling and improve subsequent cleaning.

    Science.gov (United States)

    Secker, T J; Hervé, R; Zhao, Q; Borisenko, K B; Abel, E W; Keevil, C W

    2012-01-01

    Doped diamond-like carbon (DLC) coatings offer potential antifouling surfaces against microbial and protein attachment. In particular, stainless steel surgical instruments are subject to tissue protein and resilient prion protein attachment, making decontamination methods used in sterile service departments ineffective, potentially increasing the risk of iatrogenic Creutzfeldt-Jakob disease during surgical procedures. This study examined the adsorption of proteins and prion-associated amyloid to doped DLC surfaces and the efficacy of commercial cleaning chemistries applied to these spiked surfaces, compared to titanium nitride coating and stainless steel. Surfaces inoculated with ME7-infected brain homogenate were visualised using SYPRO Ruby/Thioflavin T staining and modified epi-fluorescence microscopy before and after cleaning. Reduced protein and prion amyloid contamination was observed on the modified surfaces and subsequent decontamination efficacy improved. This highlights the potential for a new generation of coatings for surgical instruments to reduce the risk of iatrogenic CJD infection. PMID:22694725

  16. Long-term kinetics of AA amyloidosis and effects of inflammatory restimulation after disappearance of amyloid depositions in mice.

    Science.gov (United States)

    Muhammad, N; Murakami, T; Inoshima, Y; Ishiguro, N

    2015-07-01

    Amyloid A (AA) amyloidosis is characterized by extracellular pathogenic deposition of insoluble fibril protein in various body organs. Deposited amyloid generally remains in a variety of organs for long periods, but its disappearance has been reported after the precursor protein is diminished. The kinetics of AA deposition are not completely understood and, in particular, the roles of cells and cytokines in the deposition and clearance of amyloid remain unclear. In this study, we investigated the disappearance of amyloid depositions in mice over a 1-year period. AA amyloidosis was induced experimentally in mice by injecting amyloid-enhancing factor (AEF) and silver nitrate. Mice were killed at different time-points to examine the occurrence and disappearance of amyloid depositions. Maximum levels of amyloid depositions were observed at 20 days after inoculation. Clearance of amyloid depositions was observed from the 40th day onwards, with only minute traces of amyloid present by 240 days. A second inflammatory stimulus consisting of AEF and silver nitrate was given at 330 or 430 days, after amyloid depositions had disappeared almost completely. After that, serum amyloid A was overproduced and redeposition of amyloid was observed, indicating that all mice were primed for aggressive amyloid depositions. After administration of the inflammatory stimuli, the proinflammatory environment was found to have increased levels of interleukin (IL)-6, while anti-inflammatory conditions were established by IL-10 as regression of amyloid deposition occurred. These results suggest that the proinflammatory and anti-inflammatory status have key roles in both amyloid deposition and clearance. PMID:25736960

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

    Directory of Open Access Journals (Sweden)

    PRL Silva

    2007-12-01

    Full Text Available 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-transferase (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.

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

    Scientific Electronic Library Online (English)

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

    2007-12-01

    Full Text Available 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.

  19. Reducing Human Apolipoprotein E Levels Attenuates Age-Dependent A? Accumulation in Mutant Human Amyloid Precursor Protein Transgenic Mice

    OpenAIRE

    Bien-Ly, Nga; Gillespie, Anna K.; Walker, David; Yoon, Seo Yeon; Huang, Yadong

    2012-01-01

    Apolipoprotein (apo) E4 plays a major role in the pathogenesis of Alzheimer’s disease (AD). Brain amyloid-beta (A?) accumulation depends on age and apoE isoforms (apoE4 > apoE3) both in humans and in transgenic mouse models. Brain apoE levels are also isoform-dependent, but in the opposite direction (apoE4 < apoE3). Thus, one prevailing hypothesis is to increase brain apoE expression to reduce A? levels. To test this hypothesis, we generated mutant human amyloid precursor protein (hAPP) trans...

  20. Dual roles of the transmembrane protein p23/TMP21 in the modulation of amyloid precursor protein metabolism

    Directory of Open Access Journals (Sweden)

    Wieland Felix T

    2007-02-01

    Full Text Available Abstract Background Alzheimer's disease (AD is characterized by cerebral deposition of ?-amyloid (A? peptides. A? is released from ectodomain cleaved amyloid precursor protein (APP via intramembranous proteolysis by ?-secretase, a complex consisting of presenilin and a few other proteins. p23/TMP21, a member of the p24 family type I transmembrane proteins, was recently identified as a presenilin complex component capable of modulating ?-secretase cleavage. The p24 family proteins form oligomeric complexes and regulate vesicular trafficking in the early secretory pathway, but their role in APP trafficking has not been investigated. Results Here, we report that siRNA-mediated depletion of p23 in N2a neuroblastoma and HeLa cells produces concomitant knockdown of additional p24 family proteins and increases secretion of sAPP. Furthermore, intact cell and cell-free A? production increases following p23 knockdown, similar to data reported earlier using HEK293 cells. However, we find that p23 is not present in mature ?-secretase complexes isolated using an active-site ?-secretase inhibitor. Depletion of p23 and expression of a familial AD-linked PS1 mutant have additive effects on A?42 production. Knockdown of p23 expression confers biosynthetic stability to nascent APP, allowing its efficient maturation and surface accumulation. Moreover, immunoisolation analyses show decrease in co-residence of APP and the APP adaptor Mint3. Thus, multiple lines of evidence indicate that p23 function influences APP trafficking and sAPP release independent of its reported role in ?-secretase modulation. Conclusion These data assign significance to p24 family proteins in regulating APP trafficking in the continuum of bidirectional transport between the ER and Golgi, and ascribe new relevance to the regulation of early trafficking in AD pathogenesis.

  1. Endogenous C-terminal fragments of beta-amyloid precursor protein from Xenopus laevis skin exudate.

    Science.gov (United States)

    Clamagirand, Christine; El Abida, Boutaïna; Der Garabedian, P Arsene; Hanquez, Chantal; Dubost, Lionel; Marie, Arul; Rholam, Mohamed; Friguet, Bertrand; Cohen, Paul

    2007-04-01

    Using a monoclonal antibody against the entire C-terminal end of human APP(695) (643-695 sequence) and a monoclonal antibody directed against human beta[1-40] amyloid peptide (betaA), we show the existence of endogenous peptides proteolytically derived from APP in skin exudate of the non transgenic Xenopus laevis frog. The majority of the immunoreactivity is found associated with a 30 kDa molecular species. Biochemical fractionation followed by mass spectrometry identification allowed us to assign this molecular species to C-terminal APP fragments containing all or part of betaA. According to the nature of N- and C-terminal amino acids we identified endogenous beta-, gamma-, epsilon-secretase-like activities, caspase-like activity and numerous endogenous cleavage sites within the beta-amyloid sequence at same sites as those observed in human betaA sequence. All these homologies with human indicate that X. laevis skin exudate is a good natural model to study betaA metabolism. In this way, interestingly, we identified endogenous cleavages at prohormone convertase-like sites not yet described at the same sites in human. Finally, all identified peptide fragments were stably associated with a 20.2 kDa protein. These new observed features suggest new research pathways concerning human betaA metabolism and carriage of hydrophobic peptide fragments issued from APP processing. PMID:17270477

  2. Sortilin and SorLA Display Distinct Roles in Processing and Trafficking of Amyloid Precursor Protein

    DEFF Research Database (Denmark)

    Gustafsen, Camilla; Glerup, Simon

    2013-01-01

    The development and progression of Alzheimer's disease is linked to excessive production of toxic amyloid-? peptide, initiated by ?-secretase cleavage of the amyloid precursor protein (APP). In contrast, soluble APP? (sAPP?) generated by the ?-secretase is known to stimulate dendritic branching and enhance synaptic function. Regulation of APP processing, and the shift from neurotrophic to neurotoxic APP metabolism remains poorly understood, but the cellular localization of APP and its interaction with various receptors is considered important. We here identify sortilin as a novel APP interaction partner. Like the related APP receptor SorLA, sortilin is highly expressed in the CNS, but whereas SorLA mainly colocalizes with APP in the soma, sortilin interacts with APP in neurites. The presence of sortilin promotes ?-secretase cleavage of APP, unlike SorLA, which inhibits the generation of all soluble products. Also, sortilin and SorLA both bind and mediate internalization of sAPP but to different cellular compartments. The interaction involves the 6A domain of APP, present in both neuronal and non-neuronal APP isoforms. This is important as sAPP receptors described so far only bind the non-neuronal isoforms, leaving SorLA and sortilin as the only receptors for sAPP generated by neurons. Together, our findings establish sortilin, as a novel APP interaction partner that influences both production and cellular uptake of sAPP.

  3. Two amyloid precursor protein transgenic mouse models with Alzheimer?disease-like?pathology

    Science.gov (United States)

    Sturchler-Pierrat, Christine; Abramowski, Dorothee; Duke, Mairead; Wiederhold, Karl-Heinz; Mistl, Claudia; Rothacher, Sabin; Ledermann, Birgit; Bürki, Kurt; Frey, Peter; Paganetti, Paolo A.; Waridel, Caroline; Calhoun, Michael E.; Jucker, Mathias; Probst, Alphonse; Staufenbiel, Matthias; Sommer, Bernd

    1997-01-01

    Mutations in the amyloid precursor protein (APP) gene cause early-onset familial Alzheimer disease (AD) by affecting the formation of the amyloid ? (A?) peptide, the major constituent of AD plaques. We expressed human APP751 containing these mutations in the brains of transgenic mice. Two transgenic mouse lines develop pathological features reminiscent of AD. The degree of pathology depends on expression levels and specific mutations. A 2-fold overexpression of human APP with the Swedish double mutation at positions 670/671 combined with the V717I mutation causes A? deposition in neocortex and hippocampus of 18-month-old transgenic mice. The deposits are mostly of the diffuse type; however, some congophilic plaques can be detected. In mice with 7-fold overexpression of human APP harboring the Swedish mutation alone, typical plaques appear at 6 months, which increase with age and are Congo Red-positive at first detection. These congophilic plaques are accompanied by neuritic changes and dystrophic cholinergic fibers. Furthermore, inflammatory processes indicated by a massive glial reaction are apparent. Most notably, plaques are immunoreactive for hyperphosphorylated tau, reminiscent of early tau pathology. The immunoreactivity is exclusively found in congophilic senile plaques of both lines. In the higher expressing line, elevated tau phosphorylation can be demonstrated biochemically in 6-month-old animals and increases with age. These mice resemble major features of AD pathology and suggest a central role of A? in the pathogenesis of the disease. PMID:9371838

  4. Amyloid-beta protein clearance and degradation (ABCD) pathways and their role in Alzheimer's disease.

    Science.gov (United States)

    Baranello, Robert J; Bharani, Krishna L; Padmaraju, Vasudevaraju; Chopra, Nipun; Lahiri, Debomoy K; Greig, Nigel H; Pappolla, Miguel A; Sambamurti, Kumar

    2015-01-01

    Amyloid-? proteins (A?) of 42 (A?42) and 40 aa (A?40) accumulate as senile plaques (SP) and cerebrovascular amyloid protein deposits that are defining diagnostic features of Alzheimer's disease (AD). A number of rare mutations linked to familial AD (FAD) on the A? precursor protein (APP), Presenilin-1 (PS1), Presenilin- 2 (PS2), Adamalysin10, and other genetic risk factors for sporadic AD such as the ?4 allele of Apolipoprotein E (ApoE-?4) foster the accumulation of A? and also induce the entire spectrum of pathology associated with the disease. A? accumulation is therefore a key pathological event and a prime target for the prevention and treatment of AD. APP is sequentially processed by ?-site APP cleaving enzyme (BACE1) and ?-secretase, a multisubunit PS1/PS2-containing integral membrane protease, to generate A?. Although A? accumulates in all forms of AD, the only pathways known to be affected in FAD increase A? production by APP gene duplication or via base substitutions on APP and ?-secretase subunits PS1 and PS2 that either specifically increase the yield of the longer A?42 or both A?40 and A?42. However, the vast majority of AD patients accumulate A? without these known mutations. This led to proposals that impairment of A? degradation or clearance may play a key role in AD pathogenesis. Several candidate enzymes, including Insulin-degrading enzyme (IDE), Neprilysin (NEP), Endothelin-converting enzyme (ECE), Angiotensin converting enzyme (ACE), Plasmin, and Matrix metalloproteinases (MMPs) have been identified and some have even been successfully evaluated in animal models. Several studies also have demonstrated the capacity of ?-secretase inhibitors to paradoxically increase the yield of A? and we have recently established that the mechanism is by skirting A? degradation. This review outlines major cellular pathways of A? degradation to provide a basis for future efforts to fully characterize the panel of pathways responsible for A? turnover. PMID:25523424

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

    Directory of Open Access Journals (Sweden)

    Kuroda Takeshi

    2012-09-01

    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.

  6. Amyloid precursor protein enhances nav1.6 sodium channel cell surface expression.

    Science.gov (United States)

    Liu, Chao; Tan, Francis Chee Kuan; Xiao, Zhi-Cheng; Dawe, Gavin S

    2015-05-01

    Amyloid precursor protein (APP) is commonly associated with Alzheimer disease, but its physiological function remains unknown. Nav1.6 is a key determinant of neuronal excitability in vivo. Because mouse models of gain of function and loss of function of APP and Nav1.6 share some similar phenotypes, we hypothesized that APP might be a candidate molecule for sodium channel modulation. Here we report that APP colocalized and interacted with Nav1.6 in mouse cortical neurons. Knocking down APP decreased Nav1.6 sodium channel currents and cell surface expression. APP-induced increases in Nav1.6 cell surface expression were Go protein-dependent, enhanced by a constitutively active Go protein mutant, and blocked by a dominant negative Go protein mutant. APP also regulated JNK activity in a Go protein-dependent manner. JNK inhibition attenuated increases in cell surface expression of Nav1.6 sodium channels induced by overexpression of APP. JNK, in turn, phosphorylated APP. Nav1.6 sodium channel surface expression was increased by T668E and decreased by T668A, mutations of APP695 mimicking and preventing Thr-668 phosphorylation, respectively. Phosphorylation of APP695 at Thr-668 enhanced its interaction with Nav1.6. Therefore, we show that APP enhances Nav1.6 sodium channel cell surface expression through a Go-coupled JNK pathway. PMID:25767117

  7. Immune hyporesponsiveness to amyloid beta-peptide in amyloid precursor protein transgenic mice: implications for the pathogenesis and treatment of Alzheimer's disease.

    Science.gov (United States)

    Monsonego, A; Maron, R; Zota, V; Selkoe, D J; Weiner, H L

    2001-08-28

    Alzheimer's disease is a dementia that involves progressive deposition of amyloid beta-protein (Abeta) in brain regions important for memory and cognition, followed by secondary inflammation that contributes to the neuropathologic process. Immunization with Abeta can reduce cerebral Abeta burden and consequent neuropathologic changes in the brains of mice transgenic for the beta-amyloid precursor protein (APP). We found that transgenic expression of human APP in B6SJL mice, under the prion promoter, results in immune hyporesponsiveness to human Abeta, in terms of both antibody and cellular immune responses. The decreased antibody responses were related not to B cell tolerance but rather to the inability of Abeta-specific T cells to provide help for antibody production. The immune hyporesponsiveness could be overcome if T cell help was provided by coupling an Abeta B cell epitope to BSA. Our results suggest that expression of APP in transgenic mice is associated with an Abeta-specific impaired adaptive immune response that may contribute to the neuropathology. Moreover, humans with life-long elevation of brain and peripheral Abeta (e.g., patients with presenilin mutations or Down syndrome) could have reduced immune responses to Abeta vaccination. PMID:11517335

  8. Retromer binds the FANSHY sorting motif in SorLA to regulate amyloid precursor protein sorting and processing

    OpenAIRE

    Fjorback, A.W.; Seaman, M; Gustafsen, C.; Mehmedbasic, A.; Gokool, S.; Wu, C.; Militz, D.; Schmidt, V; Madsen, P.; Nyengaard, J. R.; Willnow, T E; Christensen, E I; Mobley, W.B.; Nykjaer, A; Andersen, O.M.

    2012-01-01

    sorLA is a sorting receptor for amyloid precursor protein (APP) genetically linked to Alzheimer's disease (AD). Retromer, an adaptor complex in the endosome-to-Golgi retrieval pathway, has been implicated in APP transport because retromer deficiency leads to aberrant APP sorting and processing and levels of retromer proteins are altered in AD. Here we report that sorLA and retromer functionally interact in neurons to control trafficking and amyloidogenic processing of APP. We have identified ...

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

    OpenAIRE

    Sandhu, F A; Kim, Y.; Lapan, K A; Salim, M; Aliuddin, V; Zain, S. B.(State University of New York, 12222, Albany, New York, USA)

    1996-01-01

    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. ...

  10. The Functions of the Amyloid Precursor Protein Gene and Its Derivative Peptides: II Experimental Evidence and Clinical Studies

    OpenAIRE

    Panegyres, Peter K; Atkins, Emily R.

    2011-01-01

    In vitro studies suggest Amyloid Precursor Protein Gene (APP) is involved in interaction with the extracellular matrix, neurite growth, adhesion, development, synaptic function, platelet function, and interaction with GTP binding proteins. In vivo experiments show a role in embryonic development, response to cerebral excitotoxicity and gliosis, response to brain injury including ischaemia, hypothalamic function, locomotor function, learning and memory. In vitro observations indicate A? has a...

  11. sAPP? rescues deficits in amyloid precursor protein knockout mice following focal traumatic brain injury.

    Science.gov (United States)

    Corrigan, Frances; Vink, Robert; Blumbergs, Peter C; Masters, Colin L; Cappai, Roberto; van den Heuvel, Corinna

    2012-07-01

    The amyloid precursor protein (APP) is thought to be neuroprotective following traumatic brain injury (TBI), although definitive evidence at moderate to severe levels of injury is lacking. In the current study, we investigated histological and functional outcomes in APP-/- mice compared with APP+/+ mice following a moderate focal injury, and whether administration of sAPP? restored the outcomes in knockout animals back to the wildtype state. Following moderate controlled cortical impact injury, APP-/- mice demonstrated greater impairment in motor and cognitive outcome as determined by the ledged beam and Barnes Maze tests respectively (p TBI, and that this neuroprotective activity is related to the presence of sAPP?. Importantly, it indicates that the mechanism of action of exogenously added sAPP? is independent of the presence of endogenous APP. PMID:22519988

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

    DEFF Research Database (Denmark)

    Zheng, Lin; Kågedal, Katarina

    2009-01-01

    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

  13. Functional Amyloids Signal Their Arrival

    Science.gov (United States)

    Matthew P. Badtke (Ann Arbor; University of Michigan REV)

    2009-07-21

    Amyloids have traditionally been associated with misfolded protein aggregates and debilitating neurodegenerative diseases. However, a growing number of functional amyloids have now been described that demonstrate that amyloid formation can be an integral part of normal cellular physiology. Functional amyloid production is highly regulated, and the resulting fibers serve a variety of roles for the cells that produce them. A new role for amyloid as storage reservoirs for peptide hormones within mammalian secretory granules has been discovered. More than 30 different peptide hormones have been found to form amyloids in vitro, and both rats and mice have been shown to store hormone amyloid deposits in secretory granules. Thus, the emerging evidence adds to the diverse roles of amyloid and raises intriguing questions for both the peptide hormone and the functional amyloid fields.

  14. SorLA CR-Domains Protect the Amyloid Precursor Protein against Processing

    DEFF Research Database (Denmark)

    Mehmedbasic, Arnela; Christensen, Sofie K

    2014-01-01

    SorLA is a neuronal sorting receptor that is genetically associated with Alzheimer's disease. SorLA interacts directly with the amyloid precursor protein (APP) and affects the processing of the precursor, leading to a decreased generation of the amyloid-? (A?) peptide. The sorLA complement-type repeat (CR)-domains associate in vitro with APP, but the precise molecular determinants of sorLA-APP complex formation and the mechanisms responsible for the effect of binding on APP processing have not yet been elucidated. Here, we have generated protein expression constructs for sorLA devoid of the 11 CR-domains and for two sorLA mutants harboring substitutions of the fingerprint residues in the central CR-domains. We generated SH-SY5Y cell lines that stably express these sorLA variants to study the binding and processing of APP using co-immunoprecipitation and western blotting/ELISA assays, respectively. We found that the sorLA CR-cluster is essential for interaction with APP and that deletion of the CR-cluster abolishes the protection against APP processing. Mutation of identified fingerprint residues in the sorLA CR-domains leads to changes in the O-linked glycosylation of APP when expressed in SH-SY5Y cells. Our results provide novel information on the mechanisms behind the influence of sorLA activity on APP metabolism by controlling post-translational glycosylation in the Golgi, suggesting new strategies against amyloidogenesis in Alzheimer's disease.

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

    DEFF Research Database (Denmark)

    Nielsen, Morten S; Gustafsen, Camilla

    2007-01-01

    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

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

    Science.gov (United States)

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

    2006-01-01

    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

  17. Molecular inscription of environmental information into protein suprastructures: temperature effects on unit assembly of ?-synuclein oligomers into polymorphic amyloid fibrils.

    Science.gov (United States)

    Bhak, Ghibom; Lee, Junghee; Kim, Tae-Hwan; Lee, Soonkoo; Lee, Daekyun; Paik, Seung R

    2014-12-01

    Molecular-level storage of environmental information in biological structures in tangible forms, and their subsequent transfer to the next generation, has been studied using the phenomenon of amyloidogenesis, which defines a biochemical condition generating highly ordered protein aggregates known as amyloid fibrils. ?-Synuclein oligomers shown to experience unit assembly as the formation of amyloid fibrils were used in the present study as an environment-sensing agent. With temperature varying in 2 °C intervals between 37 °C and 43 °C, the oligomeric unit assembly led to fibrillar polymorphism from a straight to a curly appearance, as assessed using TEM and small-angle neutron scattering; the different effects on the secondary structures were evaluated using attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy. The resulting diversified amyloid fibrils, which have distinctive molecular characteristics, were shown to be inherited by the next generation through the self-propagating property of amyloidogenesis. Storage of intangible temperature information in the diversified protein suprastructures and perpetuation of the stored information in the form of polymorphic amyloid fibrils could represent molecular inscription of environmental information into biological systems; this could further extend our understanding of any physiological/pathological significance of amyloidogenic polymorphism and be utilized in the area of nanobiotechnology to process various external signals. PMID:25203358

  18. Overexpression of amyloid precursor protein A4 (beta-amyloid) immunoreactivity in genetically transformed cells: implications for a cellular model of Alzheimer amyloidosis.

    OpenAIRE

    Marotta, C A; Chou, W G; Majocha, R E; Watkins, R.; LaBonne, C; Zain, S. B.(State University of New York, 12222, Albany, New York, USA)

    1989-01-01

    Among the major obstacles to clarifying molecular mechanisms involved in amyloid metabolism in Alzheimer disease has been the unavailability of laboratory models for this uniquely human disorder. The present studies were aimed at establishing genetically engineered cell lines that overexpress amyloid immunoreactivity and that may be relevant to amyloid accumulation in the Alzheimer disease brain. We used cloned amyloid cDNA that contains a region encoding A4 (beta-polypeptide) amino acids alo...

  19. Serum protein concentrations in calves with experimentally induced pneumonic pasteurellosis

    Directory of Open Access Journals (Sweden)

    Fagliari J.J.

    2003-01-01

    Full Text Available Ten healthy 2 to 4-week-old Holstein calves were randomly allotted into control and infected groups. Control calves (n=5 were inoculated intrabronchially with 5ml of Dulbecco's phosphate-buffered saline solution (DPBSS. Infected calves (n=5 were inoculated intrabronchially with 5x10(9 log-phase Mannheimia haemolytica organisms suspended in 5ml of DPBSS. Blood samples were obtained 15 minutes before and one, two, four and six hours after inoculation. Serum protein concentrations were determined by means of sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Serum concentrations of proteins with molecular weights of 125,000 D (ceruloplasmin, 60,000 D (a 1-antitrypsin, 45,000 D (haptoglobin, and 40,000 D (acid glycoprotein were significantly increased in calves with pneumonic pasteurellosis, compared with concentrations in control calves. Results indicate that acute phase proteins increase more rapidly after the onset of inflammation than previously thought. Measurement of serum protein concentrations may be useful in monitoring the progression of the induced pneumonic pasteurellosis in calves.

  20. 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.

  1. 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)

    Connolly, Mary

    2010-06-01

    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.

  2. Lead-Induced Accumulation of ?-Amyloid in the Choroid Plexus: Role of Low Density Lipoprotein Receptor Protein-1 and Protein Kinase C

    OpenAIRE

    Behl, Mamta; Zhang, Yanshu; Shi, Yunzhou; Cheng, Jixin; Du, Yansheng; Zheng, Wei

    2010-01-01

    The choroid plexus (CP), constituting the blood–cerebrospinal fluid barrier, has the capacity to remove beta-amyloid (A?) from the cerebrospinal fluid. Our previous work indicates that exposure to lead (Pb) results in A? accumulation in the CP by decreasing the expression of low density lipoprotein receptor protein-1 (LRP1), a protein involved in the transport and clearance of A?. The current study was designed to explore the relationship between A? accumulation, protein kinase C (PKC) activi...

  3. Structural and Functional Properties of Peptides Based on the N-terminus of HIV-1 gp41 and the C-terminus of the Amyloid-Beta Protein

    OpenAIRE

    Gordon, Larry M.; Nisthal, Alex; Lee, Andy B.; Eskandari, Sepehr; Ruchala, Piotr; Jung, Chun-ling; Waring, Alan J.; Mobley, Patrick W.

    2008-01-01

    Given their high alanine and glycine levels, plaque formation, ?-helix to ?-sheet interconversion and fusogenicity, FP (i.e., the N-terminal fusion peptide of HIV-1 gp41; 23 residues) and amyloids were proposed as belonging to the same protein superfamily. Here, we further test whether FP may exhibit ‘amyloid-like’ characteristics, by contrasting its structural and functional properties with those of A?(26–42), a 17-residue peptide from the C-terminus of the amyloid-beta protein resp...

  4. Protein Folding and Aggregation into Amyloid: The Interference by Natural Phenolic Compounds

    OpenAIRE

    Massimo Stefani; Stefania Rigacci

    2013-01-01

    Amyloid aggregation is a hallmark of several degenerative diseases affecting the brain or peripheral tissues, whose intermediates (oligomers, protofibrils) and final mature fibrils display different toxicity. Consequently, compounds counteracting amyloid aggregation have been investigated for their ability (i) to stabilize toxic amyloid precursors; (ii) to prevent the growth of toxic oligomers or speed that of fibrils; (iii) to inhibit fibril growth and deposition; (iv) to disassemble preform...

  5. Effects of the English (H6R) and Tottori (D7N) Familial Alzheimer Disease Mutations on Amyloid ?-Protein Assembly and Toxicity*

    OpenAIRE

    Ono, Kenjiro; Condron, Margaret M.; David B. Teplow

    2010-01-01

    Mutations in the amyloid ?-protein (A?) precursor gene cause autosomal dominant Alzheimer disease in a number of kindreds. In two such kindreds, the English and the Tottori, the mutations produce amyloid ?-proteins containing amino acid substitutions, H6R and D7N, respectively, at the peptide N terminus. To elucidate the structural and biological effects of the mutations, we began by examining monomer conformational dynamics and oligomerization. Relative to their wild type homologues, and ...

  6. Soluble amyloid precursor protein induces rapid neural differentiation of human embryonic stem cells

    DEFF Research Database (Denmark)

    Freude, Karla Kristine; Penjwini, Mahmud

    2011-01-01

    Human embryonic stem cells (hESCs) offer tremendous potential for not only treating neurological disorders but also for their ability to serve as vital reagents to model and investigate human disease. To further our understanding of a key protein involved in Alzheimer disease pathogenesis, we stably overexpressed amyloid precursor protein (APP) in hESCs. Remarkably, we found that APP overexpression in hESCs caused a rapid and robust differentiation of pluripotent stem cells toward a neural fate. Despite maintenance in standard hESC media, up to 80% of cells expressed the neural stem cell marker nestin, and 65% exhibited the more mature neural marker ?-3 tubulin within just 5 days of passaging. To elucidate the mechanism underlying the effects of APP on neural differentiation, we examined the proteolysis of APP and performed both gain of function and loss of function experiments. Taken together, our results demonstrate that the N-terminal secreted soluble forms of APP (in particular sAPP?) robustly drive neural differentiation of hESCs. Our findings not only reveal a novel and intriguing role for APP in neural lineage commitment but also identify a straightforward and rapid approach to generate large numbers of neurons from human embryonic stem cells. These novel APP-hESC lines represent a valuable tool to investigate the potential role of APP in development and neurodegeneration and allow for insights into physiological functions of this protein.

  7. Chondroitin Sulfate Accelerates Trans-Golgi-to-Surface Transport of Proteoglycan Amyloid Precursor Protein.

    Science.gov (United States)

    Mihov, Deyan; Raja, Eva; Spiess, Martin

    2015-08-01

    The amyloid precursor protein (APP) is a membrane protein implicated in the pathogenesis of Alzheimer's disease. APP is a part-time proteoglycan, as splice variants lacking exon 15 are modified by a chondroitin sulfate glycosaminoglycan (GAG) chain. Investigating the effect of the GAG chain on the trafficking of APP in non-polarized cells, we found it to increase the steady-state surface-to-intracellular distribution, to reduce the rate of endocytosis and to accelerate transport kinetics from the trans-Golgi network (TGN) to the plasma membrane. Deletion of the cytosolic domain resulted in delayed surface arrival of GAG-free APP, but did not affect the rapid export kinetics of the proteoglycan form. Protein-free GAG chains showed the same TGN-to-cell surface transport kinetics as proteoglycan APP. Endosome ablation experiments were performed to distinguish between indirect endosomal and direct pathways to the cell surface. Surprisingly, TGN-to-cell surface transport of both GAG-free and proteoglycan APP was found to be indirect via transferrin-positive endosomes. Our results show that GAGs act as alternative sorting determinants in cellular APP transport that are dominant over cytoplasmic signals and involve distinct sorting mechanisms. PMID:25951880

  8. Serum C-reactive protein levels and falciparum malaria.

    Science.gov (United States)

    Naik, P; Voller, A

    1984-01-01

    A microplate ELISA was developed to measure C-reactive protein (CRP) and it was used to establish the relationship between CRP levels and malaria. Highest serum CRP levels were found in African patients with high Plasmodium falciparum parastaemia. However, even African children with lower parasitaemia had higher CRP levels than others without parasitaemia. All African groups studied had CRP levels above those of a control UK group. PMID:6398534

  9. Serum protein profile of Crohn's disease treated with infliximab.

    Science.gov (United States)

    Gazouli, Maria; Anagnostopoulos, Athanasios K; Papadopoulou, Aggeliki; Vaiopoulou, Anna; Papamichael, Konstantinos; Mantzaris, Gerassimos; Theodoropoulos, George E; Anagnou, Nicholas P; Tsangaris, George Th

    2013-11-01

    The infliximab (IFX) has dramatically improved the treatment of Crohn's disease (CD). However, the need for predictive factors, indicative of patients' response to IFX, has yet to be met. In the current study, proteomics technologies were employed in order to monitor for differences in protein expression in a cohort of patients following IFX administration, aiming at identifying a panel of candidate protein biomarkers of CD, symptomatic of response to treatment. We enrolled 18 patients, who either had achieved clinical and serological remission (Rm, n=6), or response (Rs, n=6) and/or were PNRs (n=6), to IFX. Serum samples were subjected to two-dimensional Gel Electrophoresis. Following evaluation of densitometrical data, protein spots exhibiting differential expression among the groups, were further characterized by MALDI-TOF-MS. Identified proteins where evaluated by immunoblot analysis while functional network association was carried out to asses significance. Proteins apolipoprotein A-I (APOA1), apolipoprotein E (APOE), complement C4-B (CO4B), plasminogen (PLMN), serotransferrin (TRFE), beta-2-glycoprotein 1 (APOH), and clusterin (CLUS) were found to be up-regulated in the PNR and Rs groups whereas their levels displayed no changes in the Rm group when compared to baseline samples. Additionally, leucine-rich alpha-2-glycoprotein (A2GL), vitamin D-binding protein (VTDB), alpha-1B-glycoprotein (A1BG) and complement C1r subcomponent (C1R) were significantly increased in the serum of the Rm group. Through the incorporation of proteomics technologies, novel serum marker-molecules demonstrating high sensitivity and specificity are introduced, hence offering an innovative approach regarding the evaluation of CD patients' response to IFX therapy. PMID:23562004

  10. Absence of Nitric Oxide Synthase 3 Increases Amyloid ?-Protein Pathology in Tg-5xFAD Mice

    Directory of Open Access Journals (Sweden)

    Zishuo Ian Hu

    2013-06-01

    Full Text Available Aim: The abnormal accumulation, assembly and deposition of the amyloid ?-protein (A? are prominent pathological features of patients with Alzheimer’s disease (AD and related disorders. A number of factors in the brain can influence A? accumulation and associated pathologies. The aim of the present study was to determine the consequences of deleting nitric oxide synthase (NOS 3, the endothelial form of NOS, in Tg-5xFAD mice, a model of parenchymal AD-like amyloid pathology. Methods: Tg-5xFAD mice were bred with NOS3-/- mice. Cohorts of Tg-5xFAD mice and bigenic Tg-5xFAD/NOS3-/- mice were aged to six months followed by collection of the blood and brain tissues from the mice for biochemical and pathological analyses. Results: ELISA analyses show that the absence of NOS3 results in elevated levels of cerebral and plasma A? peptides in Tg-5xFAD mice. Immunohistochemical analyses show that the absence of NOS3 increased the amount of parenchymal A? deposition and fibrillar amyloid accumulation in Tg-5xFAD mice. The elevated levels of A? were not due to changes in the expression levels of transgene encoded human amyloid precursor protein (APP, endogenous ?-secretase, or increased proteolytic processing of APP. Conclusions: The results from this study suggest that the loss of NOS3 activity enhances A? pathology in Tg-5xFAD mice. These findings are similar to previous studies of NOS2 deletion suggesting that reduced NOS activity and NO levels enhance amyloid-associated pathologies in human APP transgenic mice.

  11. Prognostic value of Serum C-Reactive Protein in Malaria

    Directory of Open Access Journals (Sweden)

    Mitul Chhatriwala

    2014-10-01

    Full Text Available Introduction: C-Reactive protein is known as a marker of morbidity and mortality in malaria. It correlates closely with other complications in malaria and can be used to predict severity. Thus, measurement of CRP can be useful in understanding the pathogenesis of severe malaria. However, studies regarding CRP in malaria are very rare from India.Aims & objectives: The present study aimed to study CRP levels in patients with malarial infection and to find out the usefulness of CRP in assessment of disease severity.Material and Methods: The study population consisted of 50 patients with malarial infection, of them 29 were Plasmodium falciparum, 21 were P. vivax-infected and no patient found with dual infection in our study. Out of 50 patients, 17 patients required admission for various reasons and of them 5 patients died. Serum C-reactive protein concentrations were measured in all the patients. Percentage parasitemia, platelet count and Liver function tests were measured on the day of admission. Statistical tests included were Students t-test, chi-square test and Pearson correlation coefficient.Results: Serum C-reactive protein concentration was significantly elevated in all the malaria patients and it was significantly higher (P<0.01 in Plasmodium falciparum (28.81 ± 10.91 mg/L as compared to P. Vivax malaria (15.91 ± 8.73 mg/L . Admitted patients had higher CRP levels (47.80 ± 12.73 mg/L compared to patients treated on OPD basis (24.14 ± 12.17 mg/L (P < 0.0001. Also CRP levels were higher in patients with multiple complications and in the patients who died compared to survivors. A significant positive correlation was found between serum CRP and percentage parasitemia, serum bilirubin, serum alanine aminotransferase and serum aspartate aminotransferase in P. falciparum infected patients but not in P. Vivax.Conclusion: There is significant increase in C-reactive protein in malaria infected patients and can be considered a cost - effective and reliable tool in assessment of prognosis in malaria.

  12. Interaction of thorium with blood serum proteins in vivo

    International Nuclear Information System (INIS)

    The distribution of thorium in the blood serum of rats was studied in vivo, using carrier-free 234Th and 59Fe-citrate solution. The chromatographic data presented in this short communication indicate that, as with plutonium, the iron-transport protein, transferrin, is the main carrier protein for thorium. Also, like plutonium, thorium could be displaced from the transferrin complex by excess iron. Further investigations are required to determine if plutonium and thorium are bound to the same sites on the transferrin molecule as iron and whether the binding mechanisms are similar. (U.K.)

  13. Influence of transportation on serum concentrations of acute phase proteins in horse.

    Science.gov (United States)

    Casella, S; Fazio, F; Giannetto, C; Giudice, E; Piccione, G

    2012-10-01

    The modifications of Haptoglobin (Hp), Serum Amyloid A (SAA), Fibrinogen (Fbg) and White Blood Cells (WBCs) were evaluated in 15 Saddle Italian horses. Ten horses were transported covering a distance of about 320 km within 4 h with an average speed of 80 km/h (experimental group) and five horses were not subject to transportation (control group). Blood was collected via jugular venipuncture before the transportation (T0), immediately after the transportation (T1), 12 (T12), 24 (T24) and 48 (T48)hours after the transportation in experimental group and at the same time point in control group. For each parameter statistical analysis of different groups and sampling time was performed using a two-way analysis of covariance, with the data before the transportation (T0) as the covariate, by the GLM procedure of SAS. For all parameters the interaction (Group × Time) was tested and it was resulted no significant. The application of statistical analysis showed significant differences between the control group and horses subjected to transportation (P<0.01), and the influence of sampling time (P<0.05) on Hp, SAA and WBCs. These modifications appeared to be innovative showing that equine Hp, generally considered as moderate acute phase protein, increases more rapidly than the SAA after transportation-induced stress. PMID:22296939

  14. Unraveling the mysteries of serum albumin – more than just a serum protein.

    Directory of Open Access Journals (Sweden)

    AngelicaM.Merlot

    2014-08-01

    Full Text Available Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are responsible for albumin uptake. Initially, the cellular uptake and intracellular localization of albumin was unknown due to the large confinement of the protein within the vascular and interstitial compartment of the body. Studies have since assessed the intracellular localization of albumin in order to understand the mechanisms and pathways responsible for its uptake, distribution and catabolism in multiple tissues, and this is reviewed herein.

  15. Unraveling the mysteries of serum albumin—more than just a serum protein

    Science.gov (United States)

    Merlot, Angelica M.; Kalinowski, Danuta S.; Richardson, Des R.

    2014-01-01

    Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are responsible for albumin uptake. Initially, the cellular uptake and intracellular localization of albumin was unknown due to the large confinement of the protein within the vascular and interstitial compartment of the body. Studies have since assessed the intracellular localization of albumin in order to understand the mechanisms and pathways responsible for its uptake, distribution and catabolism in multiple tissues, and this is reviewed herein. PMID:25161624

  16. Unraveling the mysteries of serum albumin-more than just a serum protein.

    Science.gov (United States)

    Merlot, Angelica M; Kalinowski, Danuta S; Richardson, Des R

    2014-01-01

    Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are responsible for albumin uptake. Initially, the cellular uptake and intracellular localization of albumin was unknown due to the large confinement of the protein within the vascular and interstitial compartment of the body. Studies have since assessed the intracellular localization of albumin in order to understand the mechanisms and pathways responsible for its uptake, distribution and catabolism in multiple tissues, and this is reviewed herein. PMID:25161624

  17. The Functions of the Amyloid Precursor Protein Gene and Its Derivative Peptides: II Experimental Evidence and Clinical Studies

    Directory of Open Access Journals (Sweden)

    Peter K. Panegyres

    2011-09-01

    Full Text Available In vitro studies suggest Amyloid Precursor Protein Gene (APP is involved in interaction with the extracellular matrix, neurite growth, adhesion, development, synaptic function, platelet function, and interaction with GTP binding proteins. In vivo experiments show a role in embryonic development, response to cerebral excitotoxicity and gliosis, response to brain injury including ischaemia, hypothalamic function, locomotor function, learning and memory. In vitro observations indicate A? has a role in amyloid formation, excitotoxic neuronal injury, tachykinin interaction, endothelial vasoconstrictor response, calcium and oxidative stress, free radical interaction, cell membrane fluidity, apoptosis, astrocyte stimulation, and microglial interaction. Other studies suggest important roles for A? oligomers in synaptic function and as an antimicrobial peptide. In vivo investigations show involvement in memory function, the blood brain barrier, and tachykinin response to cerebral injury.

  18. Adaptor protein 2-mediated endocytosis of the ?-secretase BACE1 is dispensable for amyloid precursor protein processing.

    Science.gov (United States)

    Prabhu, Yogikala; Burgos, Patricia V; Schindler, Christina; Farías, Ginny G; Magadán, Javier G; Bonifacino, Juan S

    2012-06-01

    The ?-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1) is a transmembrane aspartyl protease that catalyzes the proteolytic processing of APP and other plasma membrane protein precursors. BACE1 cycles between the trans-Golgi network (TGN), the plasma membrane, and endosomes by virtue of signals contained within its cytosolic C-terminal domain. One of these signals is the DXXLL-motif sequence DISLL, which controls transport between the TGN and endosomes via interaction with GGA proteins. Here we show that the DISLL sequence is embedded within a longer [DE]XXXL[LI]-motif sequence, DDISLL, which mediates internalization from the plasma membrane by interaction with the clathrin-associated, heterotetrameric adaptor protein 2 (AP-2) complex. Mutation of this signal or knockdown of either AP-2 or clathrin decreases endosomal localization and increases plasma membrane localization of BACE1. Remarkably, internalization-defective BACE1 is able to cleave an APP mutant that itself cannot be delivered to endosomes. The drug brefeldin A reversibly prevents BACE1-catalyzed APP cleavage, ruling out that this reaction occurs in the endoplasmic reticulum (ER) or ER-Golgi intermediate compartment. Taken together, these observations support the notion that BACE1 is capable of cleaving APP in late compartments of the secretory pathway. PMID:22553349

  19. Adaptor protein 2–mediated endocytosis of the ?-secretase BACE1 is dispensable for amyloid precursor protein processing

    Science.gov (United States)

    Prabhu, Yogikala; Burgos, Patricia V.; Schindler, Christina; Farías, Ginny G.; Magadár, Javier G.; Bonifacino, Juan S.

    2012-01-01

    The ?-site amyloid precursor protein (APP)–cleaving enzyme 1 (BACE1) is a transmembrane aspartyl protease that catalyzes the proteolytic processing of APP and other plasma membrane protein precursors. BACE1 cycles between the trans-Golgi network (TGN), the plasma membrane, and endosomes by virtue of signals contained within its cytosolic C-terminal domain. One of these signals is the DXXLL-motif sequence DISLL, which controls transport between the TGN and endosomes via interaction with GGA proteins. Here we show that the DISLL sequence is embedded within a longer [DE]XXXL[LI]-motif sequence, DDISLL, which mediates internalization from the plasma membrane by interaction with the clathrin-associated, heterotetrameric adaptor protein 2 (AP-2) complex. Mutation of this signal or knockdown of either AP-2 or clathrin decreases endosomal localization and increases plasma membrane localization of BACE1. Remarkably, internalization-defective BACE1 is able to cleave an APP mutant that itself cannot be delivered to endosomes. The drug brefeldin A reversibly prevents BACE1-catalyzed APP cleavage, ruling out that this reaction occurs in the endoplasmic reticulum (ER) or ER–Golgi intermediate compartment. Taken together, these observations support the notion that BACE1 is capable of cleaving APP in late compartments of the secretory pathway. PMID:22553349

  20. An involvement of SR-B1 mediated p38 MAPK signaling pathway in serum amyloid A-induced angiogenesis in rheumatoid arthritis.

    Science.gov (United States)

    Hong, Chengcheng; Shen, Chen; Ding, Hongmei; Huang, Shanshan; Mu, Yun; Su, Huihui; Wei, Wei; Ma, Jun; Zheng, Fang

    2015-08-01

    Serum amyloid A (SAA) has been reported high expression in autoimmune diseases, such as rheumatoid arthritis (RA). However, detailed molecular mechanisms induced by SAA in the pathogenesis of RA are still unclear. Herein, we focused on the role of SAA-SR-B1 mediated p38 MAPK signaling pathway in the process of RA angiogenesis. Our results showed that both SAA and SR-B1 predominantly localized to vascular endothelial cells, lining and sublining layers in RA synovium. In a series of in vitro experiments with human umbilical vein endothelial cells (HUVECs), SAA induced the endothelial cells (ECs) proliferation, migration and tube formation. However, blockage of SR-B1 and p38 MAPK inhibited SAA-induced cells proliferation, migration and tube formation. In conclusion, our data showed a possible molecular mechanism for SAA-SR-B1 induced angiogenesis events via p38 MAPK signaling pathway. PMID:25932604

  1. 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)

    F.H.D, González; F.H, Ruipérez; J.M, Sánchez; J.C, Souza; S, Martínez-Subiela; J.J, Cerón.

    2010-12-01

    Full Text Available 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.

  2. Chronic treatment with amyloid beta(1-42) inhibits non-cholinergic high-affinity choline transport in NG108-15 cells through protein kinase C signaling.

    Science.gov (United States)

    Nováková, Jana; Mikasová, Lenka; Machová, Eva; Lisá, V?ra; Dolezal, Vladimír

    2005-11-16

    We investigated the influence of the amyloid-beta-peptide(1-42) on hemicholinum-3-sensitive high-affinity choline uptake in NG108-15 cells. RT-PCR analysis revealed the presence of mRNA for a choline transporter-like protein but not for cholinergic high-affinity choline transporter. Differentiation of cells increased both hemicholinum-3-sensitive choline uptake and high-affinity hemicholinium-3 binding. This transport was not influenced by tenfold excess of carnitine. Continuous presence of submicromolar concentrations of amyloid-beta-peptide(1-42) during differentiation resulted in a decrease of both choline uptake and hemicholinium-3 binding. These effects were not present when amyloid-beta-peptide(1-42) was added 5 min prior to measurements. Neither differentiation nor amyloid-beta-peptide(1-42) treatment changed levels of choline transporter-like protein mRNA. Protein kinase C inhibition by staurosporine or its inactivation by continuous presence of tetradecanoyl phorbol acetate prevented the inhibitory effect of amyloid-beta-peptide(1-42) treatment on choline uptake. Activation of protein kinase C by tetradecanoyl phorbol acetate during measurement had inhibitory effect on choline uptake in control but not amyloid-beta-peptide(1-42)-treated cells. The concentration of amyloid-beta-peptide(1-42) maximally effective on hemicholinium-3-sensitive choline uptake had no effect on cell growth, oxidative activity, membrane integrity, number of surface muscarinic receptors, caspase-3 and -8 activities, or uptake of deoxyglucose. Results demonstrate that long-term treatment with non-toxic concentrations of amyloid-beta-peptide(1-42) downregulates choline uptake presumably mediated by a choline transporter-like protein through activation of protein kinase C signaling. The decrease of choline uptake may have relevance to the pathogenesis of Alzheimer's disease. PMID:16256077

  3. SorLA is a molecular link for retromer-dependent sorting of the Amyloid precursor protein

    OpenAIRE

    Fjorback, Anja W.; Andersen, Olav M.

    2012-01-01

    Deficiency in the retromer sorting pathway is known to be associated with the onset of Alzheimer disease (AD), and has been suggested to involve regulation of Amyloid precursor protein (APP) trafficking. Absence of the APP sorting receptor sorLA is also associated to AD, as amyloidogenic processing of APP is increased due to missorting. Reduced activity of either retromer or sorLA thus both lead to enhanced amyloidogenic APP processing, and these pathways are therefore important factors for u...

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

    OpenAIRE

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

    2012-01-01

    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...

  5. Valsartan lowers brain ?-amyloid protein levels and improves spatial learning in a mouse model of Alzheimer disease

    OpenAIRE

    Jun WANG; Ho, Lap; Chen, Linghong; Zhao, Zhong; Zhao, Wei; Qian, Xianjuan; Humala, Nelson; Seror, Ilana; Bartholomew, Sadie; Rosendorff, Clive; Pasinetti, Giulio Maria

    2007-01-01

    Recent epidemiological evidence suggests that some antihypertensive medications may reduce the risk for Alzheimer disease (AD). We screened 55 clinically prescribed antihypertensive medications for AD-modifying activity using primary cortico-hippocampal neuron cultures generated from the Tg2576 AD mouse model. These agents represent all drug classes used for hypertension pharmacotherapy. We identified 7 candidate antihypertensive agents that significantly reduced AD-type ?-amyloid protein (A...

  6. Amyloid -protein (A) assembly: A40 and A42 oligomerize through distinct pathways

    Science.gov (United States)

    Bitan, Gal; Kirkitadze, Marina D.; Lomakin, Aleksey; Vollers, Sabrina S.; Benedek, George B.; Teplow, David B.

    2003-01-01

    Amyloid -protein (A) is linked to neuronal injury and death in Alzheimer's disease (AD). Of particular relevance for elucidating the role of A in AD is new evidence that oligomeric forms of A are potent neurotoxins that play a major role in neurodegeneration and the strong association of the 42-residue form of A, A42, with the disease. Detailed knowledge of the structure and assembly dynamics of A thus is important for the development of properly targeted AD therapeutics. Recently, we have shown that A oligomers can be cross-linked efficiently, and their relative abundances quantified, by using the technique of photo-induced cross-linking of unmodified proteins (PICUP). Here, PICUP, size-exclusion chromatography, dynamic light scattering, circular dichroism spectroscopy, and electron microscopy have been combined to elucidate fundamental features of the early assembly of A40 and A42. Carefully prepared aggregate-free A40 existed as monomers, dimers, trimers, and tetramers, in rapid equilibrium. In contrast, A42 preferentially formed pentamer/hexamer units (paranuclei) that assembled further to form beaded superstructures similar to early protofibrils. Addition of Ile-41 to A40 was sufficient to induce formation of paranuclei, but the presence of Ala-42 was required for their further association. These data demonstrate that A42 assembly involves formation of several distinct transient structures that gradually rearrange into protofibrils. The strong etiologic association of A42 with AD may thus be a result of assemblies formed at the earliest stages of peptide oligomerization.

  7. 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)

    Nielsen, EH; SØrensen, Inge Juul

    1994-01-01

    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 and the inner diameter 3.2 nm. The thickness of single and double pentamers was 4.1 and 8.7 nm, respectively. Quadruple pentamers were occasionally seen. The self-aggregation of human SAP molecules was investigated in the presence and absence of calcium ions at different concentrations. In calcium-free solutions few and mostly small SAP aggregates were seen. After addition of calcium at increasing concentration the aggregates grew in size and crystalline-like structures were formed already at 2 mM calcium. At 25 mM calcium, large aggregates with a crystalline array occasionally exhibiting cylinders predominated. Binding of the ligands heparin and heparan sulphate to SAP completely abolished the calcium-enhanced aggregation, but the distribution of the SAP molecules was affected, resulting in strands or groups of adjacent molecules. The electrophoretic mobility of SAP was moreover significantly altered after its calcium-dependent reaction with these ligands. We conclude that purified SAP has a tendency to double pentamer formation and self-aggregation also in the absence of calcium ions. However, aggregation is greatly enhanced even at low concentrations (2 mM) of calcium. SAP's tendency to self-aggregation is abolished after its binding to heparin or heparin sulphate. Furthermore, our TEM studies indicate that purified human SAP freed of its natural ligands has the double pentameric form, whereas the electrophoretic investigations suggest that SAP's interaction with low-molecular-weight natural ligands in serum prevents homodimerization and self-aggregation.

  8. Peptides of Presenilin-1 Bind the Amyloid Precursor Protein Ectodomain and Offer a Novel and Specific Therapeutic Approach to Reduce ß-Amyloid in Alzheimer’s Disease

    Science.gov (United States)

    Dewji, Nazneen N.; Singer, S. Jonathan; Masliah, Eliezer; Rockenstein, Edward; Kim, Mihyun; Harber, Martha; Horwood, Taylor

    2015-01-01

    ?-Amyloid (A?) accumulation in the brain is widely accepted to be critical to the development of Alzheimer’s disease (AD). Current efforts at reducing toxic A?40 or 42 have largely focused on modulating ?-secretase activity to produce shorter, less toxic A?, while attempting to spare other secretase functions. In this paper we provide data that offer the potential for a new approach for the treatment of AD. The method is based on our previous findings that the production of A? from the interaction between the ?-amyloid precursor protein (APP) and Presenilin (PS), as part of the ?-secretase complex, in cell culture is largely inhibited if the entire water-soluble NH2-terminal domain of PS is first added to the culture. Here we demonstrate that two small, non-overlapping water-soluble peptides from the PS-1 NH2-terminal domain can substantially and specifically inhibit the production of total A? as well as A?40 and 42 in vitro and in vivo in the brains of APP transgenic mice. These results suggest that the inhibitory activity of the entire amino terminal domain of PS-1 on A? production is largely focused in a few smaller sequences within that domain. Using biolayer interferometry and confocal microscopy we provide evidence that peptides effective in reducing A? give a strong, specific and biologically relevant binding with the purified ectodomain of APP 695. Finally, we demonstrate that the reduction of A? by the peptides does not affect the catalytic activities of ?- or ?-secretase, or the level of APP. P4 and P8 are the first reported protein site-specific small peptides to reduce A? production in model systems of AD. These peptides and their derivatives offer new potential drug candidates for the treatment of AD. PMID:25923432

  9. Structures of segments of [alpha]-synuclein fused to maltose-binding protein suggest intermediate states during amyloid formation

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Minglei; Cascio, Duilio; Sawaya, Michael R.; Eisenberg, David (UCLA)

    2011-08-29

    Aggregates of the protein {alpha}-synuclein are the main component of Lewy bodies, the hallmark of Parkinson's disease. {alpha}-Synuclein aggregates are also found in many human neurodegenerative diseases known as synucleinopathies. In vivo, {alpha}-synuclein associates with membranes and adopts {alpha}-helical conformations. The details of how {alpha}-synuclein converts from the functional native state to amyloid aggregates remain unknown. In this study, we use maltose-binding protein (MBP) as a carrier to crystallize segments of {alpha}-synuclein. From crystal structures of fusions between MBP and four segments of {alpha}-synuclein, we have been able to trace a virtual model of the first 72 residues of {alpha}-synuclein. Instead of a mostly {alpha}-helical conformation observed in the lipid environment, our crystal structures show {alpha}-helices only at residues 1-13 and 20-34. The remaining segments are extended loops or coils. All of the predicted fiber-forming segments based on the 3D profile method are in extended conformations. We further show that the MBP fusion proteins with fiber-forming segments from {alpha}-synuclein can also form fiber-like nano-crystals or amyloid-like fibrils. Our structures suggest intermediate states during amyloid formation of {alpha}-synuclein.

  10. Structural and Functional Properties of Peptides Based on the N-terminus of HIV-1 gp41 and the C-terminus of the Amyloid-Beta Protein

    Science.gov (United States)

    Gordon, Larry M.; Nisthal, Alex; Lee, Andy B.; Eskandari, Sepehr; Ruchala, Piotr; Jung, Chun-Ling; Waring, Alan J.; Mobley, Patrick W.

    2008-01-01

    Given their high alanine and glycine levels, plaque formation, ?-helix to ?-sheet interconversion and fusogenicity, FP (i.e., the N-terminal fusion peptide of HIV-1 gp41; 23 residues) and amyloids were proposed as belonging to the same protein superfamily. Here, we further test whether FP may exhibit ‘amyloid-like’ characteristics, by contrasting its structural and functional properties with those of A?(26–42), a 17-residue peptide from the C-terminus of the amyloid-beta protein responsible for Alzheimer’s. FTIR spectroscopy, electron microscopy, light scattering and predicted amyloid structure aggregation (PASTA) indicated that aqueous FP and A?(26–42) formed similar networked ?-sheet fibrils, although the FP fibril interactions were weaker. FP and A?(26–42) both lysed and aggregated human erythrocytes, with the hemolysis-onsets correlated with the conversion of ?-helix to ?-sheet for each peptide in liposomes. Congo red (CR), a marker of amyloid plaques in situ, similarly inhibited either FP- or A?(26–42)-induced hemolysis, and surface plasmon resonance indicated that this may be due to direct CR-peptide binding. These findings suggest that membrane-bound ?-sheets of FP may contribute to the cytopathicity of HIV in vivo through an amyloid-type mechanism, and support the classification of HIV-1 FP as an ‘amyloid homolog’ (or ‘amylog’). PMID:18515070

  11. Conformational Flexibility of Y145Stop Human Prion Protein Amyloid Fibrils Probed by Solid-State Nuclear Magnetic Resonance Spectroscopy

    Science.gov (United States)

    Helmus, Jonathan J.; Surewicz, Krystyna; Surewicz, Witold K.; Jaroniec, Christopher P.

    2010-01-01

    Amyloid aggregates of a C-truncated Y145Stop mutant of human prion protein, huPrP23-144, associated with a heritable amyloid angiopathy, have previously been shown to contain a compact, relatively rigid and ?-sheet-rich ?30-residue amyloid core near the C-terminus under physiologically relevant conditions. In contrast, the remaining huPrP23-144 residues display considerable conformational dynamics, as evidenced by the absence of corresponding signals in cross-polarization (CP) based solid-state NMR (SSNMR) spectra under ambient conditions and their emergence in analogous spectra recorded at low temperature on frozen fibril samples. Here we present the direct observation of residues comprising the flexible N-terminal domain of huPrP23-144 amyloid by using 2D J-coupling based magic-angle spinning (MAS) SSNMR techniques. Chemical shifts for these residues indicate that the N-terminal domain is effectively an ensemble of protein chains with random-coil-like conformations. Interestingly, a detailed analysis of signal intensities in CP-based 3D SSNMR spectra suggests that non-negligible molecular motions may also be occurring on the NMR timescale within the relatively rigid core of huPrP23-144 amyloid. To further investigate this hypothesis, quantitative measurements of backbone dipolar order parameters and transverse spin relaxation rates were performed for the core residues. The observed order parameters indicate that, on the sub-?s timescale, these residues are effectively rigid and experience only highly restricted and relatively uniform motions similar to those characteristic for well-structured regions of microcrystalline proteins. On the other hand significant variations in magnitude of transverse spin relaxation rates were noted for residues present at different locations within the core region and correlated with observed differences in spectral intensities. While interpreted only qualitatively at the present time, the extent of the observed variations in transverse relaxation rates is consistent with the presence of relatively slow, ?s-ms timescale chemical exchange type phenomena within the huPrP23-144 amyloid core. PMID:20121096

  12. Relationship between Acute Phase Proteins and Serum Fatty Acid Composition in Morbidly Obese Patients

    OpenAIRE

    Ricardo Fernandes; Bruna Teles Soares Beserra; Raphael Salles Granato Cunha; Elaine Hillesheim; Carolina de Quadros Camargo; Danielle Cristina Tonello Pequito; Isabela Coelho de Castro; Luiz Cláudio Fernandes; Everson Araújo Nunes; Erasmo Benício Santos de Moraes Trindade

    2013-01-01

    Background. Obesity is considered a low-grade inflammatory state and has been associated with increased acute phase proteins as well as changes in serum fatty acids. Few studies have assessed associations between acute phase proteins and serum fatty acids in morbidly obese patients. Objective. To investigate the relationship between acute phase proteins (C-Reactive Protein, Orosomucoid, and Albumin) and serum fatty acids in morbidly obese patients. Methods. Twenty-two morbidly obese patients ...

  13. Curli functional amyloid systems are phylogenetically widespread and display large diversity in operon and protein structure

    DEFF Research Database (Denmark)

    Dueholm, Morten S; Albertsen, Mads

    2012-01-01

    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.

  14. Amyloid-? Activates Microglia and Regulates Protein Expression in a Manner Similar to Prions.

    Science.gov (United States)

    Tu, Jian; Chen, Baian; Yang, Lifeng; Qi, Kezong; Lu, Jing; Zhao, Deming

    2015-06-01

    Prions are the only convincingly demonstrated proteinaceous infectious particle, yet recent studies find that amyloid-? peptide (A?) aggregates are capable of self-propagation, which induces amyloidosis pathology in Alzheimer's disease (AD) model mice that is similar to the self-propagation phenomenon of prions in neurons. Gliosis is a common hallmark of AD and prion diseases, in which activated microglia accumulate around abnormal protein deposits. Analyses of the characteristics of activated microglia induced by A? in comparison with those induced by prions will provide new insight into the pathogenesis of AD. Therefore, we compared the characteristics of BV-2 cells (model microglia) activated by A? fibrillar peptides (A?1-42) and prions (PrP106-126). A?1-42 and PrP106-126, as well as the supernatants of the media collected from BV-2 cells cocultured with A?1-42 and PrP106-126, were potent activators of BV-2 microglial activity, but the chemotaxis index (CI) induced by A?1-42 was significantly higher than that induced by PrP106-126 at each concentration. A?1-42 and PrP106-126 increased the proliferation index (PI) and upregulated monocyte chemoattractant protein-1 (MCP-1) and transforming growth factor beta 1 (TGF-?1) expression after 12 h of exposure. Our results show that A? activates microglia and regulates microglial protein expression in a manner similar to prions and, thus, provide new insight into the pathogenesis of AD. PMID:25869610

  15. Modeling sporadic alzheimer's disease: the insulin resistant brain state generates multiple long-term morphobiological abnormalities inclusive hyperphosphorylated tau protein and amyloid-beta. A Synthesis

    OpenAIRE

    Salkovic-Petrisic, M; Osmanovic, J; E Grünblatt; Riederer, P; Hoyer, S

    2009-01-01

    Nosologically, Alzheimer's disease (AD) is not a single disorder. Missense gene mutations involved in increased formation of the amyloid-beta protein precursor derivatives amyloid-beta (Abeta)_{1-40} and Abeta_{1-42/43} lead to autosomal dominant familial AD, found in the minority of AD cases. However, millions of subjects suffer from sporadic AD (sAD) of late onset, for which no convincing evidence suggests Abeta as the primary disease-generating compound. Environmental factors operating dur...

  16. Ternary complex formation between yttrium, phosphate and serum proteins

    International Nuclear Information System (INIS)

    In order to investigate the transport form of yttrium(III) in physiological systems the binding of radioyttrium to human serum proteins in the presence of different endogenous anions was evaluated using gel filtration (GF) and ultrafiltration (UF) techniques. Among the most endogenous anions (hydroxide, sulfate, carbonate, citrate, fluoride et.) no influence on the binding of 88Y labeled Y(III) to human serum albumin (HSA) could be observed. The binding of Y(III) to HSA was as low as in pure aqueous isotonic solution (5% GF and 20% UF). The presence of phosphate anions in the protein-metal solution, however, significantly increased the binding of Y(III) to HSA (80% GF and 95% UF). Phosphate in turn was bound to serum proteins only in the presence of Y(III) as determined with 32P labeled PO43-. The simultaneous binding of phosphate and Y(III) to HSA occurred in a stoichiometric ratio of 1:1. A ternary complex formation between Y(III), phosphate and HSA was postulated. Information about the binding mechanism could be obtained by an analysis of the pH stability of the ternary complex which is stable in the neutral pH region of 5.5 to 8.5. An estimate of the complex formation constant of the ternary compound with HSA by competitive experiments yielded lg?=10.9 (I=0.15 M; T=25 C). The same type of binding could be observed with transferrin and polyglutamic acid. Binding to the latter proved that carboxylate groups of the biomacred that carboxylate groups of the biomacromolecules are responsible for the simultaneous binding of Y(III) and phosphate. (orig.)

  17. A setup for simultaneous measurement of infrared spectra and light scattering signals: Watching amyloid fibrils grow from intact proteins

    International Nuclear Information System (INIS)

    A setup for the simultaneous measurement of mid-infrared spectra and static light scattering is described that can be used for the analysis of the formation of nanoscale and microscopic aggregates from smaller molecules to biopolymers. It can be easily integrated into sample chambers of infrared spectrometers or combined with laser beams from tunable infrared lasers. Here, its use for the analysis of the formation of amyloid fibrils from intact proteins is demonstrated. The formation of amyloid fibrils or plaques from proteins is a widespread and pathogenetic relevant process, and a number of diseases are caused and correlated with the deposition of amyloid fibrils in cells and tissues. The molecular mechanisms of these transformations, however, are still unclear. We report here the simultaneous measurement of infrared spectra and static light scattering for the analysis of fibril formation from egg-white lysozyme. The transformation of the native form into non-native forms rich in ?-sheet structure is measured by analysis of the amide I spectral region in the infrared spectra, which is sensitive for local structures. At the same time, light scattering signals at forward direction as well as the forward/backward ratio, which are sensitive for the number of scattering centers and their approximate sizes, respectively, are collected for the analysis of fibril growth. Thermodynamic and kinetic parameters as well as mechanistic information are deduced from the combination of the two complementary techniques

  18. A setup for simultaneous measurement of infrared spectra and light scattering signals: Watching amyloid fibrils grow from intact proteins

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yang; Maurer, Jürgen; Roth, Andreas; Vogel, Vitali; Winter, Ernst; Mäntele, Werner, E-mail: maentele@biophysik.uni-frankfurt.de [Institut für Biophysik, Goethe-Universität Frankfurt am Main, Max-von Laue-Straße 1, D-60438 Frankfurt am Main (Germany)

    2014-08-15

    A setup for the simultaneous measurement of mid-infrared spectra and static light scattering is described that can be used for the analysis of the formation of nanoscale and microscopic aggregates from smaller molecules to biopolymers. It can be easily integrated into sample chambers of infrared spectrometers or combined with laser beams from tunable infrared lasers. Here, its use for the analysis of the formation of amyloid fibrils from intact proteins is demonstrated. The formation of amyloid fibrils or plaques from proteins is a widespread and pathogenetic relevant process, and a number of diseases are caused and correlated with the deposition of amyloid fibrils in cells and tissues. The molecular mechanisms of these transformations, however, are still unclear. We report here the simultaneous measurement of infrared spectra and static light scattering for the analysis of fibril formation from egg-white lysozyme. The transformation of the native form into non-native forms rich in ?-sheet structure is measured by analysis of the amide I spectral region in the infrared spectra, which is sensitive for local structures. At the same time, light scattering signals at forward direction as well as the forward/backward ratio, which are sensitive for the number of scattering centers and their approximate sizes, respectively, are collected for the analysis of fibril growth. Thermodynamic and kinetic parameters as well as mechanistic information are deduced from the combination of the two complementary techniques.

  19. A setup for simultaneous measurement of infrared spectra and light scattering signals: Watching amyloid fibrils grow from intact proteins

    Science.gov (United States)

    Li, Yang; Maurer, Jürgen; Roth, Andreas; Vogel, Vitali; Winter, Ernst; Mäntele, Werner

    2014-08-01

    A setup for the simultaneous measurement of mid-infrared spectra and static light scattering is described that can be used for the analysis of the formation of nanoscale and microscopic aggregates from smaller molecules to biopolymers. It can be easily integrated into sample chambers of infrared spectrometers or combined with laser beams from tunable infrared lasers. Here, its use for the analysis of the formation of amyloid fibrils from intact proteins is demonstrated. The formation of amyloid fibrils or plaques from proteins is a widespread and pathogenetic relevant process, and a number of diseases are caused and correlated with the deposition of amyloid fibrils in cells and tissues. The molecular mechanisms of these transformations, however, are still unclear. We report here the simultaneous measurement of infrared spectra and static light scattering for the analysis of fibril formation from egg-white lysozyme. The transformation of the native form into non-native forms rich in ?-sheet structure is measured by analysis of the amide I spectral region in the infrared spectra, which is sensitive for local structures. At the same time, light scattering signals at forward direction as well as the forward/backward ratio, which are sensitive for the number of scattering centers and their approximate sizes, respectively, are collected for the analysis of fibril growth. Thermodynamic and kinetic parameters as well as mechanistic information are deduced from the combination of the two complementary techniques.

  20. Soluble amyloid precursor protein ? as blood-based biomarker of Alzheimer's disease.

    Science.gov (United States)

    Perneczky, R; Guo, L H; Kagerbauer, S M; Werle, L; Kurz, A; Martin, J; Alexopoulos, P

    2013-01-01

    The aim of this study was to explore concentrations differences of soluble amyloid precursor protein (sAPP) ? and ? in blood plasma in patients with probable Alzheimer's disease (AD) and cognitively healthy age-matched control subjects, as well as patients with behavioural variant frontotemporal dementia (bvFTD). Concentrations of sAPP? and ? were measured using enzyme-linked immunosorbent assay technology in 80 patients with probable AD, 37 age-matched control subjects and 14 patients with bvFTD. Concentration differences were explored using parametric tests. Significantly decreased plasma concentrations in the AD group compared with both the control group and the bvFTD group were detected for sAPP? (P = 0.03 for both group comparisons), but not for sAPP?. The study provides a further piece of evidence in support of sAPP? as a promising new biomarker of AD, which may potentially improve the diagnostic accuracy of existing markers and also enable a less invasive diagnostic workup. Further research is required to establish normal ranges and to replicate the results in independent cohorts including larger numbers of participants covering a wider spectrum of cognitive impairment. PMID:23423136

  1. Overexpression of amyloid precursor protein increases copper content in HEK293 cells

    International Nuclear Information System (INIS)

    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. Cu2+ 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 Cu2+ reduction and 64Cu uptake in two human HEK293 cell lines overexpressing APP. Our results indicate that Cu2+ reduction increased and cells accumulated larger levels of copper, maintaining cell viability at supra-physiological levels of Cu2+ ions. Moreover, wild-type cells exposed to both Cu2+ 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 Cu2+ reductase activity and increased 64Cu uptake. We conclude that Cu2+ reduction activity of APP facilitates copper uptake and may represent an early step in cellular copper homeostasis.

  2. Effects of conjugated linoleic acid on cleavage of amyloid precursor protein via PPAR?.

    Science.gov (United States)

    Li, Yi-Chen; Chen, Qing; Wan, Xian-Zi; Yang, Xiang-Ling; Liu, Xin; Zhong, Ling

    2011-12-01

    Conjugated linoleic acid (CLA) plays important roles in physiological conditions. The aim of present study was to explore the effects of CLA on the cleavage of amyloid precursor protein (APP) and the potential mechanism involved. The effects of CLA on intracellular APP, BACE1 (?-site APP Cleaving Enzyme1, BACE1), a disintegrin and metalloprotease (ADAM10) and extracellular sAPP? (soluble) were analyzed by RT-PCR, Western blot and ELISA in SH-SY5Y cells. Our study indicated that CLA significantly decreased the expression of BACE1 and increased the extracellular secretion of sAPP?, but not affected the levels of APP and ADAM10. The study also revealed that the nuclear receptor peroxisome proliferators activated receptor ? (PPAR?) played an important role in the CLA-induced intracellular BACE1 decrease, as well as the extracellular sAPP? increase through knockdown of PPAR? transcription using siRNA. We hypothesize that CLA acts as an agonist or ligand, which binds with PPAR? and leads to the increase in APP cleavage via ?-secretase-mediated pathway and the decrease in the deposition of A?. PMID:21800078

  3. Increased T cell reactivity to amyloid ? protein in older humans and patients with Alzheimer disease

    Science.gov (United States)

    Monsonego, Alon; Zota, Victor; Karni, Arnon; Krieger, Jeffery I.; Bar-Or, Amit; Bitan, Gal; Budson, Andrew E.; Sperling, Reisa; Selkoe, Dennis J.; Weiner, Howard L.

    2003-01-01

    Alzheimer disease (AD) is characterized by the progressive deposition of the 42-residue amyloid ? protein (A?) in brain regions serving memory and cognition. In animal models of AD, immunization with A? results in the clearance of A? deposits from the brain. However, a trial of vaccination with synthetic human A?1–42 in AD resulted in the development of meningoencephalitis in some patients. We measured cellular immune responses to A? in middle-aged and elderly healthy subjects and in patients with AD. A significantly higher proportion of healthy elderly subjects and patients with AD had strong A?-reactive T cell responses than occurred in middle-aged adults. The immunodominant A? epitopes in humans resided in amino acids 16–33. Epitope mapping enabled the identification of MHC/T cell receptor (TCR) contact residues. The occurrence of intrinsic T cell reactivity to the self-antigen A? in humans has implications for the design of A? vaccines, may itself be linked to AD susceptibility and course, and appears to be associated with the aging process. PMID:12897209

  4. Increased T cell reactivity to amyloid beta protein in older humans and patients with Alzheimer disease.

    Science.gov (United States)

    Monsonego, Alon; Zota, Victor; Karni, Arnon; Krieger, Jeffery I; Bar-Or, Amit; Bitan, Gal; Budson, Andrew E; Sperling, Reisa; Selkoe, Dennis J; Weiner, Howard L

    2003-08-01

    Alzheimer disease (AD) is characterized by the progressive deposition of the 42-residue amyloid beta protein (Abeta) in brain regions serving memory and cognition. In animal models of AD, immunization with Abeta results in the clearance of Abeta deposits from the brain. However, a trial of vaccination with synthetic human Abeta1-42 in AD resulted in the development of meningoencephalitis in some patients. We measured cellular immune responses to Abeta in middle-aged and elderly healthy subjects and in patients with AD. A significantly higher proportion of healthy elderly subjects and patients with AD had strong Abeta-reactive T cell responses than occurred in middle-aged adults. The immunodominant Abeta epitopes in humans resided in amino acids 16-33. Epitope mapping enabled the identification of MHC/T cell receptor (TCR) contact residues. The occurrence of intrinsic T cell reactivity to the self-antigen Abeta in humans has implications for the design of Abeta vaccines, may itself be linked to AD susceptibility and course, and appears to be associated with the aging process. PMID:12897209

  5. Design of LVFFARK and LVFFARK-functionalized nanoparticles for inhibiting amyloid ?-protein fibrillation and cytotoxicity.

    Science.gov (United States)

    Xiong, Neng; Dong, Xiao-Yan; Zheng, Jie; Liu, Fu-Feng; Sun, Yan

    2015-03-18

    Aggregation of amyloid ?-protein (A?) into amyloid oligomers and fibrils is pathologically linked to Alzheimer's disease (AD). Hence, the inhibition of A? aggregation is essential for the prevention and treatment of AD, but the development of potent agents capable of inhibiting A? fibrillogenesis has posed significant challenges. Herein, we designed Ac-LVFFARK-NH2 (LK7) by incorporating two positively charged residues, R and K, into the central hydrophobic fragment of A?17-21 (LVFFA) and examined its inhibitory effect on A?42 aggregation and cytotoxicity by extensive physical, biophysical, and biological analyses. LK7 was observed to inhibit A?42 fibrillogenesis in a dose-dependent manner, but its strong self-assembly characteristic also resulted in high cytotoxicity. In order to prevent the cytotoxicity that resulted from the self-assembly of LK7, the peptide was then conjugated to the surface of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) to fabricate a nanosized inhibitor, LK7@PLGA-NPs. It was found that LK7@PLGA-NPs had little cytotoxicity because the self-assembly of the LK7 conjugated on the NPs was completely inhibited. Moreover, the NPs-based inhibitor showed remarkable inhibitory capability against A?42 aggregation and significantly alleviated its cytotoxicity at a low LK7@PLGA-NPs concentration of 20 ?g/mL. At the same peptide concentration, free LK7 showed little inhibitory effect. It is considered that several synergetic effects contributed to the strong inhibitory ability of LK7@PLGA-NPs, including the enhanced interactions between A?42 and LK7@PLGA-NPs brought on by inhibiting LK7 self-assembly, restricting conformational changes of A?42, and thus redirecting A?42 aggregation into unstructured, off-pathway aggregates. The working mechanisms of the inhibitory effects of LK7 and LK7@PLGA-NPs on A?42 aggregation were proposed based on experimental observations. This work provides new insights into the design and development of potent NPs-based inhibitors against A? aggregation and cytotoxicity. PMID:25700145

  6. The Functions of the Amyloid Precursor Protein Gene and Its Derivative Peptides: I Molecular Biology and Metabolic Processing

    OpenAIRE

    Atkins, Emily R.; Panegyres, Peter K

    2011-01-01

    The amyloid precursor protein gene (APP) and its derivative peptides have important functions in the central nervous system. APP and A? fulfil criteria as neuractive peptides: presence, release and identity of action. A? is a peptide of 1 - 43 amino acids in length, derived from APP and the major component of the core of neuritic plaques found in Alzheimer’s disease. Analysis of the cDNA of A? revealed its origins from the larger precursor protein. There are at least four types of mRNA g...

  7. Self-Assembly of a Nine-Residue Amyloid-Forming Peptide Fragment of SARS Corona Virus E-Protein: Mechanism of Self Aggregation and Amyloid-Inhibition of hIAPP.

    Science.gov (United States)

    Ghosh, Anirban; Pithadia, Amit S; Bhat, Jyotsna; Bera, Supriyo; Midya, Anupam; Fierke, Carol A; Ramamoorthy, Ayyalusamy; Bhunia, Anirban

    2015-04-01

    Molecular self-assembly, a phenomenon widely observed in nature, has been exploited through organic molecules, proteins, DNA, and peptides to study complex biological systems. These self-assembly systems may also be used in understanding the molecular and structural biology which can inspire the design and synthesis of increasingly complex biomaterials. Specifically, use of these building blocks to investigate protein folding and misfolding has been of particular value since it can provide tremendous insights into peptide aggregation related to a variety of protein misfolding diseases, or amyloid diseases (e.g., Alzheimer's disease, Parkinson's disease, type-II diabetes). Herein, the self-assembly of TK9, a nine-residue peptide of the extra membrane C-terminal tail of the SARS corona virus envelope, and its variants were characterized through biophysical, spectroscopic, and simulated studies, and it was confirmed that the structure of these peptides influences their aggregation propensity, hence, mimicking amyloid proteins. TK9, which forms a beta-sheet rich fibril, contains a key sequence motif that may be critical for beta-sheet formation, thus making it an interesting system to study amyloid fibrillation. TK9 aggregates were further examined through simulations to evaluate the possible intra- and interpeptide interactions at the molecular level. These self-assembly peptides can also serve as amyloid inhibitors through hydrophobic and electrophilic recognition interactions. Our results show that TK9 inhibits the fibrillation of hIAPP, a 37 amino acid peptide implicated in the pathology of type-II diabetes. Thus, biophysical and NMR experimental results have revealed a molecular level understanding of peptide folding events, as well as the inhibition of amyloid-protein aggregation are reported. PMID:25785896

  8. The index herd with PMWS in Sweden: Presence of serum amyloid A, circovirus 2 viral load and antibody levels in healthy and PMWS-affected pigs

    Directory of Open Access Journals (Sweden)

    Allan Gordon

    2009-03-01

    Full Text Available Abstract Background Postweaning Multisystemic Wasting Syndrome (PMWS is an emerging disease in pigs of multifactorial origin, but associated to porcine circovirus type 2 (PCV2 infection. PMWS was first diagnosed in Sweden at a progeny test station that received pigs aged five weeks from 19 different nucleus herds on the day after weaning. The objective of this study was to examine, for the first time in an index outbreak of PMWS, the relationship between PCV2 virus, antibodies to PCV2 and serum amyloid a (SAA in sequentially collected serum samples from pigs with and without signs of PMWS. Methods Forty pigs of the last batch that entered the station at a mean age of 37.5 days were monitored for signs of PMWS during the first 55 days after arrival. Serum was collected on six occasions and analysed for presence of PCV2 DNA and antibodies to PCV2, as well as for levels of SAA. Results Four of the pigs (10% were concluded to have developed PMWS, with necropsy confirmation in three of them. These pigs displayed low levels of maternal antibodies to PCV2, more than 107 PCV2 viral DNA copies per ml serum and failed to mount a serological response to the virus. Starting between day 23 and 34 after arrival, an increase in PCV2 viral load was seen in all pigs, but PCV2 did not induce any SAA-response. Pigs that remained healthy seroconverted to PCV2 as the viral load was increased, regardless of initially having low or high levels of PCV2-antibodies. Conclusion In this index case of PMWS in Sweden, pigs affected by PMWS were not able to mount a relevant serum antibody response which contributed to the disease progression. The maximal PCV2 virus load was significantly higher and was also detected at an earlier stage in PMWS-affected pigs than in healthy pigs. However, a viral load above 107 PCV2 DNA copies per ml serum was also recorded in 18 out of 34 pigs without any clinical signs of PMWS, suggesting that these pigs were able to initiate a protective immune response to PCV2.

  9. Competitive protein adsorption to polymer surface from human serum

    DEFF Research Database (Denmark)

    Holmberg, Maria; Jensen, Karin Bagger Stibius

    2008-01-01

    Surface modification by "soft" plasma polymerisation to obtain a hydrophilic and non-fouling polymer surface has been validated using radioactive labelling. Adsorption to unmodified and modified polymer surfaces, from both single protein and human serum solutions, has been investigated. By using different radioisotopes, albumin and Immunoglobulin G (IgG) adsorption has been monitored simultaneously during competitive adsorption processes, which to our knowledge has not been reported in the literature before. Results show that albumin and IgG adsorption is dependent on adsorption time and on the presence and concentration of other proteins in bulk solutions during adsorption. Generally, lower albumin and IgG adsorption was observed on the modified and more hydrophilic polymer surfaces, but otherwise the modified and unmodified polymer surfaces showed the same adsorption characteristics.

  10. Neural stem cells isolated from amyloid precursor protein-mutated mice for drug discovery

    Directory of Open Access Journals (Sweden)

    Vito Antonio Baldassarro

    2013-01-01

    Full Text Available AIM: To develop an in vitro model based on neural stem cells derived from transgenic animals, to be used in the study of pathological mechanisms of Alzheimer’s disease and for testing new molecules. METHODS: Neural stem cells (NSCs were isolated from the subventricular zone of Wild type (Wt and Tg2576 mice. Primary and secondary neurosphere generation was studied, analysing population doubling and the cell yield per animal. Secondary neurospheres were dissociated and plated on MCM Gel Cultrex 2D and after 6 d in vitro (DIVs in mitogen withdrawal conditions, spontaneous differentiation was studied using specific neural markers (MAP2 and TuJ-1 for neurons, GFAP for astroglial cells and CNPase for oligodendrocytes. Gene expression pathways were analysed in secondary neurospheres, using the QIAGEN PCR array for neurogenesis, comparing the Tg2576 derived cell expression with the Wt cells. Proteins encoded by the altered genes were clustered using STRING web software. RESULTS: As revealed by 6E10 positive staining, all Tg2576 derived cells retain the expression of the human transgenic Amyloid Precursor Protein. Tg2576 derived primary neurospheres show a decrease in population doubling. Morphological analysis of differentiated NSCs reveals a decrease in MAP2- and an increase in GFAP-positive cells in Tg2576 derived cells. Analysing the branching of TuJ-1 positive cells, a clear decrease in neurite number and length is observed in Tg2576 cells. The gene expression neurogenesis pathway revealed 11 altered genes in Tg2576 NSCs compared to Wt. CONCLUSION: Tg2576 NSCs represent an appropriate AD in vitro model resembling some cellular alterations observed in vivo, both as stem and differentiated cells.

  11. Development of transgenic rats producing human ?-amyloid precursor protein as a model for Alzheimer's disease: Transgene and endogenous APP genes are regulated tissue-specifically

    Directory of Open Access Journals (Sweden)

    Chan Anthony WS

    2008-02-01

    Full Text Available Abstract Background Alzheimer's disease (AD is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD. Some instances of FAD have been linked to mutations in the ?-amyloid protein precursor (APP. Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated. Results Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (A?40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum A?42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP. Conclusion This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue-specific expression in the two transgenic rat lines and in wild-type rats contradicts our current understanding of APP gene regulation. Determination of the elements that are responsible for tissue-specific expression of APP may enable new treatment options for AD.

  12. Study on the Elution of the Adsorbed Serum Proteins on Biphasic Calcium Phosphate Ceramics

    Directory of Open Access Journals (Sweden)

    WANG Jing, CHEN Ying, ZHU Xiang-Dong, FAN Yu-Jiang, ZHANG Xing-Dong

    2013-10-01

    Full Text Available Two types of porous BCP ceramics were fabricated, using human serum as the model of protein solution. The effect of the different eluants and porous structure on the elution of the adsorbed serum proteins on BCP ceramics were investigated. The results showed that 400 mmol/L Na3PO4 solution had higher elution efficiency for the adsorbed serum proteins on BCP ceramics than 2 mol/L NaCl or 2% sodium dodecycl sufonate (SDS solution. Moreover, the porous structure of BCP ceramics had a strong impact on the elution of the adsorbed serum proteins. The microporous structure could promote the adsorption of serum proteins on BCP ceramics and slowed down their release rate, but the 3D interconnected macropores were favorable for the elution and release of serum proteins from the BCP surface.

  13. Amyloid misfolding, aggregation, and the early onset of protein deposition diseases: insights from AFM experiments and computational analyses

    Directory of Open Access Journals (Sweden)

    Yuri L. Lyubchenko

    2015-05-01

    Full Text Available The development of Alzheimer’s disease is believed to be caused by the assembly of amyloid ? proteins into aggregates and the formation of extracellular senile plaques. Similar models suggest that structural misfolding and aggregation of proteins are associated with the early onset of diseases such as Parkinson’s, Huntington’s, and other protein deposition diseases. Initially, the aggregates were structurally characterized by traditional techniques such as x-ray crystallography, NMR, electron microscopy, and AFM. However, data regarding the structures formed during the early stages of the aggregation process were unknown. Experimental models of protein deposition diseases have demonstrated that the small oligomeric species have significant neurotoxicity. This highlights the urgent need to discover the properties of these species, to enable the development of efficient diagnostic and therapeutic strategies. The oligomers exist transiently, making it impossible to use traditional structural techniques to study their characteristics. The recent implementation of single-molecule imaging and probing techniques that are capable of probing transient states have enabled the properties of these oligomers to be characterized. Additionally, powerful computational techniques capable of structurally analyzing oligomers at the atomic level advanced our understanding of the amyloid aggregation problem. This review outlines the progress in AFM experimental studies and computational analyses with a primary focus on understanding the very first stage of the aggregation process. Experimental approaches can aid in the development of novel sensitive diagnostic and preventive strategies for protein deposition diseases, and several examples of these approaches will be discussed.

  14. Amyloid precursor protein processing and A?42 deposition in a transgenic mouse model of?Alzheimer?disease

    Science.gov (United States)

    Johnson-Wood, K.; Lee, M.; Motter, R.; Hu, K.; Gordon, G.; Barbour, R.; Khan, K.; Gordon, M.; Tan, H.; Games, D.; Lieberburg, I.; Schenk, D.; Seubert, P.; McConlogue, L.

    1997-01-01

    The PDAPP transgenic mouse, which overexpresses human amyloid precursor protein (APP717V?F), has been shown to develop much of the pathology associated with Alzheimer disease. In this report, levels of APP and its amyloidogenic metabolites were measured in brain regions of transgenic mice between 4 and 18 months of age. While absolute levels of APP expression likely contribute to the rate of amyloid ?-peptide (A?) deposition, regionally specific factors also seem important, as homozygotic mice express APP levels in pathologically unaffected regions in excess of that measured in certain amyloid plaque-prone regions of heterozygotic mice. Regional levels of APP and APP-? were nearly constant at all ages, while A? levels dramatically and predictably increased in brain regions undergoing histochemically confirmed amyloidosis, most notably in the cortex and hippocampus. In hippocampus, A? concentrations increase 17-fold between the ages of 4 and 8 months, and by 18 months of age are over 500-fold that at 4 months, reaching an average level in excess of 20 nmol of A? per g of tissue. A?1–42 constitutes the vast majority of the depositing A? species. The similarities observed between the PDAPP mouse and human Alzheimer disease with regard to A?42 deposition occurring in a temporally and regionally specific fashion further validate the use of the model in understanding processes related to the disease. PMID:9037091

  15. Transcranial laser therapy alters amyloid precursor protein processing and improves mitochondrial function in a mouse model of Alzheimer's disease

    Science.gov (United States)

    McCarthy, Thomas; Yu, Jin; El-Amouri, Salim; Gattoni-Celli, Sebastiano; Richieri, Steve; De Taboada, Luis; Streeter, Jackson; Kindy, Mark S.

    2011-03-01

    Transcranial laser therapy (TLT) using a near-infrared energy laser system was tested in the 2x Tg amyloid precursor protein (APP) mouse model of Alzheimer's Disease (AD). TLT was administered 3 times/week at escalating doses, starting at 3 months of age, and was compared to a control group which received no laser treatment. Treatment sessions were continued for a total of six months. The brains were examined for amyloid plaque burden, A? peptides (A?1-40 and A?1-42 ), APP cleavage products (sAPP?, CTF?) and mitochondrial activity. Administration of TLT was associated with a significant, dose-dependent reduction in amyloid load as indicated by the numbers of A? plaques. Levels of A?1-40 and A?1-42 levels were likewise reduced in a dose-dependent fashion. All TLT doses produced an increase in brain sAPP? and a decrease in CTF? levels consistent with an increase in ?-secretase activity and a decrease in ?-secretase activity. In addition, TLT increased ATP levels and oxygen utilization in treated animals suggesting improved mitochondrial function. These studies suggest that TLT is a potential candidate for treatment of AD.

  16. Beta-amyloid precursor protein of Alzheimer disease occurs as 110- to 135-kilodalton membrane-associated proteins in neural and nonneural tissues.

    OpenAIRE

    Selkoe, D. J.; Podlisny, M. B.; Joachim, C. L.; Vickers, E A; Lee, G.; Fritz, L C; Oltersdorf, T

    1988-01-01

    Progressive cerebral deposition of extracellular filaments composed of the beta-amyloid protein (beta AP) is a constant feature of Alzheimer disease (AD). Since the gene on chromosome 21 encoding the beta AP precursor (beta APP) is not known to be altered in AD, transcriptional or posttranslational changes may underlie accelerated beta AP deposition. Using two antibodies to the predicted carboxyl terminus of beta APP, we have identified the native beta APP in brain and nonneural human tissues...

  17. Influence of chlorpyrifos oxon on the development and progression of Alzheimer's disease in amyloid precursor protein transgenic mice

    Directory of Open Access Journals (Sweden)

    Jin Yu

    2015-03-01

    Full Text Available Aim: Alzheimer's disease (AD is a devastating neurological disorder and the most common form of dementia. Until date, the cause of AD eludes us, but a number of hypotheses have been put forward to try and understand the mechanisms involved. A series of studies have indicated that environmental factors, such as pesticides, heavy metals, and others can contribute to the development and progression of AD. Based on these data, we determined the impact of pesticides (chlorpyrifos oxon [CPO] on AD-like pathogenesis in amyloid precursor protein (APP transgenic mice. Methods: APP mice were treated at various times with low-dose CPO (1 mg/kg/day, in utero (3-week of gestation, during lactation (3-week, or as young adults (continuous dosing. Results: Exposure to CPO at all times enhanced neuro-inflammation and exacerbated oxidative stress in the brain prior to amyloid deposition. CPO-treated APP mice showed a decrease in memory and learning compared with untreated APP mice; furthermore, analyses of brain tissue sections and extracts showed an increase in Ab levels and C-terminal fragment-b levels, a decrease in soluble APPa (sAPPa levels, and an increase in plaque load. In addition, CPO-treated APP transgenic mice showed a significant decrease in neurotrophic factor levels (nerve growth factor, brain-derived neurotrophic factor, and neurotrophin-3 compared to vehicle-treated APP transgenic animals. Treatment with galantamine attenuated the effects of CPO by reducing amyloid b levels and amyloid load. Conclusion: CPO accelerated and exacerbated the disease development and progression in the APP mice suggesting that pesticides may play a significant role in the pathogenesis of AD.

  18. PB1-F2 Influenza A Virus Protein Adopts a ?-Sheet Conformation and Forms Amyloid Fibers in Membrane Environments

    Science.gov (United States)

    Chevalier, Christophe; Al Bazzal, Ali; Vidic, Jasmina; Février, Vincent; Bourdieu, Christiane; Bouguyon, Edwige; Le Goffic, Ronan; Vautherot, Jean-François; Bernard, Julie; Moudjou, Mohammed; Noinville, Sylvie; Chich, Jean-François; Da Costa, Bruno; Rezaei, Human; Delmas, Bernard

    2010-01-01

    The influenza A virus PB1-F2 protein, encoded by an alternative reading frame in the PB1 polymerase gene, displays a high sequence polymorphism and is reported to contribute to viral pathogenesis in a sequence-specific manner. To gain insights into the functions of PB1-F2, the molecular structure of several PB1-F2 variants produced in Escherichia coli was investigated in different environments. Circular dichroism spectroscopy shows that all variants have a random coil secondary structure in aqueous solution. When incubated in trifluoroethanol polar solvent, all PB1-F2 variants adopt an ?-helix-rich structure, whereas incubated in acetonitrile, a solvent of medium polarity mimicking the membrane environment, they display ?-sheet secondary structures. Incubated with asolectin liposomes and SDS micelles, PB1-F2 variants also acquire a ?-sheet structure. Dynamic light scattering revealed that the presence of ?-sheets is correlated with an oligomerization/aggregation of PB1-F2. Electron microscopy showed that PB1-F2 forms amorphous aggregates in acetonitrile. In contrast, at low concentrations of SDS, PB1-F2 variants exhibited various abilities to form fibers that were evidenced as amyloid fibers in a thioflavin T assay. Using a recombinant virus and its PB1-F2 knock-out mutant, we show that PB1-F2 also forms amyloid structures in infected cells. Functional membrane permeabilization assays revealed that the PB1-F2 variants can perforate membranes at nanomolar concentrations but with activities found to be sequence-dependent and not obviously correlated with their differential ability to form amyloid fibers. All of these observations suggest that PB1-F2 could be involved in physiological processes through different pathways, permeabilization of cellular membranes, and amyloid fiber formation. PMID:20172856

  19. Pathogenic microbial amyloids: Their function and the host response

    OpenAIRE

    Garcia, MC; Lipke, PN; Klotz, SA

    2013-01-01

    Functional microbial amyloids are ubiquitous in nature and some contribute to the pathogenesis of infectious diseases. Three pathogenic microbial amyloids are compared and their contribution to the disease process explained. The recent demonstration and visualization of fungal amyloid in human invasive candidiasis is discussed. Moreover, the binding of host serum amyloid P component to Candida functional amyloid in invasive human disease is presented in light of its possible role of masking f...

  20. ALTERATIONS IN TOTAL PROTEIN CONCENTRATION, SERUM PROTEIN FRACTIONS AND ALBUMIN/GLOBULIN RATIO IN HEALTHY RABBITS

    OpenAIRE

    Nuzhat Sultana; Rahila Najam

    2013-01-01

    This study assessed the effect of oral administration of Aloe vera and was to evaluate total serum protein, albumin and globulin concentrations as well as albumin / globulin (A / G) ratio. Twenty rabbits weighing 1000 – 1800 g were divided into 2 groups. Each group consisted of ten animals. One served as control and other group served as experimental group. Results show that animals after 07, 15 and 30 days dosing of Aloe vera showed highly significant decrease in total protein and globulin a...

  1. [Amyloid ? protein suppresses hippocampal theta rhythm and induces behavioral disinhibition and spatial memory deficit in rats].

    Science.gov (United States)

    Yue, Xing-Hua; Liu, Xiao-Jie; Wu, Mei-Na; Chen, Jin-Yuan; Qi, Jin-Shun

    2014-04-25

    Hippocampal neuronal network oscillation is closely related to the memory, anxiety and behavioral inhibition of mammalian. The cognitive decline and behavioral disinhibition in the patients with Alzheimer's disease (AD) may be relevant to amyloid ? protein (A?)-induced impairment in hippocampal neuronal cooperative activity. However, it is not well known whether intrahippocampal injection of A? could induce behavioral disinhibition and neuronal network disorder, as well as cognition decline in animals. In the present study, we observed the effects of intracerebral injection of A?(1-42) on the spatial memory and behavioral inhibition of rats by using Morris water maze and elevated plus-maze tests. Further, we analyzed hippocampal theta rhythm by recording hippocampal local field potential. The results showed that: (1) bilateral hippocampal injection of A?(1-42) reduced the anxious behavior of rats, with a significant behavioral disinhibition in the elevated plus-maze test, representing as an increase in the mean entering times and mean residence time in the open arm; (2) A?(1-42) injection resulted in a significant impairment of spatial memory in rats, with significantly increased mean escape latencies in hidden platform test; (3) A?(1-42) disrupted the induction of theta rhythm induced by tail pinch, with a significant reduction in the peak power, not the peak power frequency of the theta rhythm. These experimental results indicate that intrahippocampal injection of A?(1-42) can induce behavioral disinhibition and theta rhythm suppression, as well as spatial memory impairment in rats, which suggests that the cognition deficits and behavior impairments in AD are probably associated with the A?-induced disruption of hippocampal theta rhythm and consequent down-regulation of synaptic plasticity. PMID:24777399

  2. Platelet amyloid precursor protein isoform expression in Alzheimer's disease: evidence for peripheral marker.

    Science.gov (United States)

    Vignini, A; Sartini, D; Morganti, S; Nanetti, L; Luzzi, S; Provinciali, L; Mazzanti, L; Emanuelli, M

    2011-01-01

    Alzheimer's disease (AD) is a chronic neurodegenerative disorder characterized by a progressive cognitive and memory decline. Among peripheral markers of AD, great interest has been focused on the amyloid precursor protein (APP). In this regard, platelets represent an important peripheral source of APP since it has been demonstrated that the three major isoforms, that are constituted of 770, 751 and 695 aa residues, are inserted in the membrane of resting platelets. APP 751 and APP 770 contain a Kunitz-type serine protease inhibitor domain (APP KPI) and APP 695 lacks this domain. To address this issue, we first examined the platelet APP isoform mRNAs prospectively as biomarker for the diagnosis of AD by means of real-time quantitative PCR, and then evaluated the correlation between APP mRNA expression levels and cognitive impairment of enrolled subjects. Differential gene expression measurements in the AD patient group (n=18) revealed a significant up-regulation of APP TOT (1.52-fold), APP KPI (1.32-fold), APP 770 (1.33-fold) and APP 751 (1.26-fold) compared to controls (n=22). Moreover, a statistically significant positive correlation was found between APP mRNA levels (TOT, KPI, 770 and 751) and cognitive impairment. Since AD definitive diagnosis still relies on pathological evaluation at autopsy, the present results are consistent with the hypothesis that platelet APP could be considered a potential reliable peripheral marker for studying AD and could contribute to define a signature for the presence of AD pathology. PMID:21658330

  3. Organotypic vibrosections from whole brain adult Alzheimer mice (overexpressing amyloid-precursor-protein with the Swedish-Dutch-Iowa mutations) as a model to study clearance of beta-amyloid plaques

    Science.gov (United States)

    Humpel, Christian

    2015-01-01

    Alzheimer's disease is a severe neurodegenerative disorder of the brain, pathologically characterized by extracellular beta-amyloid plaques, intraneuronal Tau inclusions, inflammation, reactive glial cells, vascular pathology and neuronal cell death. The degradation and clearance of beta-amyloid plaques is an interesting therapeutic approach, and the proteases neprilysin (NEP), insulysin and matrix metalloproteinases (MMP) are of particular interest. The aim of this project was to establish and characterize a simple in vitro model to study the degrading effects of these proteases. Organoytpic brain vibrosections (120 ?m thick) were sectioned from adult (9 month old) wildtype and transgenic mice (expressing amyloid precursor protein (APP) harboring the Swedish K670N/M671L, Dutch E693Q, and Iowa D694N mutations; APP_SDI) and cultured for 2 weeks. Plaques were stained by immunohistochemistry for beta-amyloid and Thioflavin S. Our data show that plaques were evident in 2 week old cultures from 9 month old transgenic mice. These plaques were surrounded by reactive GFAP+ astroglia and Iba1+ microglia. Incubation of fresh slices for 2 weeks with 1–0.1–0.01 ?g/ml of NEP, insulysin, MMP-2, or MMP-9 showed that NEP, insulysin, and MMP-9 markedly degraded beta-amyloid plaques but only at the highest concentration. Our data provide for the first time a potent and powerful living brain vibrosection model containing a high number of plaques, which allows to rapidly and simply study the degradation and clearance of beta-amyloid plaques in vitro. PMID:25914642

  4. Serum transferrin levels in children with protein-energy malnutrition

    Directory of Open Access Journals (Sweden)

    Selime Aydo?du

    2013-03-01

    Full Text Available Objective: Although the diagnosis of patients with severemalnutrition is easy, it is very difficult to recognize patientswith mild and moderate malnutrition. A variety of methodsattempts to develop for early diagnosis of these cases.In this study, we evaluated the serum transferrin and albuminlevels in children with mild, moderate and severeprotein-energy malnutrition (PEM.Materials and methods: Children admitted to our policlinic,aged between 3 and 25 months, 45 subjects withPEM and 39 healthy subjects (control group were evaluated.According to the Gomez, Waterlow and Kanawatisubjects with PEM were divided in 3 subgroups mild,moderate and severe PEM. Anthropometric measurementsand biochemical results of 4 groups were compared.Results: For albumin levels in mild to moderate PEMgroups, 37.7% sensitivity, and 28.5% specificity, positivepredictive value 54%; negative predictive value 16.6%was found. For severe PEM sensitivity, specificity, positivepredictive value and negative predictive value were71%, 62.5%, 45%, and 83.3% respectively.With respect to the levels of transferrin, a significant differencewas found between mild PEM-control and moderatePEM-control groups (p0.05.Conclusion: Our study results showed that albumin isa weak indicator in mild-moderate PEM. In these cases,serum transferrin level reduces before decreasing of albuminlevel, thus it may be an early sensitive finding thatcan be used as a sensitive parameter in the diagnosis ofearly stages of malnutrition.Key words: Protein energy malnutrition, children, albumin,transferrin

  5. SorLA is a molecular link for retromer-dependent sorting of the Amyloid precursor protein

    DEFF Research Database (Denmark)

    Fjorback, Anja W; Andersen, Olav Michael

    2012-01-01

    Deficiency in the retromer sorting pathway is known to be associated with the onset of Alzheimer disease (AD), and has been suggested to involve regulation of Amyloid precursor protein (APP) trafficking. Absence of the APP sorting receptor sorLA is also associated to AD, as amyloidogenic processing of APP is increased due to missorting. Reduced activity of either retromer or sorLA thus both lead to enhanced amyloidogenic APP processing, and these pathways are therefore important factors for understanding the development of AD. It is therefore key to outline the neuronal APP trafficking in order to determine the mechanisms that influence AD onset.

  6. Effect of Apolipoprotein E Allele ?4 on the Initial Phase of Amyloid ?-Protein Accumulation in the Human Brain

    OpenAIRE

    Morishima-kawashima, Maho; Oshima, Noriko; Ogata, Hiromitsu; Yamaguchi, Haruyasu; Yoshimura, Masahiro; Sugihara, Shiro; Ihara, Yasuo

    2000-01-01

    Deposition of amyloid ?-protein (A?), a hallmark of Alzheimer’s disease, occurs to some extent in the brains of most elderly individuals. We sought to learn when A? deposition begins and how deposition is affected by apolipoprotein E allele ?4, a strong risk factor for late-onset Alzheimer’s disease. Using an improved extraction protocol and specific enzyme-linked immunosorbent assay, we quantified the levels of A?40 and A?42 in the insoluble fractions of brains from 105 autopsy cas...

  7. In vivo imaging reveals sigmoidal growth kinetic of ?-amyloid plaques

    OpenAIRE

    Burgold, Steffen; Filser, Severin; Dorostkar, Mario M.; Schmidt, Boris; Herms, Jochen

    2014-01-01

    A major neuropathological hallmark of Alzheimer’s disease is the deposition of amyloid plaques in the brains of affected individuals. Amyloid plaques mainly consist of fibrillar ?-amyloid, which is a cleavage product of the amyloid precursor protein. The amyloid-cascade-hypothesis postulates A? accumulation as the central event in initiating a toxic cascade leading to Alzheimer’s disease pathology and, ultimately, loss of cognitive function. We studied the kinetics of ?-amyloid deposit...

  8. Modeling amyloids in bacteria

    OpenAIRE

    Villar-Piqué Anna; Ventura Salvador

    2012-01-01

    Abstract An increasing number of proteins are being shown to assemble into amyloid structures, self-seeding fibrillar aggregates that may lead to pathological states or play essential biological functions in organisms. Bacterial cell factories have raised as privileged model systems to understand the mechanisms behind amyloid assembly and the cellular fitness cost associated to the formation of these aggregates. In the near future, these bacterial systems will allow implementing high-throughp...

  9. Lower Total Serum Protein, Albumin and Zinc in Depression in an Iranian Population

    OpenAIRE

    Salimi, S. (PhD); M. Kianpoor; M.R. Abassi; M. Abdani; E.S. Moghaddam

    2008-01-01

    We studied associations between lowered serum zinc, total protein and albumin with depression and their correlations. In the present study serum zinc, albumin and total protein were determined in 144 depressed patients and 161 age-sex-matched healthy controls. The serum zinc concentration was significantly lower in depressive patients (64.94±13.9 ?g dL-1) than in normal controls (69.67±11.29 ?g dL-1) (p = 0.001). Albumin and total serum protein were significantly lower in depressed pat...

  10. Chemokines, macrophage inflammatory protein-2 and stromal cell-derived factor-1?, suppress amyloid ?-induced neurotoxicity

    International Nuclear Information System (INIS)

    Alzheimer's disease (AD) is characterized by a progressive cognitive decline and accumulation of neurotoxic oligomeric peptides amyloid-? (A?). Although the molecular events are not entirely known, it has become evident that inflammation, environmental and other risk factors may play a causal, disruptive and/or protective role in the development of AD. The present study investigated the ability of the chemokines, macrophage inflammatory protein-2 (MIP-2) and stromal cell-derived factor-1? (SDF-1?), the respective ligands for chemokine receptors CXCR2 and CXCR4, to suppress A?-induced neurotoxicity in vitro and in vivo. Pretreatment with MIP-2 or SDF-1? significantly protected neurons from A?-induced dendritic regression and apoptosis in vitro through activation of Akt, ERK1/2 and maintenance of metalloproteinase ADAM17 especially with SDF-1?. Intra-cerebroventricular (ICV) injection of A? led to reduction in dendritic length and spine density of pyramidal neurons in the CA1 area of the hippocampus and increased oxidative damage 24 h following the exposure. The A?-induced morphometric changes of neurons and increase in biomarkers of oxidative damage, F2-isoprostanes, were significantly inhibited by pretreatment with the chemokines MIP-2 or SDF-1?. Additionally, MIP-2 or SDF-1? was able to suppress the aberrant mislocalization of p21-activated kinase (PAK), one of the proteins involved in the maintenance of dendritic spines. Furthermore, MIP-2 also protected neurons against A? neurotoxicity in CXCR2?/? mice, potentially through observed up regulation of CXCR1 mRNA. Understanding the neuroprotective potential of chemokines is crucial in defining the role for their employment during the early stages of neurodegeneration. -- Research highlights: ? Neuroprotective ability of the chemokines MIP2 and CXCL12 against A? toxicity. ? MIP-2 or CXCL12 prevented dendritic regression and apoptosis in vitro. ? Neuroprotection through activation of Akt, ERK1/2 and maintenance of ADAM17. ? Neuroprotection of hippocampal pyramidal neurons in vivo by MIP-2 or CXCL12. ? MIP-2 or CXCL12 prevent elevation of F2-Isoprostanes against A? treatment.

  11. Serum amyloid A stimulates matrix-metalloproteinase-9 upregulation via formyl peptide receptor like-1-mediated signaling in human monocytic cells

    International Nuclear Information System (INIS)

    In the present study, we found that serum amyloid A (SAA) stimulated matrix-metalloproteinase-9 (MMP-9) upregulation at the transcription and translational levels in THP-1 cells. SAA stimulated the activation of nuclear factor ?B (NF-?B), which was required for the MMP-9 upregulation by SAA. The signaling events induced by SAA included the activation of ERK and intracellular calcium rise, which were found to be required for MMP-9 upregulation. Formyl peptide receptor like 1 (FPRL1) was found to be involved in the upregulation of MMP-9 by SAA. Among several FPRL1 agonists, including Trp-Lys-Tyr-Met-Val-D-Met (WKYMVm), SAA selectively stimulated MMP-9 upregulation. With respect to the molecular mechanisms involved in the differential action of SAA and WKYMVm, we found that SAA could not competitively inhibit the binding of 125I-labeled WKYMVm to FPRL1. Taken together, we suggest that SAA plays a role in the modulation of inflammatory and immune responses via FPRL1, by inducing MMP-9 upregulation in human monocytic cells

  12. Swedish Alzheimer mutation induces mitochondrial dysfunction mediated by HSP60 mislocalization of amyloid precursor protein (APP) and beta-amyloid

    DEFF Research Database (Denmark)

    Walls, Ken Carlson; Coskun, Pinar

    2012-01-01

    Alzheimer disease (AD) is a complex disorder that involves numerous cellular and subcellular alterations including impairments in mitochondrial homeostasis. To better understand the role of mitochondrial dysfunction in the pathogenesis of AD, we analyzed brains from clinically well-characterized human subjects and from the 3xTg-AD mouse model of AD. We find A? and critical components of the ?-secretase complex, presenilin-1, -2, and nicastrin, accumulate in the mitochondria. We used a proteomics approach to identify binding partners and show that heat shock protein 60 (HSP60), a molecular chaperone localized to mitochondria and the plasma membrane, specifically associates with APP. We next generated stable neural cell lines expressing human wild-type or Swedish APP, and provide corroborating in vitro evidence that HSP60 mediates translocation of APP to the mitochondria. Viral-mediated shRNA knockdown of HSP60 attenuates APP and A? mislocalization to the mitochondria. Our findings identify a novel interaction between APP and HSP60, which accounts for its translocation to the mitochondria.

  13. Differential effects of interleukin-1? and S100B on amyloid precursor protein in rat retinal neurons

    Directory of Open Access Journals (Sweden)

    Peter JB Anderson

    2009-02-01

    Full Text Available Peter JB Anderson1, Helena R Watts1, Sheila Jen1, Stephen M Gentleman2, Juliet A Moncaster1, et al1Department of Cellular and Molecular Neuroscience and 2Department of Clinical Neuroscience, Division of Neuroscience and Mental Health, Imperial College London, Burlington Danes Building, Hammersmith Hospital, London, UKPurpose: Interleukin-1? (IL-1? and S100B calcium binding protein B (S100B have been implicated in the pathogenesis of Alzheimer’s disease. Both are present in and around senile plaques and have been shown to increase levels of amyloid precursor protein (APP mRNA in vitro. However, it is not known how either of these substances affects APP in vivo.Methods: We have studied the effects of IL-1? and S100B on the expression and processing of APP using a retinal-vitreal model. We have also investigated the effect of amyloid beta peptide (A? on APP in the same system and the regulation of S100B production by A? and IL-1? from retinal glial cells.Results: Retinal ganglion cells constitutively express APP. However, after intravitreal injection of IL-1? or A? there was a marked reduction in APP levels as detected by Western blotting and IL-1? produced a decrease in APP immunoreactivity (IR. Nissl staining showed that the integrity of the injected retinas was unchanged after injection. Two days after S100B injection, there was a small reduction in APP-IR but this was accompanied by the appearance of some intensely stained large ganglion cells and there was some up-regulation in APP holoprotein  levels on Western blot. Seven days post-S100? injection, these large, highly stained cells had increased in number throughout the retina. Injection of A? and IL-1? also caused an increase in S100B production within the retinal Müller glial cells.Conclusion: These results support the hypothesis that S100B (a glial-derived neurotrophic factor and IL-1? (a pro-infl ammatory cytokine can modulate the expression and processing of APP in vivo and so may contribute to the progression of Alzheimer’s disease.Keywords: Alzheimer’s disease, interleukin 1?, S100B, amyloid precursor protein, amyloid-?, retina

  14. Overcoming antigen masking of anti-amyloidbeta antibodies reveals breaking of B cell tolerance by virus-like particles in amyloidbeta immunized amyloid precursor protein transgenic mice

    Science.gov (United States)

    Li, Qingyou; Cao, Chuanhai; Chackerian, Bryce; Schiller, John; Gordon, Marcia; Ugen, Kenneth E; Morgan, Dave

    2004-01-01

    Background In prior work we detected reduced anti-A? antibody titers in A?-vaccinated transgenic mice expressing the human amyloid precursor protein (APP) compared to nontransgenic littermates. We investigated this observation further by vaccinating APP and nontransgenic mice with either the wild-type human A? peptide, an A? peptide containing the "Dutch Mutation", E22Q, or a wild-type A? peptide conjugated to papillomavirus virus-like particles (VLPs). Results Anti-A? antibody titers were lower in vaccinated APP than nontransgenic mice even when vaccinated with the highly immunogenic A? E22Q. One concern was that human A? derived from the APP transgene might mask anti-A? antibodies in APP mice. To test this possibility, we dissociated antigen-antibody complexes by incubation at low pH. The low pH incubation increased the anti-A? antibody titers 20–40 fold in APP mice but had no effect in sera from nontransgenic mice. However, even after dissociation, the anti-A? titers were still lower in transgenic mice vaccinated with wild-type A? or E22Q A? relative to non-transgenic mice. Importantly, the dissociated anti-A? titers were equivalent in nontransgenic and APP mice after VLP-based vaccination. Control experiments demonstrated that after acid-dissociation, the increased antibody titer did not cross react with bovine serum albumin nor alpha-synuclein, and addition of A? back to the dissociated serum blocked the increase in antibody titers. Conclusions Circulating human A? can interfere with ELISA assay measurements of anti-A? titers. The E22Q A? peptide vaccine is more immunogenic than the wild-type peptide. Unlike peptide vaccines, VLP-based vaccines against A? abrogate the effects of A? self-tolerance. PMID:15186505

  15. Retromer Binds the FANSHY Sorting Motif in SorLA to Regulate Amyloid Precursor Protein Sorting and Processing

    DEFF Research Database (Denmark)

    Fjorback, Anja W; Seaman, Matthew

    2012-01-01

    sorLA is a sorting receptor for amyloid precursor protein (APP) genetically linked to Alzheimer's disease (AD). Retromer, an adaptor complex in the endosome-to-Golgi retrieval pathway, has been implicated in APP transport because retromer deficiency leads to aberrant APP sorting and processing and levels of retromer proteins are altered in AD. Here we report that sorLA and retromer functionally interact in neurons to control trafficking and amyloidogenic processing of APP. We have identified a sequence (FANSHY) in the cytoplasmic domain of sorLA that is recognized by the VPS26 subunit of the retromer complex. Accordingly, we characterized the interaction between the retromer complex and sorLA and determined the role of retromer on sorLA-dependent sorting and processing of APP. Mutations in the VPS26 binding site resulted in receptor redistribution to the endosomal network, similar to the situation seen in cells with VPS26 knockdown. The sorLA mutant retained APP-binding activity but, as opposed to the wild-type receptor, misdirected APP into a distinct non-Golgi compartment, resulting in increased amyloid processing. In conclusion, our data provide a molecular link between reduced retromer expression and increased amyloidogenesis as seen in patients with sporadic AD.

  16. The Lepidopteran endoribonuclease-U domain protein P102 displays dramatically reduced enzymatic activity and forms functional amyloids.

    Science.gov (United States)

    Pascale, Mariarosa; Laurino, Simona; Vogel, Heiko; Grimaldi, Annalisa; Monné, Magnus; Riviello, Lea; Tettamanti, Gianluca; Falabella, Patrizia

    2014-11-01

    Hemocytes of Heliothis virescens (F.) (Lepidoptera, Noctuidae) larvae produce a protein, P102, with a putative endoribonuclease-U domain. In previous works we have shown that P102 is involved in Lepidopteran immune response by forming amyloid fibrils, which catalyze and localize melanin deposition around non-self intruders during encapsulation, preventing harmful systemic spreading. Here we demonstrate that P102 belongs to a new class of proteins that, at least in Lepidoptera, has a diminished endoribonuclease-U activity probably due to the lack of two out of five catalytically essential residues. We show that the P102 homolog from Trichoplusia ni (Lepidoptera, Noctuidae) displays catalytic site residues identical to P102, a residual endoribonuclease-U activity and the ability to form functional amyloids. On the basis of these results as well as sequence and structural analyses, we hypothesize that all the Lepidoptera endoribonuclease-U orthologs with catalytic site residues identical to P102 form a subfamily with similar function. PMID:25043263

  17. Evaluation of three high abundance protein depletion kits for umbilical cord serum proteomics

    Directory of Open Access Journals (Sweden)

    Nie Jing

    2011-05-01

    Full Text Available Abstract Background High abundance protein depletion is a major challenge in the study of serum/plasma proteomics. Prior to this study, most commercially available kits for depletion of highly abundant proteins had only been tested and evaluated in adult serum/plasma, while the depletion efficiency on umbilical cord serum/plasma had not been clarified. Structural differences between some adult and fetal proteins (such as albumin make it likely that depletion approaches for adult and umbilical cord serum/plasma will be variable. Therefore, the primary purposes of the present study are to investigate the efficiencies of several commonly-used commercial kits during high abundance protein depletion from umbilical cord serum and to determine which kit yields the most effective and reproducible results for further proteomics research on umbilical cord serum. Results The immunoaffinity based kits (PROTIA-Sigma and 5185-Agilent displayed higher depletion efficiency than the immobilized dye based kit (PROTBA-Sigma in umbilical cord serum samples. Both the PROTIA-Sigma and 5185-Agilent kit maintained high depletion efficiency when used three consecutive times. Depletion by the PROTIA-Sigma Kit improved 2DE gel quality by reducing smeared bands produced by the presence of high abundance proteins and increasing the intensity of other protein spots. During image analysis using the identical detection parameters, 411 ± 18 spots were detected in crude serum gels, while 757 ± 43 spots were detected in depleted serum gels. Eight spots unique to depleted serum gels were identified by MALDI- TOF/TOF MS, seven of which were low abundance proteins. Conclusions The immunoaffinity based kits exceeded the immobilized dye based kit in high abundance protein depletion of umbilical cord serum samples and dramatically improved 2DE gel quality for detection of trace biomarkers.

  18. Serum protein pattern during cow pregnancy: Acute-phase proteins increase in the peripartum period.

    OpenAIRE

    Cairoli, F.; Battocchio, M.; Veronesi, Mc; Brambilla, D.; Conserva, F.; Eberini, I.; Wait, R.; Gianazza, E.

    2006-01-01

    Serum collected in a time-course mode during the pregnancy of a group of heifers was analyzed by 2-DE under various experimental conditions to optimize resolution of all protein spots. Changes in the levels of some components were detected during the last phase of pregnancy and early postpartum. These included a decrease of alpha2-HS-glycoprotein, an increase of alpha1-antichymotrypsin and, with a much larger and more abrupt variation, of orosomucoid and haptoglobin. These findings associate ...

  19. Developmental Pattern of a Serum Binding Protein for Multiplication Stimulating Activity in the Rat

    OpenAIRE

    White, Robert M.; Nissley, S. Peter; Short, Patricia A.; Rechler, Matthew M.; Fennoy, Ilene

    1982-01-01

    The concentration of multiplication stimulating activity (MSA), an insulinlike growth factor (IGF), is high in fetal rat serum. We now report that MSA is exclusively associated wth an albumin-size binding protein in fetal rat serum; the growth hormone-dependent, gamma globulin-size binding protein, which predominates in the older animal, is absent from fetal rat serum. When 125I-MSA was incubated with fetal rat serum and then gel filtered on Sephadex G-200, specific radioactivity eluted in th...

  20. Prolactin-derived amyloid in the aging pituitary gland.

    OpenAIRE

    Westermark, P.; Eriksson, L.; Engstro?m, U.; Enestro?m, S.; Sletten, K.

    1997-01-01

    Small amyloid deposits occur commonly in different organs in association with aging. As in other amyloids, the fibrils in the age-associated forms are built up by specific proteins, unique to every histological type. The amyloid proteins that have been identified in localized amyloid of human endocrine organs have all been of polypeptide hormone nature, and these include calcitonin, islet amyloid polypeptide (amylin), and atrial natriuretic factor. In the present study, we add prolactin to th...

  1. Rhenium and technetium complexes that bind to amyloid-? plaques.

    Science.gov (United States)

    Hayne, David J; North, Andrea J; Fodero-Tavoletti, Michelle; White, Jonathan M; Hung, Lin W; Rigopoulos, Angela; McLean, Catriona A; Adlard, Paul A; Ackermann, Uwe; Tochon-Danguy, Henri; Villemagne, Victor L; Barnham, Kevin J; Donnelly, Paul S

    2015-03-21

    Alzheimer's disease is associated with the presence of insoluble protein deposits in the brain called amyloid plaques. The major constituent of these deposits is aggregated amyloid-? peptide. Technetium-99m complexes that bind to amyloid-? plaques could provide important diagnostic information on amyloid-? plaque burden using Single Photon Emission Computed Tomography (SPECT). Tridentate ligands with a stilbene functional group were used to form complexes with the fac-[M(I)(CO)3](+) (M = Re or (99m)Tc) core. The rhenium carbonyl complexes with tridentate co-ligands that included a stilbene functional group and a dimethylamino substituent bound to amyloid-? present in human frontal cortex brain tissue from subjects with Alzheimer's disease. This chemistry was extended to make the analogous [(99m)Tc(I)(CO)3](+) complexes and the complexes were sufficiently stable in human serum. Whilst the lipophilicity (log?D7.4) of the technetium complexes appeared ideally suited for penetration of the blood-brain barrier, preliminary biodistribution studies in an AD mouse model (APP/PS1) revealed relatively low brain uptake (0.24% ID g(-1) at 2 min post injection). PMID:25515141

  2. Binding between lead ions and the high-abundance serum proteins.

    Science.gov (United States)

    Guo, Ming; He, Ling; Strong, P J; Wang, Hailong

    2014-10-01

    The interaction between three of the most abundant bovine serum proteins (serum albumin, transferrin and IgG) with Pb(2+) was investigated using electrochemistry. The data was used to construct a new theoretical model of Pb(2+) binding to the high-abundance serum proteins under non-ideal conditions. The binding constants (?) of Pb(2+) to the individual proteins and a mixture of proteins were measured according to a new theoretical equation (non-ideal state) as well as the McGhee-Von Hippel equation (ideal state). Differences between the models suggested that the ? values obtained using the non-ideal state model was more realistic. Protein-protein interactions and micro-environmental influences affected binding between Pb(2+) and the high-abundance serum proteins. We included a micro-environmental influence factor for the model (Fm), which accurately quantified the effect of micro-environment of the proteome of Pb(2+) binding with the serum proteins. This research provides a useful reference of theoretical and experimental work regarding heavy-metal binding interactions with serum proteins. PMID:25048942

  3. Y682G Mutation of Amyloid Precursor Protein Promotes Endo-Lysosomal Dysfunction by Disrupting APP–SorLA Interaction

    Science.gov (United States)

    La Rosa, Luca Rosario; Perrone, Lorena; Nielsen, Morten Schallburg; Calissano, Pietro; Andersen, Olav Michael; Matrone, Carmela

    2015-01-01

    The intracellular transport and localization of amyloid precursor protein (APP) are critical determinants of APP processing and ?-amyloid peptide production, thus crucially important for the pathophysiology of Alzheimer’s disease (AD). Notably, the C-terminal Y682ENPTY687 domain of APP binds to specific adaptors controlling APP trafficking and sorting in neurons. Mutation on the Y682 residue to glycine (Y682G) leads to altered APP sorting in hippocampal neurons that favors its accumulation in intracellular compartments and the release of soluble APP?. Such alterations induce premature aging and learning and cognitive deficits in APP Y682G mutant mice (APPYG/YG). Here, we report that Y682G mutation affects formation of the APP complex with sortilin-related receptor (SorLA), resulting in endo-lysosomal dysfunctions and neuronal degeneration. Moreover, disruption of the APP/SorLA complex changes the trafficking pathway of SorLA, with its consequent increase in secretion outside neurons. Mutations in the SorLA gene are a prognostic factor in AD, and changes in SorLA levels in cerebrospinal fluid are predictive of AD in humans. These results might open new possibilities in comprehending the role played by SorLA in its interaction with APP and in the progression of neuronal degeneration. In addition, they further underline the crucial role played by Y682 residue in controlling APP trafficking in neurons. PMID:25904844

  4. Y682G Mutation of Amyloid Precursor Protein Promotes Endo-Lysosomal Dysfunction by Disrupting APP-SorLA Interaction.

    Science.gov (United States)

    La Rosa, Luca Rosario; Perrone, Lorena; Nielsen, Morten Schallburg; Calissano, Pietro; Andersen, Olav Michael; Matrone, Carmela

    2015-01-01

    The intracellular transport and localization of amyloid precursor protein (APP) are critical determinants of APP processing and ?-amyloid peptide production, thus crucially important for the pathophysiology of Alzheimer's disease (AD). Notably, the C-terminal Y682ENPTY687 domain of APP binds to specific adaptors controlling APP trafficking and sorting in neurons. Mutation on the Y682 residue to glycine (Y682G) leads to altered APP sorting in hippocampal neurons that favors its accumulation in intracellular compartments and the release of soluble APP?. Such alterations induce premature aging and learning and cognitive deficits in APP Y682G mutant mice (APP (YG/YG) ). Here, we report that Y682G mutation affects formation of the APP complex with sortilin-related receptor (SorLA), resulting in endo-lysosomal dysfunctions and neuronal degeneration. Moreover, disruption of the APP/SorLA complex changes the trafficking pathway of SorLA, with its consequent increase in secretion outside neurons. Mutations in the SorLA gene are a prognostic factor in AD, and changes in SorLA levels in cerebrospinal fluid are predictive of AD in humans. These results might open new possibilities in comprehending the role played by SorLA in its interaction with APP and in the progression of neuronal degeneration. In addition, they further underline the crucial role played by Y682 residue in controlling APP trafficking in neurons. PMID:25904844

  5. Perturbations of the straight transmembrane ?-helical structure of the amyloid precursor protein affect its processing by ?-secretase.

    Science.gov (United States)

    Lemmin, Thomas; Dimitrov, Mitko; Fraering, Patrick C; Dal Peraro, Matteo

    2014-03-01

    The amyloid precursor protein (APP) is a widely expressed type I transmembrane (TM) glycoprotein present at the neuronal synapse. The proteolytic cleavage by ?-secretase of its C-terminal fragment produces amyloid-? (A?) peptides of different lengths, the deposition of which is an early indicator of Alzheimer disease. At present, there is no consensus on the conformation of the APP-TM domain at the biological membrane. Although structures have been determined by NMR in detergent micelles, their conformation is markedly different. Here we show by using molecular simulations that the APP-TM region systematically prefers a straight ?-helical conformation once embedded in a membrane bilayer. However, APP-TM is highly flexible, and its secondary structure is strongly influenced by the surrounding lipid environment, as when enclosed in detergent micelles. This behavior is confirmed when analyzing in silico the atomistic APP-TM population observed by residual dipolar couplings and double electron-electron resonance spectroscopy. These structural and dynamic features are critical in the proteolytic processing of APP by the ?-secretase enzyme, as suggested by a series of Gly(700) mutants. Affecting the hydration and flexibility of APP-TM, these mutants invariantly show an increase in the production of A?38 compared with A?40 peptides, which is reminiscent of the effect of ?-secretase modulators inhibitors. PMID:24469457

  6. Quantitative Proteomic Analysis of Serum Proteins from Oral Cancer Patients: Comparison of Two Analytical Methods

    Directory of Open Access Journals (Sweden)

    Yan Yang

    2014-08-01

    Full Text Available Serum proteomic analysis can be a valuable approach for the discovery of protein biomarkers for early detection or monitoring of a disease. In this study, two analytical methods were compared for quantification of serum proteins in patients with oral cancer. In the first approach, we quantified serum proteins between oral squamous cell carcinoma (OSCC and healthy control subjects by performing in-solution digestion of serum proteins, isobaric tags for relative and absolute quantification (iTRAQ labeling of the resulting peptides, strong cation exchange (SCX fractionation of labeled peptides and finally capillary liquid chromatography with tandem mass spectrometry (LC-MS/MS analysis of the peptides. In the second approach, we first separated serum proteins with SDS-PAGE. The gel-separated proteins were then digested with trypsin and the resulting peptides were labeled with iTRAQ and analyzed with LC-MS/MS for protein quantification. A total of 319 serum proteins were quantified with the first proteomic approach whereas a total of 281 proteins were quantified by the second proteomic approach. Most of the proteins were identified and quantified by both approaches, suggesting that these methods are similarly effective for serum proteome analysis. This study provides compelling evidence that quantitative serum proteomic analysis of OSCC is a valuable approach for identifying differentially expressed proteins in cancer patients’ circulation systems that may be used as potential biomarkers for disease detection. Further validation in large oral cancer patient populations may lead to a simple and low invasive clinical tool for OSCC diagnosis or monitoring.

  7. Structural Analysis Reveals an Amyloid Form of the Human Papillomavirus Type 16 E1?E4 Protein and Provides a Molecular Basis for Its Accumulation?

    Science.gov (United States)

    McIntosh, Pauline B.; Martin, Stephen R.; Jackson, Deborah J.; Khan, Jameela; Isaacson, Erin R.; Calder, Lesley; Raj, Kenneth; Griffin, Heather M.; Wang, Qian; Laskey, Peter; Eccleston, John F.; Doorbar, John

    2008-01-01

    The abundant human papillomavirus (HPV) type 16 E4 protein exists as two distinct structural forms in differentiating epithelial cells. Monomeric full-length 16E1?E4 contains a limited tertiary fold constrained by the N and C termini. N-terminal deletions facilitate the assembly of E1?E4 into amyloid-like fibrils, which bind to thioflavin T. The C-terminal region is highly amyloidogenic, and its deletion abolishes amyloid staining and prevents E1?E4 accumulation. Amyloid-imaging probes can detect 16E1?E4 in biopsy material, as well as 18E1?E4 and 33E1?E4 in monolayer cells, indicating structural conservation. Our results suggest a role for fibril formation in facilitating the accumulation of E1?E4 during HPV infection. PMID:18562538

  8. Proteomic Analysis of Bovine Pregnancy-specific Serum Proteins by 2D Fluorescence Difference Gel Electrophoresis

    Science.gov (United States)

    Lee, Jae Eun; Lee, Jae Young; Kim, Hong Rye; Shin, Hyun Young; Lin, Tao; Jin, Dong Il

    2015-01-01

    Two dimensional-fluorescence difference gel electrophoresis (2D DIGE) is an emerging technique for comparative proteomics, which improves the reproducibility and reliability of differential protein expression analysis between samples. The purpose of this study was to investigate bovine pregnancy-specific proteins in the proteome between bovine pregnant and non-pregnant serum using DIGE technique. Serums of 2 pregnant Holstein dairy cattle at day 21 after artificial insemination and those of 2 non-pregnant were used in this study. The pre-electrophoretic labeling of pregnant and non-pregnant serum proteins were mixed with Cy3 and Cy5 fluorescent dyes, respectively, and an internal standard was labeled with Cy2. Labeled proteins with Cy2, Cy3, and Cy5 were separated together in a single gel, and then were detected by fluorescence image analyzer. The 2D DIGE method using fluorescence CyDye DIGE flour had higher sensitivity than conventional 2D gel electrophoresis, and showed reproducible results. Approximately 1,500 protein spots were detected by 2D DIGE. Several proteins showed a more than 1.5-fold up and down regulation between non-pregnant and pregnant serum proteins. The differentially expressed proteins were identified by MALDI-TOF mass spectrometer. A total 16 protein spots were detected to regulate differentially in the pregnant serum, among which 7 spots were up-regulated proteins such as conglutinin precursor, modified bovine fibrinogen and IgG1, and 6 spots were down-regulated proteins such as hemoglobin, complement component 3, bovine fibrinogen and IgG2a three spots were not identified. The identified proteins demonstrate that early pregnant bovine serum may have several pregnancy-specific proteins, and these could be a valuable information for the development of pregnancy-diagnostic markers in early pregnancy bovine serum. PMID:25925056

  9. A role for the high-density lipoprotein receptor SR-B1 in synovial inflammation via serum amyloid-A.

    LENUS (Irish Health Repository)

    Mullan, Ronan Hugh

    2012-02-01

    Acute phase apoprotein Serum Amyloid A (A-SAA), which is strongly expressed in rheumatoid arthritis synovial membrane (RA SM), induces angiogenesis, adhesion molecule expression, and matrix metalloproteinase production through the G-coupled receptor FPRL-1. Here we report alternative signaling through the high-density lipoprotein receptor scavenger receptor-class B type 1 (SR-B1). Quantitative expression\\/localization of SR-B1 in RA SM, RA fibroblast-like cells (FLCs), and microvascular endothelial cells (ECs) was assessed by Western blotting and immunohistology\\/fluorescence. A-SAA-mediated effects were examined using a specific antibody against SR-B1 or amphipathic alpha-Helical Peptides (the SR-B1 antagonists L-37pA and D-37pA), in RA FLCs and ECs. Adhesion molecule expression and cytokine production were quantified using flow cytometry and ELISA. SR-B1 was strongly expressed in the RA SM lining layer and endothelial\\/perivascular regions compared with osteoarthritis SM or normal control synovium. Differential SR-B1 expression in RA FLC lines (n = 5) and ECs correlated closely with A-SAA, but not tumor necrosis factor alpha-induced intercellular adhesion molecule-1 upregulation. A-SAA-induced interleukin-6 and -8 production was inhibited in the presence of anti-SR-B1 in human microvascular endothelial cells and RA FLCs. Moreover, D-37pA and L-37pA inhibited A-SAA-induced vascular cell adhesion molecule-1 and intercellular adhesion molecule expression from ECs in a dose-dependent manner. As SR-B1 is expressed in RA synovial tissue and mediates A-SAA-induced pro-inflammatory pathways, a better understanding of A-SAA-mediated inflammatory pathways may lead to novel treatment strategies for RA.

  10. Competitive Protein Adsorption of Albumin and Immunoglobulin G from Human Serum onto Polymer Surfaces

    DEFF Research Database (Denmark)

    Holmberg, Maria; Hou, Xiaolin

    2010-01-01

    Competitive protein adsorption from human serum onto unmodified polyethylene terephthalate (PET) surfaces and plasma-polymerized PET surfaces, using the monomer diethylene glycol vinyl ether (DEGVE), has been investigated using radioactive labeling. Albumin and immunoglobulin G (IgG) labeled with two different iodine isotopes have been added to human serum solutions of different concentrations, and adsorption has been performed using adsorption times from approximately 5 s to 24 h. DEGVE surfaces showed indications of being nonfouling regarding albumin and IgG adsorption during competitive protein adsorption from diluted human serum solutions with relatively low protein concentrations, but the nonfouling character was weakened when less diluted human serum solutions with higher protein concentrations were used. The observed adsorption trend is independent of adsorption time, indicating that the protein concentration has a stronger influence on observed adsorption characteristics of the material than the adsorption time has.

  11. Mechanism of Inhibition of beta-site amyloid precursor protein-cleaving enzyme (BACE) by a statine-based peptide.

    Science.gov (United States)

    Marcinkeviciene, J; Luo, Y; Graciani, N R; Combs, A P; Copeland, R A

    2001-06-29

    Inhibition of beta-site amyloid precursor protein-cleaving enzyme by a statine-based inhibitor has been studied using steady state and stopped-flow methods. A slow onset rate of inhibition has been observed under steady state conditions, and a K(i) of 22 nm has been derived using progress curves analysis. Simulation of stopped-flow protein fluorescence transients provided an estimate of the K(d) for initial inhibitor binding of 660 nm. A two-step inhibition mechanism is proposed, wherein slower "tightening up" of the initial encounter complex occurs. Two hypotheses have been proposed in the literature to address the nature of the slow step in the inhibition of aspartic proteases by peptidomimetic inhibitors: a conformational change related to the "flap" movement and displacement of a catalytic water. We compared substrate and inhibitor binding rates under pre-steady-state conditions. Both ligands are likely to cause flap movement, whereas no catalytic water replacement occurs during substrate binding. Our results suggest that both ligands bind to the enzyme at a rate significantly lower than the diffusion limit, but there are additional rate limitations involved in inhibitor binding, resulting in a k(on) of 3.5 x 10(4) m(-)1 s(-)1 for the inhibitor compared with 3.5 x 10(5) m(-)1 s(-)1 for the substrate. Even though specific intermediate formation steps might be different in the productive inhibitor and substrate binding to beta-site amyloid precursor protein-cleaving enzyme, a similar final optimized conformation is achieved in both cases, as judged by the comparable free energy changes (DeltaDeltaG of 2.01 versus 1.97 kcal/mol) going from the initial to the final enzyme-inhibitor or enzyme-substrate complexes. PMID:11306583

  12. Purification of Hyaluronan Binding Proteins from Human Normal and Cancer Serum

    Directory of Open Access Journals (Sweden)

    Fekry B

    2010-04-01

    Full Text Available Background: Analysis of human cancer serum has revealed the presence of high amounts of hyaluronan (hyaluronic acid, HA when compared to human normal serum, It is well documented that HA and its receptors, known as hyaladherins (HABPs are involved in matrix regulation, cell proliferation, migration and malignant tumour progression. These hyaladherins not only interact with hyaluronan at the matrix proper but also with hyaluronan at the plasma membrane as a cell surface receptors and thus influence cell physiology including secretion of this protein into the circulatory system. Methods: Normal serum and colon cancer serum samples were included in this study, using biochemical techniques such as gel permeation, strong anion exchange chromatography, single dimension electrophoresis and western blot analysis. Results: This study is based on the clinical work of normal serum and colon cancer serum. The description of the procedure was given for the fractionation of serum proteins mainly HABPs from 20 normal and 15 colon cancer patients by using special biotinylated hyaluronan probe. Conclusion: To evaluate whether serum HABPs levels could be used as diagnostic marker for human cancer. The semi purified serum from normal and colon cancer patients showed mainly a major protein (57kDa and a minor one (30kDa by overlay experiments with b-HA probe and these results were confirmed by competition experiments with cold HA.

  13. Curcumin mitigates the fibrillation of human serum albumin and diminishes the formation of reactive oxygen species.

    Science.gov (United States)

    Mazaheri, Mansooreh; Moosavi-Movahedi, Ali Akbar; Saboury, Ali Akbar; Rezaei, Mehran Habibi; Shourian, Mostafa; Farhadi, Mohammad; Sheibani, Nader

    2015-01-01

    The formation of amyloid fibrils are thought to contribute to pathogenesis of many amyloids associated human diseases. Here the impact of curcumin on amyloid formation of human serum albumin (HSA) was studied. Incubation of HSA at 68°C under physiologic pH led to amyloid fibril formation. Thioflavin T (ThT) fluorescence was used for determination of amyloid fibril formation. Atomic force microscopy experiments indicated different fibril structure of HSA incubated with or without curcumin. The monitoring of the changes in reactive oxygen species (ROS) levels upon incubation of curcumin with HSA showed a significant decrease in ROS levels. Similar experiments were also carried out in the presence of aflatoxin M1 (AFM1) and lead (Pb) ions. Our results indicated that AFM1 and Pb ions promote the fibrillation of HSA and accelerate ROS production, which were inhibited in the presence of curcumin. Thus, curcumin mitigates protein fibrillation activity and diminishes ROS generation. PMID:25666039

  14. Accumulation of Intraneuronal ?-Amyloid 42 Peptides Is Associated with Early Changes in Microtubule-Associated Protein 2 in Neurites and Synapses

    OpenAIRE

    Takahashi, Reisuke H.; Capetillo-Zarate, Estibaliz; Lin, Michael T; Milner, Teresa A; Gouras, Gunnar K.

    2013-01-01

    Pathologic aggregation of ?-amyloid (A?) peptide and the axonal microtubule-associated protein tau protein are hallmarks of Alzheimer's disease (AD). Evidence supports that A? peptide accumulation precedes microtubule-related pathology, although the link between A? and tau remains unclear. We previously provided evidence for early co-localization of A?42 peptides and hyperphosphorylated tau within postsynaptic terminals of CA1 dendrites in the hippocampus of AD transgenic mice. Here, we ...

  15. Accumulation of Intraneuronal beta-Amyloid 42 Peptides Is Associated with Early Changes in Microtubule-Associated Protein 2 in Neurites and Synapses

    OpenAIRE

    Takahashi, Reisuke H.; Capetillo-Zarate, Estibaliz; Lin, Michael T; Milner, Teresa A; Gouras, Gunnar

    2013-01-01

    Pathologic aggregation of beta-amyloid (A beta) peptide and the axonal microtubule-associated protein tau protein are hallmarks of Alzheimer's disease (AD). Evidence supports that A beta peptide accumulation precedes microtubule-related pathology, although the link between A beta and tau remains unclear. We previously provided evidence for early co-localization of A beta 42 peptides and hyperphosphorylated tau within postsynaptic terminals of CA1 dendrites in the hippocampus of AD transge...

  16. Testing the Neurovascular Hypothesis of Alzheimer’s Disease: LRP-1 Antisense Reduces Blood-Brain Barrier Clearance, Increases Brain Levels of Amyloid-? Protein, and Impairs Cognition

    OpenAIRE

    Jaeger, Laura B.; Dohgu, Shinya; Hwang, Mark C.; Farr, Susan A.; Murphy, M Paul; Fleegal-DeMotta, Melissa A.; Lynch, Jessica L.; Robinson, Sandra M.; Niehoff, Michael L.; Johnson, Steven N.; Kumar, Vijaya B.; Banks, William A.

    2009-01-01

    Decreased clearance is the main reason amyloid-? protein (A?) in increased in the brains of patients with Alzheimer’s disease (AD). The neurovascular hypothesis states that this decreased clearance is caused by impairment of low density lipoprotein receptor related protein-1 (LRP-1), the major brain-to-blood transporter of A? at the blood-brain barrier (BBB). As deletion of the LRP-1 gene is a lethal mutation, we tested the neurovascular hypothesis by developing a cocktail of phosphorothioate...

  17. Functional Amyloid Formation within Mammalian Tissue.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available Amyloid is a generally insoluble, fibrous cross-beta sheet protein aggregate. The process of amyloidogenesis is associated with a variety of neurodegenerative diseases including Alzheimer, Parkinson, and Huntington disease. We report the discovery of an unprecedented functional mammalian amyloid structure generated by the protein Pmel17. This discovery demonstrates that amyloid is a fundamental nonpathological protein fold utilized by organisms from bacteria to humans. We have found that Pmel17 amyloid templates and accelerates the covalent polymerization of reactive small molecules into melanin-a critically important biopolymer that protects against a broad range of cytotoxic insults including UV and oxidative damage. Pmel17 amyloid also appears to play a role in mitigating the toxicity associated with melanin formation by sequestering and minimizing diffusion of highly reactive, toxic melanin precursors out of the melanosome. Intracellular Pmel17 amyloidogenesis is carefully orchestrated by the secretory pathway, utilizing membrane sequestration and proteolytic steps to protect the cell from amyloid and amyloidogenic intermediates that can be toxic. While functional and pathological amyloid share similar structural features, critical differences in packaging and kinetics of assembly enable the usage of Pmel17 amyloid for normal function. The discovery of native Pmel17 amyloid in mammals provides key insight into the molecular basis of both melanin formation and amyloid pathology, and demonstrates that native amyloid (amyloidin may be an ancient, evolutionarily conserved protein quaternary structure underpinning diverse pathways contributing to normal cell and tissue physiology.

  18. Serum protein identification and quantification of the corona of 5, 15 and 80 nm gold nanoparticles

    International Nuclear Information System (INIS)

    When nanoparticles (NP) enter the body they come into contact with body fluids containing proteins which can adsorb to their surface. These proteins may influence the NP interactions with the biological vicinity, eventually determining their biological fate inside the body. Adsorption of the most abundantly binding proteins was studied after an in vitro 24 hr incubation of monodisperse, negatively charged 5, 15 and 80 nm gold spheres (AuNP) in mouse serum by a two-step analysis: proteomic protein identification and quantitative protein biochemistry. The adsorbed proteins were separated from non-adsorbed proteins by centrifugation and gel electrophoresis and identified using a MALDI-TOF-MS-Proteomics-Analyzer. Quantitative analysis of proteins in gel bands by protein densitometry, required the focus on predominantly binding serum proteins. Numerous proteins adsorbed to the AuNP depending on their size, e.g. apolipoproteins or complement C3. The qualitative and quantitative amount of adsorbed proteins differed between 5, 15 and 80 nm AuNP. Band intensities of adsorbed proteins decreased with increasing AuNP sizes based not only on their mass but also on their surface area. Summarizing, the AuNP surface is covered with serum proteins containing transport and immune related proteins among others. Hence, protein binding depends on the size, surface area and curvature of the AuNP. (paper)

  19. Amyloid precursor-like protein 2 (APLP2) affects the actin cytoskeleton and increases pancreatic cancer growth and metastasis.

    Science.gov (United States)

    Pandey, Poomy; Rachagani, Satyanarayana; Das, Srustidhar; Seshacharyulu, Parthasarathy; Sheinin, Yuri; Naslavsky, Naava; Pan, Zenggang; Smith, Brittney L; Peters, Haley L; Radhakrishnan, Prakash; McKenna, Nicole R; Giridharan, Sai Srinivas Panapakkam; Haridas, Dhanya; Kaur, Sukhwinder; Hollingsworth, Michael A; MacDonald, Richard G; Meza, Jane L; Caplan, Steve; Batra, Surinder K; Solheim, Joyce C

    2015-02-10

    Amyloid precursor-like protein 2 (APLP2) is aberrantly expressed in pancreatic cancer. Here we showed that APLP2 is increased in pancreatic cancer metastases, particularly in metastatic lesions found in the diaphragm and intestine. Examination of matched human primary tumor-liver metastasis pairs showed that 38.1% of the patients had positive APLP2 expression in both the primary tumor and the corresponding liver metastasis. Stable knock-down of APLP2 expression (with inducible shRNA) in pancreatic cancer cells reduced the ability of these cells to migrate and invade. Loss of APLP2 decreased cortical actin and increased intracellular actin filaments in pancreatic cancer cells. Down-regulation of APLP2 decreased the weight and metastasis of orthotopically transplanted pancreatic tumors in nude mice. PMID:25576918

  20. Characterisation of the effect of knockout of the amyloid precursor protein on outcome following mild traumatic brain injury.

    Science.gov (United States)

    Corrigan, Frances; Vink, Robert; Blumbergs, Peter C; Masters, Colin L; Cappai, Roberto; van den Heuvel, Corinna

    2012-04-27

    The amyloid precursor protein (APP) increases following traumatic brain injury (TBI), although the functional significance of this remains unclear largely because the functions of the subsequent APP metabolites are so different: A? is neurotoxic whilst sAPP? is neuroprotective. To investigate this further, APP wildtype and knockout mice were subjected to mild diffuse TBI and their outcomes compared. APP knockout mice displayed significantly worse cognitive and motor deficits, as demonstrated by the Barnes Maze and rotarod respectively, than APP wildtype mice. This was associated with a significant increase in hippocampal and cortical cell loss, as well as axonal injury, in APP knockout mice and an impaired neuroreparative response as indicated by diminished GAP-43 immunoreactivity when compared to APP wildtype mice. This study is the first to demonstrate that endogenous APP is beneficial following mild TBI, suggesting that the upregulation of APP observed following injury is an acute protective response. PMID:22424792

  1. A new molecular explanation for age-related neurodegeneration : The Tyr682 residue of amyloid precursor protein

    DEFF Research Database (Denmark)

    Matrone, Carmela; Matrone, Carmela

    2013-01-01

    Emerging evidence supports the role for the intracellular domains of amyloid precursor protein (APP) in the physiology and function of APP. In this short report, I discuss the hypothesis that mutation of Tyr682 on the Y682 ENPTY687 C-terminal motif of APP may be directly or indirectly associated with alterations in APP functioning and activity, leading to neuronal defects and deficits. Mutation of Tyr682 induces an early and progressive age-dependent cognitive and locomotor decline that is associated with a loss of synaptic connections, a decrease in cholinergic tone, and defects in NGF signaling. These findings support a model in which APP-C-terminal domain exerts a pathogenic function in neuronal development and decline, and suggest that Tyr682 potentially could modulate the properties of APP metabolites in humans.

  2. The prion protein regulates beta-amyloid-mediated self-renewal of neural stem cells in vitro.

    Science.gov (United States)

    Collins, Steven J; Tumpach, Carolin; Li, Qiao-Xin; Lewis, Victoria; Ryan, Timothy M; Roberts, Blaine; Drew, Simon C; Lawson, Victoria A; Haigh, Cathryn L

    2015-01-01

    The beta-amyloid (A?) peptide and the A?-oligomer receptor, prion protein (PrP), both influence neurogenesis. Using in vitro murine neural stem cells (NSCs), we investigated whether A? and PrP interact to modify neurogenesis. A? imparted PrP-dependent changes on NSC self-renewal, with PrP-ablated and wild-type NSCs displaying increased and decreased cell growth, respectively. In contrast, differentiation of A?-treated NSCs into mature cells was unaffected by PrP expression. Such marked PrP-dependent differences in NSC growth responses to A? provides further evidence of biologically significant interactions between these two factors and an important new insight into regulation of NSC self-renewal in vivo. PMID:25884827

  3. Structural insight into the mechanism of amyloid precursor protein recognition by ?-secretase 1: A molecular dynamics study.

    Science.gov (United States)

    Chakraborty, Sandipan; Basu, Soumalee

    2015-07-01

    ?-secretase 1 (BACE1) initiates the proteolysis of amyloid precursor protein (APP) to generate A?, aggregation of which has been considered to be the main histopathological feature of Alzheimer's Disease. Here, we have explored the conformational switching of BACE1 during APP recognition using molecular dynamics simulation thereby suggesting the recognition to be a conformational selection process. Free BACE1 is highly flexible and exists as an ensemble of conformations. The ?-hairpin flap that covers the active site of BACE1 visits numerous conformations during the simulation. Essential dynamics reveal that concerted movements in several loops including the flap region lead to a conformational switching from open to closed form. During the simulation, free BACE1 visits both the open and closed forms multiple times. Binding of APP to the BACE1 cavity shifts the equilibrium towards a stable complex stabilized by strong electrostatic surface complementarity along with several van der Waals and hydrogen bonding interactions. PMID:25863345

  4. Cellular Prion Protein Mediates Impairment of Synaptic Plasticity by Amyloid-? Oligomers

    OpenAIRE

    Laurén, Juha; Gimbel, David A.; Nygaard, Haakon B.; Gilbert, John W.; Strittmatter, Stephen M

    2009-01-01

    A pathological hallmark of Alzheimer’s disease (AD) is an accumulation of insoluble plaque containing the amyloid-? peptide (A?) of 40–42 aa residues1. Prefibrillar, soluble oligomers of A? have been recognized to be early and key intermediates in AD-related synaptic dysfunction2–9. At nanomolar concentrations, soluble A?-oligomers block hippocampal long-term potentiation7, cause dendritic spine retraction from pyramidal cells5,8 and impair rodent spatial memory2. Soluble A?-oligom...

  5. Systematic studies of the interaction between amyloid beta-protein and lipids

    OpenAIRE

    Song, Haipeng

    2005-01-01

    The amyloid peptide (Aß), a normal constituent of neuronal and non-neuronal cells, has been shown to be a major component of the extracellular plaque of Alzheimer’s disease (AD). The interaction of Aß peptides with the lipid matrix of neuronal cell membranes plays an important role in the pathogenesis of AD. In this study, we have developed peptide-tethered artificial lipid membranes by the Langmuir-Blodgett and Langmuir-Schaefer methods. Anti-Aß40-mAb labeled with a fluorophore was used...

  6. Blood-Serum Proteins of Rattus rattus and Rattus norvegicus (Mammalia: Rodentia) in Turkey

    OpenAIRE

    Yi?g?i?t, Nuri; Veri?mli?, Reyhan; C?olak, Ercu?ment; So?zen, Mustafa

    2001-01-01

    The blood serum proteins of 35 live specimens of Rattus rattus and Rattus norvegicus collected from ten localities in Turkey were examined by SDS-PAGE (sodium dodecyl sulphate - polyacrylamide gel electrophoresis). Globulin, albumin, postalbumin, and prealbumin proteins in blood serum showed different electrophoretic patterns in populations of R. rattusand R. norvegicus.Because of the variations in electrophoretic patterns of both species, these patterns do not seem to be diagnostic char...

  7. Crystallization and preliminary crystallographic studies of the copper-binding domain of the amyloid precursor protein of Alzheimer’s disease

    International Nuclear Information System (INIS)

    The binding of Cu2+ ions to the copper-binding domain of the amyloid precursor protein of Alzheimer’s disease reduces the production of the amyloid ? peptide, which is centrally involved in Alzheimer’s disease. Structural studies of the copper-binding domain will provide a basis for structure-based drug design that might prove useful in treating this devastating disease. Alzheimer’s disease is thought to be triggered by production of the amyloid ? (A?) peptide through proteolytic cleavage of the amyloid precursor protein (APP). The binding of Cu2+ to the copper-binding domain (CuBD) of APP reduces the production of A? in cell-culture and animal studies. It is expected that structural studies of the CuBD will lead to a better understanding of how copper binding causes A? depletion and will define a potential drug target. The crystallization of CuBD in two different forms suitable for structure determination is reported here

  8. Radioprotective properties of certain nitrogenous compounds heterocyclic on the serum proteins of irradiated mice

    International Nuclear Information System (INIS)

    The results obtained from this study suggest the following: the concentration of total serum proteins in mice is very little changed during all the treatments carried out, while protein fractions showed significant alterations. The concentrations of various serum proteins remain almost constant under normal conditions. Intraperitoneal administration of imidazole or benzimidazole at the mentioned doses induces rapid quantitative changes in the serum which are recovered in about 3 days Whole-body X-irradiation at 750 roentgens creates slow but progressive and persisting serious changes in a concentration of serum protein fractions which end by death of animals at the 8 - 10. day after irradiation. Whole-body X-irradiation of imidazole or benzimidazole protected animals results in quantitative rapid changes in concentration of serum protein fractions, for about four days after which a slow but steady restoration begins. The concentration approaches the normal levels towards the 10. day after irradiation. Imidazole and benzimidazole were proved to be good radio-protectants against the effects of radiation on serum protein fractions. Benzimidazole seems to surpass imidazole. (authors)

  9. Peptide sequences that target proteins to lysosomes for enhanced degradation during serum withdrawal

    International Nuclear Information System (INIS)

    Ribonuclease A (RNase A) microinjected into human diploid fibroblasts is degraded with a half-life of 80-100 h in the presence of serum, but its half-life declines to 40 h when cells are deprived of serum. This increased degradation in response to serum withdrawal results from an increased rate of uptake of microinjected RNase A from the cytosol into lysosomes. The cells' ability to recognize RNase A for this enhanced degradation is based on some feature of amino acids 1-20. Covalent linkage of S-peptide to other proteins results in enhanced lysosomal degradation of the conjugate in response to serum withdrawal. The authors have further defined the essential region of RNase S-peptide to be within residues 7-11, KFERQ. Coinjection of radiolabeled RNase A and excess unlabeled pentapeptide specifically blocks the enhanced degradation of RNase A. Affinity-purified polyclonal IgGs raised against KFERQ immunoprecipitate 25-30% of radiolabeled cytosolic proteins from human fibroblasts. Pulse-chase experiments indicate that these proteins are preferentially degraded upon serum withdrawal while degradation of nonimmuno-precipitated proteins is unaffected. These results suggest that peptide sequences similar to KFERQ are contained within cellular proteins and that such sequences target proteins to lysosomes for enhanced degradation during serum withdrawal

  10. Amyloid Beta as a Modulator of Synaptic Plasticity

    OpenAIRE

    Parihar, Mordhwaj S.; Brewer, Gregory J.

    2010-01-01

    Alzheimer’s disease is associated with synapse loss, memory dysfunction and pathological accumulation of amyloid beta in plaques. However, an exclusively pathological role for amyloid beta is being challenged by new evidence for an essential function of amyloid beta at the synapse. Amyloid beta protein exists in different assembly states in the central nervous system and plays distinct roles ranging from synapse and memory formation to memory loss and neuronal cell death. Amyloid beta is pr...

  11. Potential Natural Products for Alzheimer's Disease: Targeted Search Using the Internal Ribosome Entry Site of Tau and Amyloid-? Precursor Protein.

    Science.gov (United States)

    Tasi, Yun-Chieh; Chin, Ting-Yu; Chen, Ying-Ju; Huang, Chun-Chih; Lee, Shou-Lun; Wu, Tzong-Yuan

    2015-01-01

    Overexpression of the amyloid precursor protein (APP) and the hyperphosphorylation of the tau protein are vital in the understanding of the cause of Alzheimer's disease (AD). As a consequence, regulation of the expression of both APP and tau proteins is one important approach in combating AD. The APP and tau proteins can be targeted at the levels of transcription, translation and protein structural integrity. This paper reports the utilization of a bi-cistronic vector containing either APP or tau internal ribosome entry site (IRES) elements flanked by ?-galactosidase gene (cap-dependent) and secreted alkaline phosphatase (SEAP) (cap-independent) to discern the mechanism of action of memantine, an N-methyl-d-aspartate (NMDA) receptor antagonist. Results indicate that memantine could reduce the activity of both the APP and tau IRES at a concentration of ~10 ?M (monitored by SEAP activity) without interfering with the cap-dependent translation as monitored by the ?-galactosidase assay. Western blot analysis of the tau protein in neuroblastoma (N2A) and rat hippocampal cells confirmed the halting of the expression of the tau proteins. We also employed this approach to identify a preparation named NB34, extracts of Boussingaultia baselloides (madeira-vine) fermented with Lactobacillus spp., which can function similarly to memantine in both IRES of APP and Tau. The water maze test demonstrated that NB34 could improve the spatial memory of a high fat diet induced neurodegeneration in apolipoprotein E-knockout (ApoE-/-) mice. These results revealed that the bi-cistronic vector provided a simple, and effective platform in screening and establishing the mechanistic action of potential compounds for the treatment and management of AD. PMID:25903151

  12. Depletion of Amyloid Precursor Protein (APP) causes G0 arrest in non-small cell lung cancer (NSCLC) cells.

    Science.gov (United States)

    Sobol, Anna; Galluzzo, Paola; Weber, Megan J; Alani, Sara; Bocchetta, Maurizio

    2015-06-01

    We recently reported that Amyloid Precursor Protein (APP) regulates global protein synthesis in a variety of human dividing cells, including non-small cell lung cancer (NSCLC) cells. More specifically, APP depletion causes an increase of both cap- and IRES-dependent translation. Since growth and proliferation are tightly coupled processes, here, we asked what effects artificial downregulation of APP could have elicited in NSCLC cells proliferation. APP depletion caused a G0/G1 arrest through destabilization of the cyclin-C protein and reduced pRb phosphorylation at residues Ser802/811. siRNA to cyclin-C mirrored the cell cycle distribution observed when silencing APP. Cells arrested in G0/G1 (and with augmented global protein synthesis) increased their size and underwent a necrotic cell death due to cell membrane permeabilization. These phenotypes were reversed by overexpression of the APP C-terminal domain, indicating a novel role for APP in regulating early cell cycle entry decisions. It is seems that APP moderates the rate of protein synthesis before the cell clears growth factors- and nutrients-dependent checkpoint in mid G1. Our results raise questions on how such processes interact in the context of (at least) dividing NSCLC cells. The data presented here suggest that APP, although required for G0/G1 transitions, moderates the rate of protein synthesis before the cell fully commits to cell cycle progression following mechanisms, which seem additional to concurrent signals deriving from the PI3-K/Akt/mTORC-1 axis. APP appears to play a central role in regulating cell cycle entry with the rate of protein synthesis; and its loss-of-function causes cell size abnormalities and death. PMID:25502341

  13. Serum protein binding of diazepam in maternal and foetal serum during pregnancy.

    OpenAIRE

    Lee, J. N.; Chen, S. S.; Richens, A.; Menabawey, M.; Chard, T.

    1982-01-01

    1 The serum binding capacity for diazepam was significantly lower in pregnancy and there was a linear correlation with gestational age. 2 The binding of diazepam was not correlated to albumin during pregnancy. 3 In cord sera there was a significantly reduced binding capacity for diazepam with albumin levels of less than 40 g/l.

  14. Comparison of functional properties of 34% and 80% whey protein and milk serum protein concentrates.

    Science.gov (United States)

    Luck, P J; Vardhanabhuti, B; Yong, Y H; Laundon, T; Barbano, D M; Foegeding, E A

    2013-09-01

    This study compared the functional properties of serum protein concentrate (SPC) with whey protein concentrate (WPC) made from the same milk and with commercial WPC. The experimental SPC and WPC were produced at 34% or 80% protein from the same lot of milk. Protein contents of WPC and SPC were comparable; however, fat content was much lower in SPC compared with WPC and commercial WPC. The effect of drying methods (freeze vs. spray drying) was studied for 34% WPC and SPC. Few differences due to drying method were found in turbidity and gelation; however, drying method made a large difference in foam formation for WPC but not SPC. Between pH 3 and 7, SPC was found to have lower turbidity than WPC; however, protein solubility was similar between SPC and WPC. Foaming and gelation properties of SPC were better than those of WPC. Differences in functional properties may be explained by differences in composition and extent of denaturation or aggregation. PMID:23871371

  15. ALTERATIONS IN TOTAL PROTEIN CONCENTRATION, SERUM PROTEIN FRACTIONS AND ALBUMIN/GLOBULIN RATIO IN HEALTHY RABBITS

    Directory of Open Access Journals (Sweden)

    Nuzhat Sultana

    2013-08-01

    Full Text Available This study assessed the effect of oral administration of Aloe vera and was to evaluate total serum protein, albumin and globulin concentrations as well as albumin / globulin (A / G ratio. Twenty rabbits weighing 1000 – 1800 g were divided into 2 groups. Each group consisted of ten animals. One served as control and other group served as experimental group. Results show that animals after 07, 15 and 30 days dosing of Aloe vera showed highly significant decrease in total protein and globulin and highly significant decrease in Albumin after 15 and 30 days of dosing of Aloe vera in comparison to control animals group. It is concluded that the long-term use of Aloe vera may cause hypoglobinemia and hypoalbuminemia at 30 days of dosing and it could be due to the liver diseases, evidence of hepatotoxicity induced Aloe vera also reported in previous studies.

  16. Monoacylated Cellular Prion Proteins Reduce Amyloid-?-Induced Activation of Cytoplasmic Phospholipase A2 and Synapse Damage

    Directory of Open Access Journals (Sweden)

    Ewan West

    2015-06-01

    Full Text Available Alzheimer’s disease (AD is a progressive neurodegenerative disease characterized by the accumulation of amyloid-? (A? and the loss of synapses. Aggregation of the cellular prion protein (PrPC by A? oligomers induced synapse damage in cultured neurons. PrPC is attached to membranes via a glycosylphosphatidylinositol (GPI anchor, the composition of which affects protein targeting and cell signaling. Monoacylated PrPC incorporated into neurons bound “natural A?”, sequestering A? outside lipid rafts and preventing its accumulation at synapses. The presence of monoacylated PrPC reduced the A?-induced activation of cytoplasmic phospholipase A2 (cPLA2 and A?-induced synapse damage. This protective effect was stimulus specific, as treated neurons remained sensitive to ?-synuclein, a protein associated with synapse damage in Parkinson’s disease. In synaptosomes, the aggregation of PrPC by A? oligomers triggered the formation of a signaling complex containing the cPLA2.a process, disrupted by monoacylated PrPC. We propose that monoacylated PrPC acts as a molecular sponge, binding A? oligomers at the neuronal perikarya without activating cPLA2 or triggering synapse damage.

  17. Agitation of amyloid proteins to speed aggregation measured by ThT fluorescence: a call for standardization.

    Science.gov (United States)

    Batzli, Kiersten M; Love, Brian J

    2015-03-01

    This retrospective study of protein aggregation measured by Thioflavin T (ThT) fluorescence assay in published literature has assessed protein sensitivity to denaturing conditions that include elevated temperatures, fluctuations in pH, and concentration and, in particular, agitation to induce amyloid structure formation. The dynamic tracking of fluorescence shows a sigmoidal evolution as aggregates form; the resulting kinetics of association have been analyzed to explore the range of aggregation behavior which occurs based on environmental parameters. Comparisons between the experimental results of different groups have been historically difficult due to subtleties of experimental procedures including denaturing temperature, protein type and concentration, formulation differences, and how agitation is achieved. While it is clear that agitation has a strong influence on the driving force for aggregation, the use of magnetic stirring bar or shaker table rotational speed is insufficient to characterize the degree of turbulence produced during shear. The pathway forward in resolving dependence of aggregate formation on shear may require alternative methodologies or better standardization of the experimental protocols. PMID:25579934

  18. Transcriptional regulation of human FE65, a ligand of Alzheimer's disease amyloid precursor protein, by Sp1.

    LENUS (Irish Health Repository)

    Yu, Hoi-Tin

    2010-03-01

    FE65 is a neuronal-enriched adaptor protein that binds to the Alzheimer\\'s disease amyloid precursor protein (APP). FE65 forms a transcriptionally active complex with the APP intracellular domain (AICD). The precise gene targets for this complex are unclear but several Alzheimer\\'s disease-linked genes have been proposed. Additionally, evidence suggests that FE65 influences APP metabolism. The mechanism by which FE65 expression is regulated is as yet unknown. To gain insight into the regulatory mechanism, we cloned a 1.6 kb fragment upstream of the human FE65 gene and found that it possesses particularly strong promoter activity in neurones. To delineate essential regions in the human FE65 promoter, a series of deletion mutants were generated. The minimal FE65 promoter was located between -100 and +5, which contains a functional Sp1 site. Overexpression of the transcription factor Sp1 potentiates the FE65 promoter activity. Conversely, suppression of the FE65 promoter was observed in cells either treated with an Sp1 inhibitor or in which Sp1 was knocked down. Furthermore, reduced levels of Sp1 resulted in downregulation of endogenous FE65 mRNA and protein. These findings reveal that Sp1 plays a crucial role in transcriptional control of the human FE65 gene.

  19. Amyloid protein-mediated differential DNA methylation status regulates gene expression in Alzheimer’s disease model cell line

    International Nuclear Information System (INIS)

    Highlights: ? Genome-wide DNA methylation pattern in Alzheimer’s disease model cell line. ? Integrated analysis of CpG methylation and mRNA expression profiles. ? Identify three Swedish mutant target genes; CTIF, NXT2 and DDR2 gene. ? The effect of Swedish mutation on alteration of DNA methylation and gene expression. -- Abstract: The Swedish mutation of amyloid precursor protein (APP-sw) has been reported to dramatically increase beta amyloid production through aberrant cleavage at the beta secretase site, causing early-onset Alzheimer’s disease (AD). DNA methylation has been reported to be associated with AD pathogenesis, but the underlying molecular mechanism of APP-sw-mediated epigenetic alterations in AD pathogenesis remains largely unknown. We analyzed genome-wide interplay between promoter CpG DNA methylation and gene expression in an APP-sw-expressing AD model cell line. To identify genes whose expression was regulated by DNA methylation status, we performed integrated analysis of CpG methylation and mRNA expression profiles, and identified three target genes of the APP-sw mutant; hypomethylated CTIF (CBP80/CBP20-dependent translation initiation factor) and NXT2 (nuclear exporting factor 2), and hypermethylated DDR2 (discoidin domain receptor 2). Treatment with the demethylating agent 5-aza-2?-deoxycytidine restored mRNA expression of these three genes, implying methylation-dependent transcriptional regulation. The profound alteration in the methylation status was detected at the ?435, ?295, and ?271 CpG sites of CTIF, and at the ?505 to ?341 region in the promoter of DDR2. In the promoter region of NXT2, only one CpG site located at ?432 was differentially unmethylated in APP-sw cells. Thus, we demonstrated the effect of the APP-sw mutation on alteration of DNA methylation and subsequent gene expression. This epigenetic regulatory mechanism may contribute to the pathogenesis of AD.

  20. Characterization of the beta amyloid precursor protein-like gene in the central nervous system of the crab Chasmagnathus. Expression during memory consolidation

    Directory of Open Access Journals (Sweden)

    Fustiñana Maria

    2010-09-01

    Full Text Available Abstract Background Human ?-amyloid, the main component in the neuritic plaques found in patients with Alzheimer's disease, is generated by cleavage of the ?-amyloid precursor protein. Beyond the role in pathology, members of this protein family are synaptic proteins and have been associated with synaptogenesis, neuronal plasticity and memory, both in vertebrates and in invertebrates. Consolidation is necessary to convert a short-term labile memory to a long-term and stable form. During consolidation, gene expression and de novo protein synthesis are regulated in order to produce key proteins for the maintenance of plastic changes produced during the acquisition of new information. Results Here we partially cloned and sequenced the beta-amyloid precursor protein like gene homologue in the crab Chasmagnathus (cappl, showing a 37% of identity with the fruit fly Drosophila melanogaster homologue and 23% with Homo sapiens but with much higher degree of sequence similarity in certain regions. We observed a wide distribution of cappl mRNA in the nervous system as well as in muscle and gills. The protein localized in all tissues analyzed with the exception of muscle. Immunofluorescence revealed localization of cAPPL in associative and sensory brain areas. We studied gene and protein expression during long-term memory consolidation using a well characterized memory model: the context-signal associative memory in this crab species. mRNA levels varied at different time points during long-term memory consolidation and correlated with cAPPL protein levels Conclusions cAPPL mRNA and protein is widely distributed in the central nervous system of the crab and the time course of expression suggests a role of cAPPL during long-term memory formation.

  1. Pro-Inflammatory S100A8 and S100A9 Proteins: Self-Assembly into Multifunctional Native and Amyloid Complexes

    Directory of Open Access Journals (Sweden)

    Ludmilla A. Morozova-Roche

    2012-03-01

    Full Text Available S100A8 and S100A9 are EF-hand Ca2+ binding proteins belonging to the S100 family. They are abundant in cytosol of phagocytes and play critical roles in numerous cellular processes such as motility and danger signaling by interacting and modulating the activity of target proteins. S100A8 and S100A9 expression levels increased in many types of cancer, neurodegenerative disorders, inflammatory and autoimmune diseases and they are implicated in the numerous disease pathologies. The Ca2+ and Zn2+-binding properties of S100A8/A9 have a pivotal influence on their conformation and oligomerization state, including self-assembly into homo- and heterodimers, tetramers and larger oligomers. Here we review how the unique chemical and conformational properties of individual proteins and their structural plasticity at the quaternary level account for S100A8/A9 functional diversity. Additional functional diversification occurs via non-covalent assembly into oligomeric and fibrillar amyloid complexes discovered in the aging prostate and reproduced in vitro. This process is also regulated by Ca2+and Zn2+-binding and effectively competes with the formation of the native complexes. High intrinsic amyloid-forming capacity of S100A8/A9 proteins may lead to their amyloid depositions in numerous ailments characterized by their elevated expression patterns and have additional pathological significance requiring further thorough investigation.

  2. Binding of radioiodinated human ?-endorphin to serum proteins from rats and humans, determined by several methods

    International Nuclear Information System (INIS)

    Binding of immunoreactive radioiodinated human ?-endorphin (125I-?-EP) to rat serum was demonstrated by gel filtration of 125I-?-EP in pooled rat serum on Sephadex G-200. Two radioactive peaks associated with proteins eluted from the column. The first peak eluted at the void volume containing lipoproteins, ?2- and ?2-macroglobulins, and the second peak at the fraction of albumin. Binding of 125I-?-EP to albumin was directly proved by gel filtration of 125I-?-EP in buffer containing 4% human serum albumin on Sephadex G-200. Equilibrium dialysis was not applicable to investigating the interaction of 125I-?-EP with serum proteins, because of the intense nonspecific adsorption to the semi-permeable membrane and the degradation of the peptide during dialysis. Therefore, in order to quantitatively evaluate the binding of 125I-?-EP in sera from rats and humans, the authors utilized four other methods (ultrafiltration, charcoal adsorption, polyethylene glycol precipitation and equilibrium gel filtration). These methods corresponded well with each other and indicated 35-44% binding of 125I-?-EP in rat serum. Binding of 125I-?-EP in normal human serum was 36%, determined by ultrafiltration. Serum protein binding of 125I-?-EP was concentration independent over the concentration range studied (1-1000 nM). 23 references, 4 figures, 1 table 4 figures, 1 table

  3. Mitogen-activated protein kinase signaling pathways promote low-density lipoprotein receptor-related protein 1-mediated internalization of beta-amyloid protein in primary cortical neurons.

    Science.gov (United States)

    Yang, Wei-Na; Ma, Kai-Ge; Qian, Yi-Hua; Zhang, Jian-Shui; Feng, Gai-Feng; Shi, Li-Li; Zhang, Zhi-Chao; Liu, Zhao-Hui

    2015-07-01

    Mounting evidence suggests that the pathological hallmarks of Alzheimer's disease (AD) are caused by the intraneuronal accumulation of beta-amyloid protein (A?). Reuptake of extracellular A? is believed to contribute significantly to the intraneuronal A? pool in the early stages of AD. Published reports have claimed that the low-density lipoprotein receptor-related protein 1 (LRP1) mediates A?1-42 uptake and lysosomal trafficking in GT1-7 neuronal cells and mouse embryonic fibroblast non-neuronal cells. However, there is no direct evidence supporting the role of LRP1 in A? internalization in primary neurons. Our recent study indicated that p38 MAPK and ERK1/2 signaling pathways are involved in regulating ?7 nicotinic acetylcholine receptor (?7nAChR)-mediated A?1-42 uptake in SH-SY5Y cells. This study was designed to explore the regulation of MAPK signaling pathways on LRP1-mediated A? internalization in neurons. We found that extracellular A?1-42 oligomers could be internalized into endosomes/lysosomes and mitochondria in cortical neurons. A?1-42 and LRP1 were also found co-localized in neurons during A?1-42 internalization, and they could form A?1-42-LRP1 complex. Knockdown of LRP1 expression significantly decreased neuronal A?1-42 internalization. Finally, we identified that p38 MAPK and ERK1/2 signaling pathways regulated the internalization of A?1-42 via LRP1. Therefore, these results demonstrated that LRP1, p38 MAPK and ERK1/2 mediated the internalization of A?1-42 in neurons and provided evidence that blockade of LRP1 or inhibitions of MAPK signaling pathways might be a potential approach to lowering brain A? levels and served a potential therapeutic target for AD. PMID:25936756

  4. Amyloid Deposits in Senile Vertebral Arteries, Immunohistological and Ultrastructural Findings

    Directory of Open Access Journals (Sweden)

    S. Doostkam

    2008-01-01

    Full Text Available In a study on amyloid deposits in vertebral arteries, many elderly patients showed amyloid deposits in the perivascular tissue. These proved to be senile systemic amyloidosis of the transthyretin-type by immunohistochemistry. Amyloid deposits were also found in the arterial wall. These intramural amyloid deposits showed significant affinity to elastic material of the arterial wall. The intramural amyloid deposits did not react with any of the known or available antibodies to amyloid subtypes. Only a polyclonal antibody to human elastin could mark this type of amyloid. It may therefore be assumed that the precursor protein of this amyloid is derived from elastin molecules. By electron microscopy, the light microscopic amyloid deposits were of fibrillary structure, typical for amyloid with a direct contact to elastic material.

  5. A panel of regulated proteins in serum from patients with cervical intraepithelial neoplasia and cervical cancer.

    Science.gov (United States)

    Boichenko, Alexander P; Govorukhina, Natalia; Klip, Harry G; van der Zee, A G J; Güzel, Co?kun; Luider, Theo M; Bischoff, Rainer

    2014-11-01

    We developed a discovery-validation mass-spectrometry-based pipeline to identify a set of proteins that are regulated in serum of patients with cervical intraepithelial neoplasia (CIN) and squamous cell cervical cancer using iTRAQ, label-free shotgun, and targeted mass-spectrometric quantification. In the discovery stage we used a "pooling" strategy for the comparative analysis of immunodepleted serum and revealed 15 up- and 26 down-regulated proteins in patients with early- (CES) and late-stage (CLS) cervical cancer. The analysis of nondepleted serum samples from patients with CIN, CES, an CLS and healthy controls showed significant changes in abundance of alpha-1-acid glycoprotein 1, alpha-1-antitrypsin, serotransferrin, haptoglobin, alpha-2-HS-glycoprotein, and vitamin D-binding protein. We validated our findings using a fast UHPLC/MRM method in an independent set of serum samples from patients with cervical cancer or CIN and healthy controls as well as serum samples from patients with ovarian cancer (more than 400 samples in total). The panel of six proteins showed 67% sensitivity and 88% specificity for discrimination of patients with CIN from healthy controls, a stage of the disease where current protein-based biomarkers, for example, squamous cell carcinoma antigen (SCCA), fail to show any discrimination. Additionally, combining the six-protein panel with SCCA improves the discrimination of patients with CES and CLS from healthy controls. PMID:25232869

  6. New Insight into Toxicity due to Oligomeric Amyloid-b Peptide and Alpha-Synuclein Protein induced by Copper(II ion

    Directory of Open Access Journals (Sweden)

    Keita Abe

    2015-07-01

    Full Text Available It is well known that Alzheimer’s and Parkinson’ s diseases are closely related with the aggregated forms of amyloid-b peptide and alpha-synuclein (a-syn protein, respectively, and the recent work shows that neurotoxicity due to oligomeric a-syn requires the presence of copper but not iron ion. In this article, we have proposed the new insight into the toxicity due to the oligomeric amyloid-b peptide and a-syn protein induced by the copper(II ion, based on the Nishida Reaction which is specific for the binuclear copper(II compounds where two copper(II ions are in close vicinity, and that copper(II-peroxide adduct exhibits strong electrophilicity towards several organic compounds similar to the singlet oxygen (1Dg.

  7. Quantification of gamma-secretase modulation differentiates inhibitor compound selectivity between two substrates Notch and amyloid precursor protein

    Directory of Open Access Journals (Sweden)

    Yang Ting

    2008-11-01

    Full Text Available Abstract Background Deposition of amyloid-? protein (A? is a major pathological hallmark of Alzheimer's disease (AD. A? is generated from ?-secretase cleavage of amyloid precursor protein (APP. In addition to APP, ?-secretase also cleaves other type I integral membrane proteins, including the Notch receptor, a key molecule involved in embryonic development. Results To explore selective ?-secretase inhibitors, a combination of five methods was used to systematically determine these inhibitors' profiles on the ?-secretase cleavage of APP and Notch. When two potent ?-secretase inhibitors, compound E (cpd E and DAPT, were used in a conventional in vitro ?-secretase activity assay, cpd E completely blocked A? generation from the cleavage of substrate APP C100, but only had a minor effect on Notch cleavage and NICD generation. Next, cpd E and DAPT were applied to HEK293 cells expressing a truncated Notch substrate Notch?E. Both cpd E and DAPT were more potent in blocking A? generation than NICD generation. Third, a reporter construct was created that carried the NICD targeting promoter with three Su(H binding sequences followed by the luciferase gene. We found that the inhibition of NICD generation by cpd E and DAPT was consistent with the reduced expression of luciferase gene driven by this Notch targeting promoter. Fourth, levels of "Notch-A?-like" (N?* peptide derived from two previously reported chimeric APP with its transmembrane domain or the juxtamembrane portion replaced by the Notch sequence were quantified. Measurement of N?* peptides by ELISA confirmed that EC50's of cpd E were much higher for N?* than A?. Finally, the expression levels of Notch target gene her6 in cpd E or DAPT-treated zebrafish were correlated with the degree of tail curvature due to defective somitogenesis, a well characterized Notch phenotype in zebrafish. Conclusion Our ELISA-based quantification of A? and N?* in combination with the test in zebrafish provides a novel approach for efficient cell-based screening and in vivo validation of APP selective ?-secretase inhibitors.

  8. Usefulness of labeled human serum albumin scintigraphy in protein-losing enteropathy: report of two cases

    International Nuclear Information System (INIS)

    Protein-losing enteropathy is a rare entity characterized by a gastrointestinal loss of proteins. 99mTc-labeled Human Serum Albumin Scintigraphy is a simple exam that can detect these losses. We report two cases in which scintigraphy proved an important input. (authors)

  9. Effects of supraphysiologic temperature and broth dilution on serum protein binding.

    OpenAIRE

    White, R. L.; Kays, M. B.; Armstrong, T. A.; Friedrich, L. V.

    1990-01-01

    Pooled human serum, unheated (UU) and heated at 56 degrees C for 30 (HU-30) and 60 (HU-60) min, and these media diluted 1:1 with broth (UD, HD-30, and HD-60) were used to assess the effects of supraphysiologic temperature and broth dilution on the serum protein binding of ceftriaxone. Protein binding was determined by ultrafiltration and subsequent high-performance liquid chromatography analysis. Significant differences in protein binding between UU and HU-60 (P less than 0.05), UD and HD-60 ...

  10. Methyl-binding domain protein-based DNA isolation from human blood serum combines DNA analyses and serum-autoantibody testing

    Directory of Open Access Journals (Sweden)

    Jungbauer Christof

    2011-09-01

    Full Text Available Abstract Background Circulating cell free DNA in serum as well as serum-autoantibodies and the serum proteome have great potential to contribute to early cancer diagnostics via non invasive blood tests. However, most DNA preparation protocols destroy the protein fraction and therefore do not allow subsequent protein analyses. In this study a novel approach based on methyl binding domain protein (MBD is described to overcome the technical difficulties of combining DNA and protein analysis out of one single serum sample. Methods Serum or plasma samples from 98 control individuals and 54 breast cancer patients were evaluated upon silica membrane- or MBD affinity-based DNA isolation via qPCR targeting potential DNA methylation markers as well as by protein-microarrays for tumor-autoantibody testing. Results In control individuals, an average DNA level of 22.8 ± 25.7 ng/ml was detected applying the silica membrane based protocol and 8.5 ± 7.5 ng/ml using the MBD-approach, both values strongly dependent on the serum sample preparation methods used. In contrast to malignant and benign tumor serum samples, cell free DNA concentrations were significantly elevated in sera of metastasizing breast cancer patients. Technical evaluation revealed that serum upon MBD-based DNA isolation is suitable for protein-array analyses when data are consistent to untreated serum samples. Conclusion MBD affinity purification allows DNA isolations under native conditions retaining the protein function, thus for example enabling combined analyses of DNA methylation and autoantigene-profiles from the same serum sample and thereby improving minimal invasive diagnostics.

  11. Luteolin Isolated from the Medicinal Plant Elsholtzia rugulosa (Labiatae) Prevents Copper-Mediated Toxicity in ?-Amyloid Precursor Protein Swedish Mutation Overexpressing SH-SY5Y Cells

    OpenAIRE

    Guanhua Du; Lin Li; Xi Lan; Hailin Qin; Li De Zhang; Ailin Liu; Fanrui Meng; Rui Liu

    2011-01-01

    Luteolin, a 3’,4’,5,7-tetrahydroxyflavone, is a plant flavonoid and pharmacologically active agent that has been isolated from several plant species. In the present study, the effects of luteolin obtained from the medicinal plant Elsholtzia rugulosa and the related mechanisms were examined in an Alzheimer's disease (AD) cell model. In this model, copper was used to exacerbate the neurotoxicity in ?-amyloid precursor protein Swedish mutation stably overexpressed SH-SY5Y cells (named “AP...

  12. Lewy Bodies Contain Altered ?-Synuclein in Brains of Many Familial Alzheimer’s Disease Patients with Mutations in Presenilin and Amyloid Precursor Protein Genes

    OpenAIRE

    Lippa, Carol F.; Fujiwara, Hideo; Mann, David M. A.; Giasson, Benoit; Baba, Minami; Schmidt, Marie L.; Nee, Linda E.; O’connell, Brendan; Pollen, Dan A.; St George-hyslop, Peter; Ghetti, Bernardino; Nochlin, David; Bird, Thomas D.; Cairns, Nigel J.; Lee, Virginia M. -y

    1998-01-01

    Missense mutations in the ?-synuclein gene cause familial Parkinson’s disease (PD), and ?-synuclein is a major component of Lewy bodies (LBs) in sporadic PD, dementia with LBs (DLB), and the LB variant of Alzheimer’s disease (AD). To determine whether ?-synuclein is a component of LBs in familial AD (FAD) patients with known mutations in presenilin (n = 65) or amyloid precursor protein (n = 9) genes, studies were conducted with antibodies to ?-, ?-, and ?-synuclein. LBs were detecte...

  13. Amyloid precursor-like protein 2 suppresses irradiation-induced apoptosis in Ewing sarcoma cells and is elevated in immune-evasive Ewing sarcoma cells

    OpenAIRE

    Peters, Haley L; Yan, Ying; Nordgren, Tara M.; Cutucache, Christine E; Joshi, Shantaram S.; Solheim, Joyce C.

    2013-01-01

    Despite surgery, chemotherapy, and radiotherapy treatments, the children, adolescents, and young adults who are diagnosed with metastasized Ewing sarcoma face a dismal prognosis. Amyloid precursor-like protein 2 (APLP2) has recently been implicated in the survival of cancer cells and in our current study, APLP2’s contribution to the survival of Ewing sarcoma cells was examined. APLP2 was readily detected in all Ewing sarcoma cell lines analyzed by western blotting, with the TC71 Ewing sarco...

  14. Heparan sulphate proteoglycan and the low-density lipoprotein receptor-related protein 1 constitute major pathways for neuronal amyloid-? uptake

    OpenAIRE

    Kanekiyo, Takahisa; ZHANG, JUAN; Liu, Qiang; Liu, Chia-Chen; Zhang, Lijuan; Bu, Guojun

    2011-01-01

    Alzheimer’s disease (AD) is a progressive and irreversible neurodegenerative disorder in which the aggregation and deposition of amyloid-? (A?) peptides in the brain are central to its pathogenesis. In healthy brains, A? is effectively metabolized with little accumulation. Cellular uptake and subsequent degradation of A? is one of the major pathways for its clearance in the brain. Increasing evidence has demonstrated significant roles for the low-density lipoprotein receptor-related protein 1...

  15. Low-density lipoprotein receptor-related protein-1 : a serial clearance homeostatic mechanism controlling Alzheimer's amyloid ?-peptide elimination from the brain

    OpenAIRE

    Zlokovic, Berislav V.; Deane, Rashid; Sagare, Abhay P.; Bell, Robert D; Winkler, Ethan A

    2010-01-01

    Low-density lipoprotein receptor-related protein-1 (LRP1), a member of the LDL receptor family, has major roles in the cellular transport of cholesterol, endocytosis of forty structurally diverse ligands, transcytosis of ligands across the blood-brain barrier, and transmembrane and nuclear signaling. Recent evidence indicates that LRP1 regulates brain and systemic clearance of Alzheimer's disease (AD) amyloid ?-peptide (A?). According to the two hit vascular hypothesis for AD, vascular damage...

  16. Chronic treatment with amyloid beta(1-42) inhibits non-cholinergic high-affinity choline transport in NG108-15 cells through protein kinase C signaling.

    Czech Academy of Sciences Publication Activity Database

    Nováková, Jana; Mikasová, Lenka; Machová, Eva; Lisá, V?ra; Doležal, Vladimír

    2005-01-01

    Ro?. 1062, ?. 1-2 (2005), s. 101-110. ISSN 0006-8993 R&D Projects: GA AV ?R(CZ) IAA5011206; GA MŠk(CZ) LC554 Grant ostatní: Lipidiet(XE) QLK1-CT-2002-00172 Institutional research plan: CEZ:AV0Z50110509 Keywords : choline transporter * beta-amyloid * protein kinase C Subject RIV: ED - Physiology Impact factor: 2.296, year: 2005

  17. The Functions of the Amyloid Precursor Protein Gene and Its Derivative Peptides: I Molecular Biology and Metabolic Processing

    Directory of Open Access Journals (Sweden)

    Emily R Atkins

    2011-06-01

    Full Text Available The amyloid precursor protein gene (APP and its derivative peptides have important functions in the central nervous system. APP and A? fulfil criteria as neuractive peptides: presence, release and identity of action. A? is a peptide of 1 - 43 amino acids in length, derived from APP and the major component of the core of neuritic plaques found in Alzheimer’s disease. Analysis of the cDNA of A? revealed its origins from the larger precursor protein. There are at least four types of mRNA generated by alternative splicing of exons 7 and 8. Exon 7 encodes a 57 amino acid sequence found in the extracellular domain with major homology to the Kunitz-type of serine protease inhibitors. APP is cleaved by three secretases known as ?, ?, and ? secretase which act on APP at different sites producing various fragments of differing amino acid length. The ? secretase is a macromolecular enzyme complex composed of presenilin 1, 2 and other molecular constitutents essential for its function.

  18. Detection of ?-amyloid peptide (1–16) and amyloid precursor protein (APP770) using spectroscopic ellipsometry and QCM techniques: A step forward towards Alzheimers disease diagnostics

    OpenAIRE

    Mustafa, M. K.; Nabok, Alexei; Parkinson, D.; Tothill, Ibtisam E.; Salam, Faridah; Tsargorodskaya, A.

    2010-01-01

    A highly sensitive method of spectroscopic ellipsometry in total internal reflection mode (TIRE) was exploited for detecting ?-amyloid peptide (A?1–16) in the direct immune reaction with monoclonal DE2 antibodies (raised against A?1–16) electrostatically immobilised on the surface of gold. A rapid detection of A?1–16 in a wide range of concentrations from 5 ?g/ml down to 0.05 ng/ml was achieved using a cost-effective and label-free direct immunoassay format. TIRE dynamic spectral m...

  19. Dynamics of Abeta turnover and deposition in different beta-amyloid precursor protein transgenic mouse models following gamma-secretase inhibition.

    Science.gov (United States)

    Abramowski, Dorothee; Wiederhold, Karl-Heinz; Furrer, Ulrich; Jaton, Anne-Lise; Neuenschwander, Anton; Runser, Marie-Josephine; Danner, Simone; Reichwald, Julia; Ammaturo, Domenico; Staab, Dieter; Stoeckli, Markus; Rueeger, Heinrich; Neumann, Ulf; Staufenbiel, Matthias

    2008-11-01

    Human beta-amyloid precursor protein (APP) transgenic mice are commonly used to test potential therapeutics for Alzheimer's disease. We have characterized the dynamics of beta-amyloid (Abeta) generation and deposition following gamma-secretase inhibition with compound LY-411575 [N(2)-[(2S)-2-(3,5-difluorophenyl)-2-hydroxyethanoyl]-N(1)-[(7S)-5-methyl-6-oxo-6,7-dihydro-5H-dibenzo[b,d]azepin-7-yl]-L-alaninamide]. Kinetic studies in preplaque mice distinguished a detergent-soluble Abeta pool in brain with rapid turnover (half-lives for Abeta40 and Abeta42 were 0.7 and 1.7 h) and a much more stable, less soluble pool. Abeta in cerebrospinal fluid (CSF) reflected the changes in the soluble brain Abeta pool, whereas plasma Abeta turned over more rapidly. In brain, APP C-terminal fragments (CTF) accumulated differentially. The half-lives for gamma-secretase degradation were estimated as 0.4 and 0.1 h for C99 and C83, respectively. Three different APP transgenic lines responded very similarly to gamma-secretase inhibition regardless of the familial Alzheimer's disease mutations in APP. Amyloid deposition started with Abeta42, whereas Abeta38 and Abeta40 continued to turn over. Chronic gamma-secretase inhibition lowered amyloid plaque formation to a different degree in different brain regions of the same mice. The extent was inversely related to the initial amyloid load in the region analyzed. No evidence for plaque removal below baseline was obtained. gamma-Secretase inhibition led to a redistribution of intracellular Abeta and an elevation of CTFs in neuronal fibers. In CSF, Abeta showed a similar turnover as in preplaque animals demonstrating its suitability as marker of newly generated, soluble Abeta in plaque-bearing brain. This study supports the use of APP transgenic mice as translational models to characterize Abeta-lowering therapeutics. PMID:18687920

  20. Formation of toxic peptides in irradiated rats and binding thereof with blood serum proteins

    International Nuclear Information System (INIS)

    Whole-body ?-irradiation of rats with a dose of 9.0 Gy caused a 1.5-fold and a 5-fold increase in excretion of bas peptides (molecular mass of 500-2000) in urea on the 2nd and 5th postirradiation days, respectively. These peptides possessed toxic activity and ability to form complexes with macroglobulins, immunoglobulins, and blood serum albumins, in particular. Irradiation decreased binding ability of serum proteins, and preliminary washing thereof by ultrafiltration increased it

  1. Serum beta 2-microglobulin and C reactive protein concentrations in viral infections.

    OpenAIRE

    Cooper, E. H.; Forbes, M. A.; Hambling, M. H.

    1984-01-01

    Serum beta 2-microglobulin concentrations were assayed in a number of virus diseases. Infectious mononucleosis, cytomegalovirus, and influenza A were associated with pronounced increases in serum beta 2-microglobulin concentration. Smaller increases, with values generally less than 4 mg/l, were noted in other viral infections. Apart from in acute influenza A, the C reactive protein and beta 2-microglobulin responses were not associated.

  2. The binding of 3-14C-coumarin to serum proteins in the rat

    International Nuclear Information System (INIS)

    Like other benzo-pyrones, coumarin has been shown to bind to serum proteins. Under in vitro conditions irrespective of the dose which ranged from 50 - 3.31 ?gm/ml, the binding remained at approximately 40%. With intravenous administration an average of 4.6% was bound to serum albumins. With intraperitoneal administration an average of 4% of the injected dose entered the vascular system of which an average of 36.5% was in the bound form. (author)

  3. Association between serum levels of C-reactive protein and personality traits in women

    OpenAIRE

    Anckarsäter Henrik; Landén Mikael; Holm Göran; Rosmond Roland; Baghaei Fariba; Henningsson Susanne; Ekman Agneta

    2008-01-01

    Abstract Background While low-grade inflammation has consistently been observed in subjects with depression, studies on the possible relationship between inflammation and other aspects of brain function are as yet sparse. In this study, we aimed to investigate the possible association between serum levels of the inflammation marker C-reactive protein (CRP) and personality traits. Methods In this study, serum levels of high-sensitivity CRP were determined by ELISA in a population of 270 42-yea...

  4. Intraoperative values of S-100 protein, myelin basic protein, lactate, and albumin in the CSF and serum of neurosurgical patients

    OpenAIRE

    Vries, J.; Thijssen, W.; Snels, S.; Menovsky, T.; Peer, N.; Lamers, K.

    2001-01-01

    OBJECTIVES—To assess the concentrations of S-100 protein, myelin basic protein (MBP), and lactate, and the (CSF)/serum albumin ratio (Qalb) during intracranial neurosurgical procedures.?METHODS—Samples of CSF from 91 patients with various CNS diseases were obtained by aspiration of cisternal CSF at the beginning of surgery (before starting surgical manipulation of the brain) and concentrations of S-100 protein, MBP, and lactate, and Qalb were determined. At the same...

  5. Kinetic studies on beta-site amyloid precursor protein-cleaving enzyme (BACE). Confirmation of an iso mechanism.

    Science.gov (United States)

    Toulokhonova, Larisa; Metzler, William J; Witmer, Mark R; Copeland, Robert A; Marcinkeviciene, Jovita

    2003-02-14

    The steady-state kinetic mechanism of beta-amyloid precursor protein-cleaving enzyme (BACE)-catalyzed proteolytic cleavage was evaluated using product and statine- (Stat(V)) or hydroxyethylene-containing (OM99-2) peptide inhibition data, solvent kinetic isotope effects, and proton NMR spectroscopy. The noncompetitive inhibition pattern observed for both cleavage products, together with the independence of Stat(V) inhibition on substrate concentration, suggests a uni-bi-iso kinetic mechanism. According to this mechanism, the enzyme undergoes multiple conformation changes during the catalytic cycle. If any of these steps are rate-limiting to turnover, an enzyme form preceding the rate-limiting conformational change should accumulate. An insignificant solvent kinetic isotope effect (SKIE) on k(cat)/K(m), a large inverse solvent kinetic isotope effect on k(cat), and the absence of any SKIE on the inhibition onset by Stat(V) during catalysis together indicate that the rate-limiting iso-step occurs after formation of a tetrahedral intermediate. A moderately short and strong hydrogen bond (at delta 13.0 ppm and phi of 0.6) has been observed by NMR spectroscopy in the enzyme-hydroxyethylene peptide (OM99-2) complex that presumably mimics the tetrahedral intermediate of catalysis. Collapse of this intermediate, involving multiple steps and interconversion of enzyme forms, has been suggested to impose a rate limitation, which is manifested in a significant SKIE on k(cat). Multiple enzyme forms and their distribution during catalysis were evaluated by measuring the SKIE on the noncompetitive (mixed) inhibition constants for the C-terminal reaction product. Large, normal SKIE values were observed for these inhibition constants, suggesting that both kinetic and thermodynamic components contribute to the K(ii) and K(is) expressions, as has been suggested for other iso-mechanism featuring enzymes. We propose that a conformational change related to the reprotonation of aspartates during or after the bond-breaking event is the rate-limiting segment in the catalytic reaction of beta-amyloid precursor protein-cleaving enzyme, and ligands binding to other than the ground-state forms of the enzyme might provide inhibitors of greater pharmacological relevance. PMID:12458195

  6. Serum eosinophil granule proteins predict asthma risk in allergic rhinitis

    OpenAIRE

    Nielsen, Lars Peter; Peterson, Christer; Dahl, Ronald

    2009-01-01

    Background: Allergic rhinitis is a common disease, in which some patients will deteriorate or develop asthma. It is important to characterize these patients, thereby offering the possibility for prevention. This study evaluated eosinophil parameters as potential indicators of deteriorating allergic airway disease. Methods: The subjects of the study included all patients who suffered seasonal allergic rhinitis and had participated in a study 6 years earlier, in which blood eosinophils, serum e...

  7. Serum Protein Profiles of Juvenile Ring-Necked Pheasants Vaccinated or Not Against Newcastle Disease

    OpenAIRE

    Elizabeth Moreira dos Santos Schmidt; Antonio Carlos Paulillo; Rosangela Locatelli-Dittrich; Olair Beltrame; Janine Denadai

    2009-01-01

    The aim of this study was to investigate blood protein parameters in juvenile ring-necked pheasants using Ulster 2C, B1 and LaSota vaccines strains of the Newcastle disease virus. Total serum protein, albumin and globulin concentrations showed significantly differences among vaccinated and non-vaccinated birds. Significant variations were not observed in the analyses in relation to protein electrophoresis profile.

  8. Interaction of the amyloid precursor protein-like protein 1 (APLP1) E2 domain with heparan sulfate involves two distinct binding modes

    Energy Technology Data Exchange (ETDEWEB)

    Dahms, Sven O., E-mail: sdahms@fli-leibniz.de [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany); Mayer, Magnus C. [Freie Universität Berlin, Thielallee 63, 14195 Berlin (Germany); Miltenyi Biotec GmbH, Robert-Koch-Strasse 1, 17166 Teterow (Germany); Roeser, Dirk [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany); Multhaup, Gerd [McGill University Montreal, Montreal, Quebec H3G 1Y6 (Canada); Than, Manuel E., E-mail: sdahms@fli-leibniz.de [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany)

    2015-03-01

    Two X-ray structures of APLP1 E2 with and without a heparin dodecasaccharide are presented, revealing two distinct binding modes of the protein to heparan sulfate. The data provide a mechanistic explanation of how APP-like proteins bind to heparan sulfates and how they specifically recognize nonreducing structures of heparan sulfates. Beyond the pathology of Alzheimer’s disease, the members of the amyloid precursor protein (APP) family are essential for neuronal development and cell homeostasis in mammals. APP and its paralogues APP-like protein 1 (APLP1) and APP-like protein 2 (APLP2) contain the highly conserved heparan sulfate (HS) binding domain E2, which effects various (patho)physiological functions. Here, two crystal structures of the E2 domain of APLP1 are presented in the apo form and in complex with a heparin dodecasaccharide at 2.5 Å resolution. The apo structure of APLP1 E2 revealed an unfolded and hence flexible N-terminal helix ?A. The (APLP1 E2){sub 2}–(heparin){sub 2} complex structure revealed two distinct binding modes, with APLP1 E2 explicitly recognizing the heparin terminus but also interacting with a continuous heparin chain. The latter only requires a certain register of the sugar moieties that fits to a positively charged surface patch and contributes to the general heparin-binding capability of APP-family proteins. Terminal binding of APLP1 E2 to heparin specifically involves a structure of the nonreducing end that is very similar to heparanase-processed HS chains. These data reveal a conserved mechanism for the binding of APP-family proteins to HS and imply a specific regulatory role of HS modifications in the biology of APP and APP-like proteins.

  9. Interaction of the amyloid precursor protein-like protein 1 (APLP1) E2 domain with heparan sulfate involves two distinct binding modes

    International Nuclear Information System (INIS)

    Two X-ray structures of APLP1 E2 with and without a heparin dodecasaccharide are presented, revealing two distinct binding modes of the protein to heparan sulfate. The data provide a mechanistic explanation of how APP-like proteins bind to heparan sulfates and how they specifically recognize nonreducing structures of heparan sulfates. Beyond the pathology of Alzheimer’s disease, the members of the amyloid precursor protein (APP) family are essential for neuronal development and cell homeostasis in mammals. APP and its paralogues APP-like protein 1 (APLP1) and APP-like protein 2 (APLP2) contain the highly conserved heparan sulfate (HS) binding domain E2, which effects various (patho)physiological functions. Here, two crystal structures of the E2 domain of APLP1 are presented in the apo form and in complex with a heparin dodecasaccharide at 2.5 Å resolution. The apo structure of APLP1 E2 revealed an unfolded and hence flexible N-terminal helix ?A. The (APLP1 E2)2–(heparin)2 complex structure revealed two distinct binding modes, with APLP1 E2 explicitly recognizing the heparin terminus but also interacting with a continuous heparin chain. The latter only requires a certain register of the sugar moieties that fits to a positively charged surface patch and contributes to the general heparin-binding capability of APP-family proteins. Terminal binding of APLP1 E2 to heparin specifically involves a structure of the nonreducing end that is very similar to heparanase-processed HS chains. These data reveal a conserved mechanism for the binding of APP-family proteins to HS and imply a specific regulatory role of HS modifications in the biology of APP and APP-like proteins

  10. ANTIAMNESIC POTENTIAL OF SOLASODINE AGAINST ?-AMYLOID PROTEIN INDUCED AMNESIA IN MICE

    Directory of Open Access Journals (Sweden)

    Desai Alpesh B

    2011-05-01

    Full Text Available Alzheimer’s disease (AD, the most common form of dementia in the elderly population, is characterized by an insidious onset with memory impairment and an inexorable progression of cognitive decline. Nootropic agents are a heterogeneous groups of drugs developed for use in dementia and other cerebral disorders. Nootropics agents are being primarily used to improve memory, mood and behavior. However, the resulting adverse effects associated with these agents have limited their use. Therefore, it is worthwhile to explore the utility of traditional medicines for the treatment of various cognitive disorders. The present study was undertaken to assess the potential of solasodine on ?-amyloid induced amnesia in mice. Elevated plus maze (EPM and Morris water maze (MWM was employed to evaluate learning and memory parameters. Piracetam was used as the standard drug. Solasodine (1, 2 and 4 mg/kg, p.o. was screened for claimed potential in mice. Solasodine improved both short term memory and long term memory when assessed on Elevated pluz maze and Morris Water maze respectively. Hence, solasodine might prove to be a useful memory restorative agent in the treatment of dementia seen in the Alzheimer’s disease.

  11. Binding of colchicine and thiocolchicoside to human serum proteins and blood cells.

    Science.gov (United States)

    Sabouraud, A; Chappey, O; Dupin, T; Scherrmann, J M

    1994-08-01

    The binding of 3H-colchicine and its derivative 3H-thiocolchicoside to human serum, purified human proteins and blood cells was studied by equilibrium dialysis and centrifugation. Binding of colchicine and thiocolchicoside to human serum was 38.9 C +/- 4.7 and 12.8 C +/- 5.3%, respectively, essentially to albumin. Protein binding was not dependent on the concentration of either drug between 10(-10) and 10(-5)M. The binding of colchicine and thiocolchicoside to isolated erythrocytes (55 C +/- 5.6 and 16.5 C +/- 2.1%, respectively) decreased markedly in the presence of human serum proteins, i.e. in whole blood (38.7 C +/- 3.1 and 3.4 C +/- 0.8%). Binding of colchicine and thiocolchicoside to other blood cells was very low C thiocolchicoside to be pharmacokinetically sensitive to binding displacement by drug interactions. PMID:7981928

  12. Effect of acute irradiation on concentration of some serum proteins in rats

    International Nuclear Information System (INIS)

    The relationship is studied between the effect of a dose of acute radiation and the character of changes in the serum concentration of prealbumin, albumin, A1-globulin, A1-macroglobulin (A1M), transferrin, IgG, haptoglobin, ceruloplasmin, hemopexin, and the C3 component of the complement in rats irradiated with doses of 1.91 Gy, 5.26 Gy and 6.70 Gy. With proteins whose serum concentrations decreased, the decrease was found to depend on the radiation dose. With proteins whose concentration increased as a result of irradiation, the most effective dose was found to be 5.26 Gy. The concentration of the serum proteins was determined by 2-dimensional immunoelectrophoresis. (author)

  13. The use of proteomics in identifying differentially expressed serum proteins in humans with type 2 diabetes

    Directory of Open Access Journals (Sweden)

    Göransson Michael

    2006-12-01

    Full Text Available Abstract Background The aim of the study was to optimize protocols for finding and identifying serum proteins that are differentially expressed in persons with normal glucose tolerance (NGT compared to individuals with type 2 diabetes mellitus (T2DM. Serum from persons with NGT and persons with T2DM was profiled using ProteinChip arrays and time-of-flight mass spectra were generated by surface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS. Results Mass spectra from NGT- and T2DM-groups were compared. Fifteen proteins ranging from 5 to 79 kDa were differentially expressed (p Conclusion Protocols for protein profiling by SELDI-TOF MS and protein identification by fractionation, SDS-PAGE and PMF were optimized for serum from humans with T2DM. With these protocols differentially expressed proteins were discovered and identified when serum from NGT- and T2DM-individuals was analyzed.

  14. Metabolism of homologous and heterologous serum proteins in garter snakes (Thamnophis ordinoides)

    International Nuclear Information System (INIS)

    The half-life (Tsub(1/2) of serum immunoglobulin (Ig) and albumin from snakes and mammals were determined in both garter snakes (Thamnophis ordinoides) and mice (Mus musculus). Metabolism of serum proteins in snakes was similar to mammalian protein metabolism in that homologous serum albumin had shorter Tsub(1/2) (16 days) than IgG (38 days). Also, reptilian and mammalian serum proteins had a relatively longer Tsub(1/2) when injected into closely related species. Thus mammalian serum Ig (rabbit gamma globulin (RGG)) had a shorter Tsub(1/2) (6.3 days) in snake than did homologous snake IgG (38 days), whereas in mice, RGG had a longer Tsub(1/2) (3.8 days) than snake Ig (0.9 days). Differences between metabolism of homologous and heterologous albumins were apparent only in snakes in which the Tsub(1/2) of homologous albumin was approximately 8-fold greater than mammalian albumin. These results indicate that metabolism of both Ig and albumin in snakes is regulated by specific receptors whereas albumin receptors have been difficult to demonstrate in mammals. The results of this study suggest that one of the factors determining the metabolism of a protein is its foreignness to the host perhaps because of receptor cross reactions. (author)

  15. Identification of a Compound That Disrupts Binding of Amyloid-? to the Prion Protein Using a Novel Fluorescence-based Assay.

    Science.gov (United States)

    Risse, Emmanuel; Nicoll, Andrew J; Taylor, William A; Wright, Daniel; Badoni, Mayank; Yang, Xiaofan; Farrow, Mark A; Collinge, John

    2015-07-01

    The prion protein (PrP) has been implicated both in prion diseases such as Creutzfeldt-Jakob disease, where its monomeric cellular isoform (PrP(C)) is recruited into pathogenic self-propagating polymers of misfolded protein, and in Alzheimer disease, where PrP(C) may act as a receptor for synaptotoxic oligomeric forms of amyloid-? (A?). There has been considerable interest in identification of compounds that bind to PrP(C), stabilizing its native fold and thereby acting as pharmacological chaperones to block prion propagation and pathogenesis. However, compounds binding PrP(C) could also inhibit the binding of toxic A? species and may have a role in treating Alzheimer disease, a highly prevalent dementia for which there are currently no disease-modifying treatments. However, the absence of a unitary, readily measurable, physiological function of PrP makes screening for ligands challenging, and the highly heterogeneous nature of A? oligomer preparations makes conventional competition binding assays difficult to interpret. We have therefore developed a high-throughput screen that utilizes site-specifically fluorescently labeled protein to identify compounds that bind to PrP and inhibit both A? binding and prion propagation. Following a screen of 1,200 approved drugs, we identified Chicago Sky Blue 6B as the first small molecule PrP ligand capable of inhibiting A? binding, demonstrating the feasibility of development of drugs to block this interaction. The interaction of Chicago Sky Blue 6B was characterized by isothermal titration calorimetry, and its ability to inhibit A? binding and reduce prion levels was established in cell-based assays. PMID:25995455

  16. Effect of chemical radioprotectors on serum proteins of rats exposed to gamma radiation

    International Nuclear Information System (INIS)

    Paper electrophoretic fractions of serum proteins were made prior to and at one, three and seven days following exposure of rats to 11 Gy whole body ?-irradiation. During the early period of post irradiation an increase in total proteins, a decrease in albumin and ?-globulin fractions and an increase in ? and ? fractions were observed. On 7th day of post irradiation there was a drastic decrease in total proteins and in all the fractions. Similar studies carried out in animals pre-treated with radioprotectors (other than MPG) reduced considerably the changes produced by ?-irradiation on total proteins and its fractions except albumin. Combinations of ?-mercaptropropionylglycine + 5-hydroxy L-tryptophan (5-HTP), 2-aminoethylisothiuronium bromide hydrochloride (AET) + 5-HTP and the optimum dose of AET (200 mg/kg body wt.) gave better protection than 5-HTP and accelerated the normalisation process of the altered serum proteins of irradiated rats. (author)

  17. Tracer diffusion coefficients of proteins by means of holographic relaxation spectroscopy: application to bovine serum albumin

    International Nuclear Information System (INIS)

    Holographic relaxation spectroscopy has been used to measure tracer diffusion coefficients for photochromically labeled bovine serum albumin in solutions having total bovine serum albumin concentrations in the range 3.25 to 257 g/liter. In the limit of zero concentration, the diffusion coefficient was found to be 5.9 X 10(-7) cm2/s and the initial slope was zero. The concentration dependence of the diffusion coefficient was not significantly affected by the fraction of protein molecules which were labeled. Holographic relaxation spectroscopy permits rapid, accurate determination of tracer diffusion coefficients for proteins in mixtures

  18. Inhibition of AMP-activated protein kinase signaling alleviates impairments in hippocampal synaptic plasticity induced by amyloid ?.

    Science.gov (United States)

    Ma, Tao; Chen, Yiran; Vingtdeux, Valerie; Zhao, Haitian; Viollet, Benoit; Marambaud, Philippe; Klann, Eric

    2014-09-01

    The AMP-activated protein kinase (AMPK) is a Ser/Thr kinase that is activated in response to low-energy states to coordinate multiple signaling pathways to maintain cellular energy homeostasis. Dysregulation of AMPK signaling has been observed in Alzheimer's disease (AD), which is associated with abnormal neuronal energy metabolism. In the current study we tested the hypothesis that aberrant AMPK signaling underlies AD-associated synaptic plasticity impairments by using pharmacological and genetic approaches. We found that amyloid ? (A?)-induced inhibition of long-term potentiation (LTP) and enhancement of long-term depression were corrected by the AMPK inhibitor compound C (CC). Similarly, LTP impairments in APP/PS1 transgenic mice that model AD were improved by CC treatment. In addition, A?-induced LTP failure was prevented in mice with genetic deletion of the AMPK ?2-subunit, the predominant AMPK catalytic subunit in the brain. Furthermore, we found that eukaryotic elongation factor 2 (eEF2) and its kinase eEF2K are key downstream effectors that mediate the detrimental effects of hyperactive AMPK in AD pathophysiology. Our findings describe a previously unrecognized role of aberrant AMPK signaling in AD-related synaptic pathophysiology and reveal a potential therapeutic target for AD. PMID:25186765

  19. Effects of yokukansan and donepezil on learning disturbance and aggressiveness induced by intracerebroventricular injection of amyloid ? protein in mice.

    Science.gov (United States)

    Sekiguchi, Kyoji; Imamura, Sachiko; Yamaguchi, Takuji; Tabuchi, Masahiro; Kanno, Hitomi; Terawaki, Kiyoshi; Kase, Yoshio; Ikarashi, Yasushi

    2011-04-01

    The effects of yokukansan and donepezil on learning disturbance and aggressiveness were examined in amyloid ? protein (A?)-injected mice. Intellicage tests showed that both yokukansan and donepezil ameliorated A?-induced learning disturbance, but the ameliorating effect of donepezil was not enhanced by concomitant administration of yokukansan. On the other hand, a social interaction test showed that A?-induced aggressiveness was ameliorated by yokukansan, but not by donepezil. Co-administration of both drugs also ameliorated aggressiveness, as did yokukansan alone. In vitro binding assays revealed that yokukansan did not bind to choline receptors or transporters. In vitro enzyme assays revealed that yokukansan did not affect choline acetyltransferase activity or inhibit acetylcholinesterase activity, as did donepezil. These results suggest that yokukansan might ameliorate aggressiveness without interfering with the pharmacological efficacy (antidementia effect) of donepezil and also that concomitant administration of yokukansan might be useful for amelioration of aggressiveness, which was not lessened by donepezil. The difference in the efficacies of both drugs may be due to a difference in their pharmacological mechanisms. PMID:20803480

  20. The Kunitz-protease inhibitor domain in amyloid precursor protein reduces cellular mitochondrial enzymes expression and function.

    Science.gov (United States)

    Chua, Li-Min; Lim, Mei-Li; Wong, Boon-Seng

    2013-08-01

    Mitochondrial dysfunction is a prominent feature of Alzheimer's disease (AD) and this can be contributed by aberrant metabolic enzyme function. But, the mechanism causing this enzymatic impairment is unclear. Amyloid precursor protein (APP) is known to be alternatively spliced to produce three major isoforms in the brain (APP695, APP751, APP770). Both APP770 and APP751 contain the Kunitz Protease Inhibitory (KPI) domain, but the former also contain an extra OX-2 domain. APP695 on the other hand, lacks both domains. In AD, up-regulation of the KPI-containing APP isoforms has been reported. But the functional contribution of this elevation is unclear. In the present study, we have expressed and compared the effect of the non-KPI containing APP695 and the KPI-containing APP751 on mitochondrial function. We found that the KPI-containing APP751 significantly decreased the expression of three major mitochondrial metabolic enzymes; citrate synthase, succinate dehydrogenase and cytochrome c oxidase (COX IV). This reduction lowers the NAD(+)/NADH ratio, COX IV activity and mitochondrial membrane potential. Overall, this study demonstrated that up-regulation of the KPI-containing APP isoforms is likely to contribute to the impairment of metabolic enzymes and mitochondrial function in AD. PMID:23872114

  1. Solid-state NMR analysis of the {beta}-strand orientation of the protofibrils of amyloid {beta}-protein

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Takashi [Graduate School of Science, Kyoto University, Kyoto 606-8502 (Japan); Masuda, Yuichi, E-mail: masuda@mail.pharm.tohoku.ac.jp [Graduate School of Science, Kyoto University, Kyoto 606-8502 (Japan); Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578 (Japan); Irie, Kazuhiro [Graduate School of Agriculture, Kyoto University, Kyoto 606-8502 (Japan); Akagi, Ken-ichi; Monobe, Youko; Imazawa, Takayoshi [Section of Laboratory Equipment, Division of Biomedical Research, National Institute of Biomedical Innovation, Osaka 567-0085 (Japan); Takegoshi, K. [Graduate School of Science, Kyoto University, Kyoto 606-8502 (Japan)

    2012-11-30

    Highlights: Black-Right-Pointing-Pointer The supramolecular structure of A{beta}42 protofibrils was analyzed by solid-state NMR. Black-Right-Pointing-Pointer The Ala-21 residue in the A{beta}42 protofibrils is included in a slightly disordered {beta}-strand. Black-Right-Pointing-Pointer The A{beta}42 protofibrils do not form intermolecular in-register parallel {beta}-sheets. -- Abstract: Alzheimer's disease (AD) is caused by abnormal deposition (fibrillation) of a 42-residue amyloid {beta}-protein (A{beta}42) in the brain. During the process of fibrillation, the A{beta}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{beta}42 protofibrils, the intermolecular proximity of the Ala-21 residues in the A{beta}42 protofibrils was analyzed by {sup 13}C-{sup 13}C rotational resonance experiments in the solid state. Unlike the A{beta}42 fibrils, an intermolecular {sup 13}C-{sup 13}C correlation was not found in the A{beta}42 protofibrils. This result suggests that the {beta}-strands of the A{beta}42 protofibrils are not in an in-register parallel orientation. A{beta}42 monomers would assemble to form protofibrils with the {beta}-strand conformation, then transform into fibrils by forming intermolecular parallel {beta}-sheets.

  2. The low-density lipoprotein receptor-related protein 1 and amyloid-? clearance in Alzheimer’s disease

    Directory of Open Access Journals (Sweden)

    Takahisa Kanekiyo

    2014-05-01

    Full Text Available Accumulation and aggregation of amyloid-? (A? peptides in the brain trigger the development of progressive neurodegeneration and dementia associated with Alzheimer’s disease (AD. Perturbation in A? clearance, rather than A? production, is likely the cause of sporadic, late-onset AD, which accounts for the majority of AD cases. Since cellular uptake and subsequent degradation constitute a major A? clearance pathway, the receptor-mediated endocytosis of A? has been intensely investigated. Among A? receptors, the low-density lipoprotein receptor-related protein 1 (LRP1 is one of the most studied receptors. LRP1 is a large endocytic receptor for more than 40 ligands, including apolipoprotein E (apoE, ?2-macroglobulin and A?. Emerging in vitro and in vivo evidence demonstrates that LRP1 is critically involved in brain A? clearance. LRP1 is highly expressed in a variety of cell types in the brain including neurons, vascular cells and glial cells, where LRP1 functions to maintain brain homeostasis and control A? metabolism. LRP1-mediated endocytosis regulates cellular A? uptake by binding to A? either directly or indirectly through its co-receptors or ligands. Furthermore, LRP1 regulates several signaling pathways, which also likely influences A? endocytic pathways. In this review, we discuss how LRP1 regulates the brain A? clearance and how this unique endocytic receptor participates in AD pathogenesis. Understanding of the mechanisms underlying LRP1-mediated A? clearance should enable the rational design of novel diagnostic and therapeutic strategies for AD.

  3. Serum Advanced Oxidation Protein Products in Oral Squamous Cell Carcinoma: Possible Markers of Diagnostic Significance

    Directory of Open Access Journals (Sweden)

    Abhishek Singh Nayyar

    2013-07-01

    Full Text Available Background: The aim of this study was to measure the concentrations (levels ofserum total proteins and advanced oxidation protein products as markers of oxidantmediated protein damage in the sera of patients with oral cancers.Methods: The study consisted of the sera analyses of serum total protein andadvanced oxidation protein products’ levels in 30 age and sex matched controls, 60patients with reported pre-cancerous lesions and/or conditions and 60 patients withhistologically proven oral squamous cell carcinoma. One way analyses of variance wereused to test the difference between groups. To determine which of the two groups’ meanswere significantly different, the post-hoc test of Bonferroni was used. The results wereaveraged as mean ± standard deviation. In the above test, P values less than 0.05 weretaken to be statistically significant. The normality of data was checked before thestatistical analysis was performed.Results: The study revealed statistically significant variations in serum levels ofadvanced oxidation protein products (P<0.001. Serum levels of total protein showedextensive variations; therefore the results were largely inconclusive and statisticallyinsignificant.Conclusion: The results emphasize the need for more studies with larger samplesizes to be conducted before a conclusive role can be determined for sera levels of totalprotein and advanced oxidation protein products as markers both for diagnosticsignificance and the transition from the various oral pre-cancerous lesions and conditionsinto frank oral cancers.

  4. Serum immune-related proteins are differentially expressed during hibernation in the American black bear.

    Science.gov (United States)

    Chow, Brian A; Donahue, Seth W; Vaughan, Michael R; McConkey, Brendan; Vijayan, Mathilakath M

    2013-01-01

    Hibernation is an adaptation to conserve energy in the face of extreme environmental conditions and low food availability that has risen in several animal phyla. This phenomenon is characterized by reduced metabolic rate (?25% of the active basal metabolic rate in hibernating bears) and energy demand, while other physiological adjustments are far from clear. The profiling of the serum proteome of the American black bear (Ursus americanus) may reveal specific proteins that are differentially modulated by hibernation, and provide insight into the remarkable physiological adaptations that characterize ursid hibernation. In this study, we used differential gel electrophoresis (DIGE) analysis, liquid chromatography coupled to tandem mass spectrometry, and subsequent MASCOT analysis of the mass spectra to identify candidate proteins that are differentially expressed during hibernation in captive black bears. Seventy serum proteins were identified as changing by ±1.5 fold or more, out of which 34 proteins increased expression during hibernation. The majority of identified proteins are involved in immune system processes. These included ?2-macroglobulin, complement components C1s and C4, immunoglobulin ? and J chains, clusterin, haptoglobin, C4b binding protein, kininogen 1, ?2-HS-glycoprotein, and apoplipoproteins A-I and A-IV. Differential expression of a subset of these proteins identified by proteomic analysis was also confirmed by immunodetection. We propose that the observed serum protein changes contribute to the maintenance of the hibernation phenotype and health, including increased capacities for bone maintenance and wound healing during hibernation in bears. PMID:23825529

  5. Amyloid protein-mediated differential DNA methylation status regulates gene expression in Alzheimer's disease model cell line

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Hye Youn; Choi, Eun Nam [Department of Biochemistry, School of Medicine, Ewha Womans University, 911-1 Mok-6-dong, Yangcheon-ku, Seoul 158-710 (Korea, Republic of); Ahn Jo, Sangmee [Department of Pharmacy, College of Pharmacy, Dankook University, San 29 Anseo-dong, Dongnam-gu, Cheonan-si, Chungnam 330-714 (Korea, Republic of); Oh, Seikwan [Department of Neuroscience and TIDRC, School of Medicine, Ewha Womans University, 911-1 Mok-6-dong, Yangcheon-ku, Seoul 158-710 (Korea, Republic of); Ahn, Jung-Hyuck, E-mail: ahnj@ewha.ac.kr [Department of Biochemistry, School of Medicine, Ewha Womans University, 911-1 Mok-6-dong, Yangcheon-ku, Seoul 158-710 (Korea, Republic of)

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer Genome-wide DNA methylation pattern in Alzheimer's disease model cell line. Black-Right-Pointing-Pointer Integrated analysis of CpG methylation and mRNA expression profiles. Black-Right-Pointing-Pointer Identify three Swedish mutant target genes; CTIF, NXT2 and DDR2 gene. Black-Right-Pointing-Pointer The effect of Swedish mutation on alteration of DNA methylation and gene expression. -- Abstract: The Swedish mutation of amyloid precursor protein (APP-sw) has been reported to dramatically increase beta amyloid production through aberrant cleavage at the beta secretase site, causing early-onset Alzheimer's disease (AD). DNA methylation has been reported to be associated with AD pathogenesis, but the underlying molecular mechanism of APP-sw-mediated epigenetic alterations in AD pathogenesis remains largely unknown. We analyzed genome-wide interplay between promoter CpG DNA methylation and gene expression in an APP-sw-expressing AD model cell line. To identify genes whose expression was regulated by DNA methylation status, we performed integrated analysis of CpG methylation and mRNA expression profiles, and identified three target genes of the APP-sw mutant; hypomethylated CTIF (CBP80/CBP20-dependent translation initiation factor) and NXT2 (nuclear exporting factor 2), and hypermethylated DDR2 (discoidin domain receptor 2). Treatment with the demethylating agent 5-aza-2 Prime -deoxycytidine restored mRNA expression of these three genes, implying methylation-dependent transcriptional regulation. The profound alteration in the methylation status was detected at the -435, -295, and -271 CpG sites of CTIF, and at the -505 to -341 region in the promoter of DDR2. In the promoter region of NXT2, only one CpG site located at -432 was differentially unmethylated in APP-sw cells. Thus, we demonstrated the effect of the APP-sw mutation on alteration of DNA methylation and subsequent gene expression. This epigenetic regulatory mechanism may contribute to the pathogenesis of AD.

  6. Serum autoantibodies to neurofilament proteins in sporadic amyotrophic lateral sclerosis.

    Science.gov (United States)

    Couratier, P; Yi, F H; Preud'homme, J L; Clavelou, P; White, A; Sindou, P; Vallat, J M; Jauberteau, M O

    1998-02-01

    Anti-neurofilament (NF) autoantibodies were searched for by enzyme-linked immunosorbent assays (ELISA) in the serum from 85 sporadic amyotrophic lateral sclerosis (ALS) patients, 98 healthy controls and 79 patients with unrelated immunological diseases (Guillain-Barré syndrome, myasthenia gravis and multiple sclerosis). ELISA cutoff value was determined as mean control levels +2 SD and it corresponded to a specificity of 94%. Such high level antibodies were detected in 24.7% of ALS patients contrasting with 12.6% of neurological controls (P<0.05) and only 6.1% of healthy subjects (P<5.10[-4]). In ALS, anti-NF antibodies were significantly associated with a slow evolution, as measured by the mean time spent in the initial functional states. They did not relate with age, sex and clinical form. The predominant isotype of the anti-NF antibodies was IgM lambda by ELISA. In contrast to negative sera, indirect immunohistochemical studies demonstrated that most sera positive for anti-NF antibodies reacted with axons with predominant isotypes restricted to IgM lambda. By using Western blotting, small amounts of serum monoclonal IgM were found with a high frequency in anti-NF antibody-positive patients. These results suggest the possible involvement of anti-NF antibodies in an autoimmune process in a subgroup of ALS patients. PMID:9562303

  7. Prophylaxis and Treatment of Alzheimer's Disease by Delivery of an Adeno-Associated Virus Encoding a Monoclonal Antibody Targeting the Amyloid Beta Protein

    OpenAIRE

    Shimada, Masaru; Abe, Shinya; Takahashi, Toru; Shiozaki, Kazumasa; Okuda, Mitsue; Mizukami, Hiroaki; Klinman, Dennis M.; Ozawa, Keiya; Okuda, Kenji

    2013-01-01

    We previously reported on a monoclonal antibody (mAb) that targeted amyloid beta (Aß) protein. Repeated injection of that mAb reduced the accumulation of Aß protein in the brain of human Aß transgenic mice (Tg2576). In the present study, cDNA encoding the heavy and light chains of this mAb were subcloned into an adeno-associated virus type 1 (AAV) vector with a 2A/furin adapter. A single intramuscular injection of 3.0×1010 viral genome of these AAV vectors into C57BL/6 mice generated seru...

  8. Insulin amyloid fibrillation at above 100°C: New insights into protein folding under extreme temperatures

    OpenAIRE

    Arora, Anubhav; Ha, Chanki; Park, Chan Beum

    2004-01-01

    To investigate the folding behavior of amyloidogenic proteins under extreme temperatures, the kinetics of fibrillation and accompanying secondary structure transitions of bovine insulin were studied for temperatures ranging up to 140°C. The presence of extreme heat stress had traditionally been associated with irreversible denaturation of protein while the initial steps of such a denaturation process may be common with a fibril formation pathway of amyloidogenic proteins. The present work de...

  9. Agregados amiloides: rol en desórdenes de conformación proteica / Amyloid aggregates: role in protein misfolding disorders

    Scientific Electronic Library Online (English)

    Claudia, Duran-Aniotz; Inés, Moreno-Gonzalez; Rodrigo, Morales.

    2013-04-01

    Full Text Available [...] Abstract in english Misfolding and aggregation of proteins are the main features of a group of diseases termed Protein Misfolding Disorders (PMDs). PMDs include Alzheimer's disease and Transmissible Spongiform Encephalopathies, among many others. The deposition of protein aggregates is the main responsible for tissue d [...] amage and the consequent clinical signs generated in such disorders. In this review, we will focus in the role of protein aggregates in these diseases and in the putative mechanisms by which they exert their toxicity.

  10. Canine cancer screening via ultraviolet absorbance and fluorescence spectroscopy of serum proteins

    Science.gov (United States)

    Dickerson, Bryan D.; Geist, Brian L.; Spillman, William B., Jr.; Robertson, John L.

    2007-11-01

    A cost-effective optical cancer screening and monitoring technique was demonstrated in a pilot study of canine serum samples and was patented for commercialization. Compared to conventional blood chemistry analysis methods, more accurate estimations of the concentrations of albumin, globulins, and hemoglobin in serum were obtained by fitting the near UV absorbance and photoluminescence spectra of diluted serum as a linear combination of component reference spectra. Tracking these serum proteins over the course of treatment helped to monitor patient immune response to carcinoma and therapy. For cancer screening, 70% of dogs with clinical presentation of cancer displayed suppressed serum hemoglobin levels (below 20 mg/dL) in combination with atypical serum protein compositions, that is, albumin levels outside of a safe range (from 4 to 8 g/dL) and globulin levels above or below a more normal range (from 1.7 to 3.7 g/dL). Of the dogs that met these criteria, only 20% were given a false positive label by this cancer screening test.

  11. Evaluation of a type 1 diabetes serum cohort by SELDI-TOF MS protein profiling

    DEFF Research Database (Denmark)

    Albrethsen, J.; Kaas, A.

    2009-01-01

    Proteomics analysis of serum from patients with type 1 diabetes (T1D) may lead to novel biomarkers for prediction of disease and for patient monitoring. However, the serum proteome is highly sensitive to sample processing and before proteomics biomarker research serum cohorts should preferably be examined for potential bias between sample groups. S ELDI-TOF MS protein profiling was used for preliminary evaluation of a biological-bank with 766 serum samples from 270 patients with T1D, collected at 18 different paediatric centers representing 15 countries in Europe and Japan over 2 years (2000-2002). Samples collected 1 (n = 270), 6 (n = 248), and 12 (n = 248) months after T1D diagnosis were grouped across centers and compared. The serum protein profiles varied with collection site and day of analysis; however, markers of sample processing were not systematically different between samples collected at different times after diagnosis. Three members of the apolipoprotein family increased with time in patient serum collected 1, 6, and 12 months after diagnosis (ANOVA, p

  12. Clinical significance of serum sex hormones protein and lipid determination in patients with ulcerative colitis

    International Nuclear Information System (INIS)

    Objective: To investigate the relationships between changes of serum sex hormones levels and protein-lipid metabolism in patients with ulcerative colitis. Methods: Serum levels of estradiol (E2) pregnenedione (P), prolactin(PRL), luteinizing hormone (LH), follicle-stimulating hormone (FSH) (with CLIA), sree testos (T, with RIA) and total-protein (TP), albumin (Alb), globulin (G), albumin/globulinratio (A/G) total-cholesterd (TC), high density lipoprotein cholesterols (LDL-C) (with biochemistry were determined in 72 patients) with ulcerative colitis and 72 controls. Results: The serum levels of T, LH, FSH, TP, Alb, A/G, TC, LDL-C in patients with ulcerative colitis were significantly lower than those in controls (P2, PRL in patients with ulcerative colitis were significantly higher than those in controls (P2 were negatively correlated with TP, A/G and TC (P2 levels in the female sex (P>0.05) as well as between LH, FSH and T levels in the male sex (P>0.05). Conclusion: The abnormal serum levels of sex hormone might contribute to the development of hypoproteinaemia and lowered lipid levels in patients with ulcerative colitis. Treatment with correction of serum sex hormones levels mightrection of serum sex hormones levels might be beneficial to the patients. (authors)

  13. A preliminary study on the serum protein response in canine babesiosis : research communication

    Directory of Open Access Journals (Sweden)

    R.G. Lobetti

    2012-07-01

    Full Text Available Total serum protein, albumin, globulin, globulin fractions (alpha, beta and gamma globulins and an acute-phase protein ((alpha1-acid glycoprotein were evaluated in dogs with naturally occurring mild (Group 1, severe (Group 2 or complicated babesiosis (Group 3. Results showed that the total serumprotein, albumin, A/G ratio and (alphaglobulins were statistically different between Groups 1 and 2. There was no statistical difference between groups with total, (betaand (gamma globulins. The findings from this study suggest that dogs with mild and severe babesiosis had low total serum proteins, albumin, A/G ratio and (alpha globulins; dogs with complicated babesiosis showed no typical serum protein changes or patterns; and that there was no evidence of an acute-phase response detectable on serum protein electrophoresis in any of the 3 groups. A marked acute-phase response was, however, present, as measured by the (alpha1-acid glycoprotein, in all 3 groups. As this was a retrospective study, the possibility that the observed responses were due in part to concurrent disease could not be excluded.

  14. The correlation study of serum retinol binding protein 4 and atherosclerosis in type 2 diabetes

    International Nuclear Information System (INIS)

    Objective: To evaluate the association of Retinol binding protein 4(RBP4) with carotid atherosclerosis in type 2 diabetes mellitus (T2DM). Methods: ELISA was used to detect the serum RBP4 level in 100 T2DM in 39 of which carotid atherosclerosis was observed and in the 30 controls, transthyretin (TTR) and other clinical index were also tested. Results: Serum RBP4 level and RBP4/ TTR were significantly higher in T2DM with/without carotid atherosclerosis than that in the controls (P0, OR>1, P<0.1). Conclusion: Serum RBP4 may be a novel risk factor for diabetes with atherosclerosis. Importantly, the RBP4/TTR ratio is more valuable than serum RBP4 concentration to used to evaluate risk factor for carotid atherosclerosis in T2DM. (authors)

  15. Influence of human diets containing casein, soy protein isolate, and soy protein concentrate on serum cholesterol and lipoproteins in humans, rabbits and rats

    OpenAIRE

    Raaij, J.M.A., van

    1982-01-01

    It is well known that feeding animals such as rabbits with semipurified diets containing animal proteins, as for example casein, results in hypercholesterolemia and atherosclerosis. On the other hand, diets containing vegetable proteins such as soybean protein maintain low levels of serum cholesterol. Little is known about the effects of the type of protein in the diet in humans.This thesis deals with the effects of casein and soy protein on serum cholesterol and lipoproteins, as observed in ...

  16. Ratanasampil (Tibetan Medicine, RNSP Reduces ?-Amyloid Protein (A? and Pro-Inflammatory Factor Levels and Improves Cognitive Functions in Mild-to-Moderate Alzheimer’s Disease (AD Patients Living at High Altitude

    Directory of Open Access Journals (Sweden)

    Aiqin Zhu

    2012-02-01

    Full Text Available Ratanasampil (RNSP is a traditional Tibetan medicine used for the treatment of stroke and cerebrovascular diseases. Previous discoveries that RNSP can reduce ?-amyloid protein levels and increase learning and memory in Alzheimer’s mouse models (Tg2576 led us to investigate whether RNSP can improve cognitive functions in Alzheimer’s patients. In this study, 146 AD patients living in Qinghai province received either one gram or 0.33 gram daily of RNSP for 16 weeks. Placebo patients received Piracetam. Serum A?40 and A?42 levels were measured at the beginning of the study and after 4 and 16 weeks of treatment. Compared to the same group before treatment, MMSE scores, ADAS-cog scores and ADL scores were significantly improved (p < 0.01, p < 0.01 and p < 0.05, respectively in patients after 16-week treatment with high-dose RNSP No. significant differences were observed in either the low-dose RNSP or placebo groups (p > 0.05, p > 0.05. After 16-week treatment, serum TNF-?, IL-1?, IL-6 and A?42 levels were significantly decreased (p < 0. 01 in the high-dose RNSP group, whereas no significant differences were found in the low-dose and placebo groups. The A?42/A?40 ratio was significantly decreased after 4-week and 16-week treatment in the high-dose RNSP group (p < 0. 05, p < 0.01. Furthermore, serum A?42 concentrations had a strong positive correlation with TNF-?, IL-1? and IL-6 levels. There were no observable adverse effects in either treatment or control groups. We conclude that further clinical trials of RNSP in Alzheimer disease are warranted.

  17. PuF, an antimetastatic and developmental signaling protein, interacts with the Alzheimer’s amyloid-? precursor protein via a tissue-specific proximal regulatory element (PRE

    Directory of Open Access Journals (Sweden)

    Lahiri Debomoy K

    2013-01-01

    Full Text Available Abstract Background Alzheimer’s disease (AD is intimately tied to amyloid-? (A? peptide. Extraneuronal brain plaques consisting primarily of A? aggregates are a hallmark of AD. Intraneuronal A? subunits are strongly implicated in disease progression. Protein sequence mutations of the A? precursor protein (APP account for a small proportion of AD cases, suggesting that regulation of the associated gene (APP may play a more important role in AD etiology. The APP promoter possesses a novel 30 nucleotide sequence, or “proximal regulatory element” (PRE, at ?76/?47, from the +1 transcription start site that confers cell type specificity. This PRE contains sequences that make it vulnerable to epigenetic modification and may present a viable target for drug studies. We examined PRE-nuclear protein interaction by gel electrophoretic mobility shift assay (EMSA and PRE mutant EMSA. This was followed by functional studies of PRE mutant/reporter gene fusion clones. Results EMSA probed with the PRE showed DNA-protein interaction in multiple nuclear extracts and in human brain tissue nuclear extract in a tissue-type specific manner. We identified transcription factors that are likely to bind the PRE, using competition gel shift and gel supershift: Activator protein 2 (AP2, nm23 nucleoside diphosphate kinase/metastatic inhibitory protein (PuF, and specificity protein 1 (SP1. These sites crossed a known single nucleotide polymorphism (SNP. EMSA with PRE mutants and promoter/reporter clone transfection analysis further implicated PuF in cells and extracts. Functional assays of mutant/reporter clone transfections were evaluated by ELISA of reporter protein levels. EMSA and ELISA results correlated by meta-analysis. Conclusions We propose that PuF may regulate the APP gene promoter and that AD risk may be increased by interference with PuF regulation at the PRE. PuF is targeted by calcium/calmodulin-dependent protein kinase II inhibitor 1, which also interacts with the integrins. These proteins are connected to vital cellular and neurological functions. In addition, the transcription factor PuF is a known inhibitor of metastasis and regulates cell growth during development. Given that APP is a known cell adhesion protein and ferroxidase, this suggests biochemical links among cell signaling, the cell cycle, iron metabolism in cancer, and AD in the context of overall aging.

  18. Serum concentration and interaction properties of MBL/ficolin associated protein-1

    DEFF Research Database (Denmark)

    Skjoedt, Mikkel-Ole; Hummelshoj, Tina

    2011-01-01

    Recently, a novel protein named MBL/ficolin associated protein-1 (MAP-1) derived from the MASP1 gene through differential splicing was identified. In the present study, we established biochemical characteristics, determined the serum level and assessed the interactions between the lectin complement pathway (LCP) recognition molecules and MAP-1. We expressed recombinant MAP-1 in CHO DG44 cells, developed a quantitative ELISA assay based on a MAP-1 specific monoclonal capture antibody and measured the serum levels in 100 Danish blood donors. In addition we assessed the association properties between MAP-1 and Ficolin-2, -3 and MBL in serum using ELISA and density gradient ultra centrifugation. When recombinant MAP-1 was subjected to N-glycosidase F treatment the molecular mass decreased from ?45kDa to ?40kDa equivalent with the calculated molecular mass from the deduced amino acid sequence without the signal peptide. We found that serum MAP-1 was very stable when subjected to repeated freeze and thaw cycles. The mean serum concentration of MAP-1 was found to be 240ng/ml (range: 115-466ng/ml). MAP-1 was predominantly found in complex with Ficolin-3 and to a lesser degree with Ficolin-2 and MBL and by use of density gradient ultra centrifugation we could show that the major part of serum MAP-1 circulates in complex with the LCP molecules. In conclusion, these results show that MAP-1 is a highly stable glycosylated human serum protein found in complex with Ficolin-3, Ficolin-2 and MBL.

  19. Serum Adenosine deaminase activity and C-reactive protein levels in unstable angina

    Directory of Open Access Journals (Sweden)

    Rani Surekha

    2003-01-01

    Full Text Available In unstable angina (USA patients, immunological responses contributing to inflammation play a vital role in plaque rupture and thrombosis causing stroke. In the present study an attempt is made to estimate the levels of adenosine deaminase activity, an immunoenzyme marker and C-reactive protein, a marker of inflammation in USA patients. 45 patients presenting USA and 50 age and sex matched healthy controls were included in the study. Serum ADA activity was measured spectrophotometrically at 630nm and serum C-reactive protein was detected using Avitex CRP kit, which is a rapid latex agglutination test. The Mean ADA levels were 41.15 ± 11.04 in patients and 20.71±5.63 in controls and 66.6% of patients and none of the controls were positive to CRP. The present study observed the importance of ADA as a serum marker in addition to CRP for assessing the immune response in USA patients.

  20. Elevated serum levels of heat shock protein 70 can be detected after radiofrequency ablation

    OpenAIRE

    Haen, Sebastian P.; Gouttefangeas, Cécile; Schmidt, Diethard; Boss, Andreas; Clasen, Stephan; von Herbay, Alexandra; Kosan, Bora; Aebert, Hermann; Pereira, Philippe L.; Rammensee, Hans-Georg

    2011-01-01

    Due to their adjuvant effect and their ability to chaperone tumor-associated peptides, heat shock proteins constitute a potent alarm signal for the immune system and can lead to activation of anti-tumor T-cell immunity. Radiofrequency ablation has been reported to induce heat shock protein expression especially that of heat shock protein 70 in sublethally damaged tumor cells. In this study, we evaluated the release of heat shock protein 70 into the serum of cancer-bearing patients directly af...

  1. Electrophoretic studies on serum protein binding of radiopharmaceuticals for hepatobiliary scan

    International Nuclear Information System (INIS)

    Protein binding analyses of verious radiocompounds for hepatobiliary scintigraphy were performed on electrophoresis using a cellulose acetate menbrane in 0.05 M, pH 8.5 barbital buffer solution. The menbranes obtained were exposed tightly to X-ray film for making autoradiograms which were followed by a Microphotometry for semi-quantitative analysis. All of four sup(99m)Tc-labeled hepatobiliary agents, sup(99m)Tc-DM-IDA, sup(99m)Tc-DE-IDA, sup(99m)Tc-PI and sup(99m)Tc-PB-IDA, migrated to an anode, at the further distance than that of sup(99m)Tc-HSA employed as a control. These radioagents showed poor affinity to the albumin in the serum proteins. The other hand, 131I-labeled agents, 131I-BSP and 131I-RB, represented a very good binding to the human serum albumin. In further analysis, very interesting phenomena were shown in protein binding of 131I-BSP and 131I-RB. There was a increase of unbound compornent of 131I-BSP noted after adding of non-radioactive BSP to the human serum mixture with it. However, 131I-RB was not affected in it's protein affinity by adding of non-radioactive BSP. The other hand, hyperbilirubinemia (22 mg/dl) affected a protein affinity of 131I-RB to serum albumin but gave no significant affection to that of 131I-BSP. These results suggest that radioagents for hepatobiliary scan are not only qualitative but also quantitative, specific and competitivelso quantitative, specific and competitive in protein binding to a human serum albumin. (author)

  2. Revision of the biodistribution of uranyl in serum: is fetuin-A the major protein target?

    Science.gov (United States)

    Basset, Christian; Averseng, Olivier; Ferron, Pierre-Jean; Richaud, Nicolas; Hagège, Agnès; Pible, Olivier; Vidaud, Claude

    2013-05-20

    Uranium is a natural actinide present as uranyl U(VI) species in aqueous environments. Its toxicity is considered to be chemical rather than radiotoxicological. Whatever the route of entry, uranyl reaches the blood, is partly eliminated via the kidneys, and accumulated in the bones. In serum, its speciation mainly involves carbonate and proteins. Direct identification of labile uranyl-protein complexes is extremely difficult because of the complexity of this matrix. Thus, until now the biodistribution of the metal in serum has not been described, and therefore, little is known about the metal transport mechanisms leading to bone accumulation. A rapid screening method based on a surface plasmon resonance (SPR) technique was used to determine the apparent affinities for U(VI) of the major serum proteins. A first biodistribution of uranyl was obtained by ranking the proteins according to the criteria of both their serum concentrations and affinities for this metal. Despite its moderate concentration in serum, fetuin-A (FETUA) was shown to exhibit an apparent affinity within the 30 nM range and to carry more than 80% of the metal. This protein involved in bone mineralization aroused interest in characterizing the U(VI) and FETUA interaction. Using complementary chromatographic and spectroscopic approaches, we demonstrated that the protein can bind 3 U(VI) at different binding sites exhibiting Kd from ?30 nM to 10 ?M. Some structural modifications and functional properties of FETUA upon uranyl complexation were also controlled. To our knowledge, this article presents the first identification of a uranyl carrier involved in bone metabolism along with the characterization of its metal binding sites. PMID:23527557

  3. High-protein diets in hyperlipidemia : effect of wheat gluten on serum lipids, uric acid, and renal function

    OpenAIRE

    Jenkins, D.J.A.; Kendall, C.W.C.; Vidgen, E.; Augustin, L.S.A.; Erk, M., van; Geelen, A; Parker, T.; Faulkner, D.; Vuksan, V; Josse, R.G.; Leiter, L.A.; Connelly, P.W.

    2001-01-01

    BACKGROUND: The metabolic effects of diets high in vegetable protein have not been assessed despite much recent interest in the effect of soy proteins in reducing serum cholesterol. OBJECTIVE: We assessed the metabolic effects of diets high in vegetable protein (specifically, wheat gluten) on serum lipids, uric acid concentrations, and renal function. DESIGN: Twenty hyperlipidemic men and women consumed isoenergetic test (high-protein) and control metabolic diets for 1 mo in a randomized cros...

  4. Electrostatics controls the formation of amyloid-like superstructures in protein aggregation

    CERN Document Server

    Foderà, Vito; Lattuada, Marco; Donald, Athene M

    2013-01-01

    The possibility for proteins to aggregate in different superstructures, i.e. large-scale polymorphism, has been widely observed, but an understanding of the physico-chemical mechanisms behind it is still out of reach. Here we present a theoretical model for the description of a generic aggregate formed from an ensemble of charged proteins. The model predicts the formation of multi-fractal structures with the geometry of the growth determined by the electrostatic interactions between single proteins. The model predictions are successfully verified in comparison with experimental curves for aggregate growth. The model is general and is able to predict aggregate morphologies occurring both in vivo and in vitro. Our findings provide for the first time a unifying and general framework where the physical interactions between single proteins, the aggregate morphology and the growth kinetics are connected into a single model in excellent agreement with the experimental data.

  5. A preliminary study on the serum protein response in canine babesiosis : research communication

    OpenAIRE

    Lobetti, R. G.; Mo?hr, A. J.; Dippenaar, T.; Myburgh, E.

    2012-01-01

    Total serum protein, albumin, globulin, globulin fractions (alpha, beta and gamma globulins) and an acute-phase protein ((alpha)1-acid glycoprotein) were evaluated in dogs with naturally occurring mild (Group 1), severe (Group 2) or complicated babesiosis (Group 3). Results showed that the total serumprotein, albumin, A/G ratio and (alpha)globulins were statistically different between Groups 1 and 2. There was no statistical difference between groups with total, (beta)and (gamma) globulins. T...

  6. In Vivo Studies of Serum C-reactive Protein Turnover in Rabbits

    OpenAIRE

    Chelladurai, Mohanathasan; Macintyre, Stephen S.; Kushner, Irving

    1983-01-01

    We determined the plasma half-life of the acute phase protein C-reactive protein (CRP) both in normal rabbits and in rabbits that had received inflammatory stimuli. Rabbit CRP was purified from acute phase serum by Cx-polysaccharide affinity chromatography, radiolabeled, and rendered pyrogen-free. Six unstimulated rabbits were injected intravenously with 1251-CRP prepared by the lactoperoxidase method and four were injected with CRP labeled by methylation using [14C]formaldehyde. Blood sample...

  7. Targeted quantification of low ng/mL level proteins in human serum without immunoaffinity depletion

    OpenAIRE

    Shi, Tujin; Sun, Xuefei; Gao, Yuqian; Fillmore, Thomas L.; Schepmoes, Athena A.; Zhao, Rui; He, Jintang; Moore, Ronald J.; Kagan, Jacob; Rodland, Karin D.; Liu, Tao; Liu, Alvin Y.; Smith, Richard D.; Tang, Keqi; Camp, David G.

    2013-01-01

    We recently reported an antibody-free targeted protein quantification strategy, termed high-pressure, high-resolution separations with intelligent selection and multiplexing (PRISM) for achieving significantly enhanced sensitivity using selected reaction monitoring (SRM) mass spectrometry. Integrating PRISM with front-end IgY14 immunoaffinity depletion, sensitive detection of targeted proteins at 50–100 pg/mL levels in human blood plasma/serum was demonstrated. However, immunoaffinity deple...

  8. Serum protein profiles as potential biomarkers for infectious disease status in pigs

    Directory of Open Access Journals (Sweden)

    Koene Miriam GJ

    2012-03-01

    Full Text Available Abstract Background In veterinary medicine and animal husbandry, there is a need for tools allowing the early warning of diseases. Preferably, tests should be available that warn farmers and veterinarians during the incubation periods of disease and before the onset of clinical signs. The objective of this study was to explore the potential of serum protein profiles as an early biomarker for infectious disease status. Serum samples were obtained from an experimental pig model for porcine circovirus-associated disease (PCVAD, consisting of Porcine Circovirus type 2 (PCV2 infection in combination with either Porcine Parvovirus (PPV or Porcine Reproductive and Respiratory Syndrome virus (PRRSV. Sera were collected before and after onset of clinical signs at day 0, 5 and 19 post infection. Serum protein profiles were evaluated against sera from non-infected control animals. Results Protein profiles were generated by SELDI-TOF mass spectrometry in combination with the Proteominer™ technology to enrich for low-abundance proteins. Based on these protein profiles, the experimentally infected pigs could be classified according to their infectious disease status. Before the onset of clinical signs 88% of the infected animals could be classified correctly, after the onset of clinical sigs 93%. The sensitivity of the classification appeared to be high. The protein profiles could distinguish between separate infection models, although specificity was moderate to low. Classification of PCV2/PRRSV infected animals was superior compared to PCV2/PPV infected animals. Limiting the number of proteins in the profiles (ranging from 568 to 10 had only minor effects on the classification performance. Conclusions This study shows that serum protein profiles have potential for detection and identification of viral infections in pigs before clinical signs of the disease become visible.

  9. Effect of feed supplement on Milk Production, Fat % Total Serum Protein and Minerals in Lactating Buffalo

    OpenAIRE

    Verma, R.K.; Praveen Kumar; A. Adil and G.K. Arya

    2009-01-01

    A study was carried out to see the effect of feed supplement “Khurak” on milk yielding buffalo. The buffaloes were divided in two group. One group was offered “Khurak” as feed supplement for 7 days. Significant increase was observed in milk production, Total serum protein and calcium in khurak supplemented group (Treatment group). [Vet. World 2009; 2(5.000): 193-194

  10. Discovery of potential serum protein biomarkers for lymph-node metastasis in oral cancer.

    Science.gov (United States)

    Chai, Yang D; Zhang, Lifeng; Yang, Yan; Su, Trent; Charugundla, Prashant; Ai, Jiye; Messadi, Diana; Wong, David T; Hu, Shen

    2014-09-16

    Background: The purpose of our study is to identify serum protein biomarkers for node-positive oral squamous cell carcinoma (OSCC). Biomarkers indicating lymph node metastasis provides valuable classification methodology to optimize treatment plans for patients with OSCC. Methods: Quantitative serum proteomic analysis of OSCCs with either node-positive or node-negative disease was performed with tandem mass spectrometry and isobaric tagging for relative and absolute quantitation (iTRAQ). Immunoassays were used to validate a panel of candidate protein biomarkers and receiver operating characteristic analysis was used to evaluate the performance of the candidate biomarkers. Results: A total of 282 serum proteins were quantified between node-positive and node-negative OSCCs with the proteomic approach. Four candidate biomarkers, gelsolin, fibronectin, angiotensinogen and haptoglobin, were validated in an independent group of patients with node-positive or node-negative OSCC. The best candidate biomarker, gelsolin, yielded a receiver operating characteristic value of 89 % for node-positive OSCC, although the sample size for validation is relatively small. Fibronectin, gelsolin and angiotensinogen were also found to be differentially expressed between cancer cell lines of node-positive and node-negative cancer origin. Conclusion: Our studies suggest that testing of serum protein biomarkers might help detect lymph node metastasis of oral cancer. Due to limited sample size in our studies, long-term longitudinal studies with large populations of individuals with oral cancer are needed to validate these potential biomarkers. Head Neck, 2014. PMID:25223295

  11. Presence of non-fibrillar amyloid beta protein in skin biopsies of Alzheimer's disease (AD), Down's syndrome and non-AD normal persons

    DEFF Research Database (Denmark)

    Wen, G Y; Wisniewski, H M

    1994-01-01

    A total of 66 skin biopsies from persons with Alzheimer's disease (AD) or Down's syndrome (DS) and from persons without AD were used in this study. The age range was from 7 to 89 years. Positive immunoreactivity of skin biopsies to monoclonal antibody 4G8, which is reactive to amino acid residue 17-24 of synthetic amyloid beta protein (A beta), and 4G8-Fab (the antigen-binding fragment of 4G8 IgG, reactive only to amyloid plaque) was observed in the epidermis-dermis junction or the basement membrane of the epidermis and in some blood vessels of the biopsy skins of 13/18 (72%) AD, 9/10 (90%) DS, and 14/38 (37%) non-AD control cases. The Fisher exact probability test revealed a significant difference (P = 0.0415 one-tailed) in immunoreactivity between AD and age-matched controls. There was also a significant difference (P = 0.0152 one-tailed; P = 0.0200 two-tailed) between DS and age-matched control in the same test. Immuno-gold electron microscopy examination of these cases with positive immunoreactivity revealed that the gold particles were deposited along the basement membrane of the epidermis. Amyloid fibrils were not observed in the regions with gold particles. Results of this study suggest that A beta is associated with the basement membrane of skin and is present in amorphous, non-fibrillar form as soluble A beta.

  12. Immunoprecipitation of amyloid fibrils by the use of an antibody that recognizes a generic epitope common to amyloid fibrils.

    Science.gov (United States)

    Greiner, Erin R; Kelly, Jeffery W; Palhano, Fernando L

    2014-01-01

    Amyloid fibrils are associated with many maladies, including Alzheimer's disease (AD). The isolation of amyloids from natural materials is very challenging because the extreme structural stability of amyloid fibrils makes it difficult to apply conventional protein science protocols to their purification. A protocol to isolate and detect amyloids is desired for the diagnosis of amyloid diseases and for the identification of new functional amyloids. Our aim was to develop a protocol to purify amyloid from organisms, based on the particular characteristics of the amyloid fold, such as its resistance to proteolysis and its capacity to be recognized by specific conformational antibodies. We used a two-step strategy with proteolytic digestion as the first step followed by immunoprecipitation using the amyloid conformational antibody LOC. We tested the efficacy of this method using as models amyloid fibrils produced in vitro, tissue extracts from C. elegans that overexpress A? peptide, and cerebrospinal fluid (CSF) from patients diagnosed with AD. We were able to immunoprecipitate A?(1-40) amyloid fibrils, produced in vitro and then added to complex biological extracts, but not ?-synuclein and gelsolin fibrils. This method was useful for isolating amyloid fibrils from tissue homogenates from a C. elegans AD model, especially from aged worms. Although we were able to capture picogram quantities of A?(1-40) amyloid fibrils produced in vitro when added to complex biological solutions, we could not detect any A? amyloid aggregates in CSF from AD patients. Our results show that although immunoprecipitation using the LOC antibody is useful for isolating A?(1-40) amyloid fibrils, it fails to capture fibrils of other amyloidogenic proteins, such as ?-synuclein and gelsolin. Additional research might be needed to improve the affinity of these amyloid conformational antibodies for an array of amyloid fibrils without compromising their selectivity before application of this protocol to the isolation of amyloids. PMID:25144803

  13. PB1-F2 influenza A virus protein adopts a beta-sheet conformation and forms amyloid fibers in membrane environments.

    Science.gov (United States)

    Chevalier, Christophe; Al Bazzal, Ali; Vidic, Jasmina; Février, Vincent; Bourdieu, Christiane; Bouguyon, Edwige; Le Goffic, Ronan; Vautherot, Jean-François; Bernard, Julie; Moudjou, Mohammed; Noinville, Sylvie; Chich, Jean-François; Da Costa, Bruno; Rezaei, Human; Delmas, Bernard

    2010-04-23

    The influenza A virus PB1-F2 protein, encoded by an alternative reading frame in the PB1 polymerase gene, displays a high sequence polymorphism and is reported to contribute to viral pathogenesis in a sequence-specific manner. To gain insights into the functions of PB1-F2, the molecular structure of several PB1-F2 variants produced in Escherichia coli was investigated in different environments. Circular dichroism spectroscopy shows that all variants have a random coil secondary structure in aqueous solution. When incubated in trifluoroethanol polar solvent, all PB1-F2 variants adopt an alpha-helix-rich structure, whereas incubated in acetonitrile, a solvent of medium polarity mimicking the membrane environment, they display beta-sheet secondary structures. Incubated with asolectin liposomes and SDS micelles, PB1-F2 variants also acquire a beta-sheet structure. Dynamic light scattering revealed that the presence of beta-sheets is correlated with an oligomerization/aggregation of PB1-F2. Electron microscopy showed that PB1-F2 forms amorphous aggregates in acetonitrile. In contrast, at low concentrations of SDS, PB1-F2 variants exhibited various abilities to form fibers that were evidenced as amyloid fibers in a thioflavin T assay. Using a recombinant virus and its PB1-F2 knock-out mutant, we show that PB1-F2 also forms amyloid structures in infected cells. Functional membrane permeabilization assays revealed that the PB1-F2 variants can perforate membranes at nanomolar concentrations but with activities found to be sequence-dependent and not obviously correlated with their differential ability to form amyloid fibers. All of these observations suggest that PB1-F2 could be involved in physiological processes through different pathways, permeabilization of cellular membranes, and amyloid fiber formation. PMID:20172856

  14. The mechanism of membrane disruption by cytotoxic amyloid oligomers formed by prion protein(106-126) is dependent on bilayer composition.

    Science.gov (United States)

    Walsh, Patrick; Vanderlee, Gillian; Yau, Jason; Campeau, Jody; Sim, Valerie L; Yip, Christopher M; Sharpe, Simon

    2014-04-11

    The formation of fibrillar aggregates has long been associated with neurodegenerative disorders such as Alzheimer and Parkinson diseases. Although fibrils are still considered important to the pathology of these disorders, it is now widely understood that smaller amyloid oligomers are the toxic entities along the misfolding pathway. One characteristic shared by the majority of amyloid oligomers is the ability to disrupt membranes, a commonality proposed to be responsible for their toxicity, although the mechanisms linking this to cell death are poorly understood. Here, we describe the physical basis for the cytotoxicity of oligomers formed by the prion protein (PrP)-derived amyloid peptide PrP(106-126). We show that oligomers of this peptide kill several mammalian cells lines, as well as mouse cerebellar organotypic cultures, and we also show that they exhibit antimicrobial activity. Physical perturbation of model membranes mimicking bacterial or mammalian cells was investigated using atomic force microscopy, polarized total internal reflection fluorescence microscopy, and NMR spectroscopy. Disruption of anionic membranes proceeds through a carpet or detergent model as proposed for other antimicrobial peptides. By contrast, when added to zwitterionic membranes containing cholesterol-rich ordered domains, PrP(106-126) oligomers induce a loss of domain separation and decreased membrane disorder. Loss of raft-like domains may lead to activation of apoptotic pathways, resulting in cell death. This work sheds new light on the physical mechanisms of amyloid cytotoxicity and is the first to clearly show membrane type-specific modes of action for a cytotoxic peptide. PMID:24554723

  15. Critical analysis of the use of ?-site amyloid precursor protein-cleaving enzyme 1 inhibitors in the treatment of Alzheimer's disease

    Directory of Open Access Journals (Sweden)

    Evin G

    2014-01-01

    Full Text Available Genevieve Evin,1,2 Adel Barakat21Oxidation Biology Laboratory, Mental Health Research Institute, Florey Institute of Neuroscience and Mental Health, University of Melbourne, 2Department of Pathology, University of Melbourne, Parkville, VIC, AustraliaAbstract: Alzheimer's disease (AD is the major cause of dementia in the elderly and an unmet clinical challenge. A variety of therapies that are currently under development are directed to the amyloid cascade. Indeed, the accumulation and toxicity of amyloid-? (A? is believed to play a central role in the etiology of the disease, and thus rational interventions are aimed at reducing the levels of A? in the brain. Targeting ?-site amyloid precursor protein-cleaving enzyme (BACE-1 represents an attractive strategy, as this enzyme catalyzes the initial and rate-limiting step in A? production. Observation of increased levels of BACE1 and enzymatic activity in the brain, cerebrospinal fluid, and platelets of patients with AD and mild cognitive impairment supports the potential benefits of BACE1 inhibition. Numerous potent inhibitors have been generated, and many of these have been proved to lower A? levels in the brain of animal models. Over 10 years of intensive research on BACE1 inhibitors has now culminated in advancing half a dozen of these drugs into human trials, yet translating the in vitro and cellular efficacy of BACE1 inhibitors into preclinical and clinical trials represents a challenge. This review addresses the promises and also the potential problems associated with BACE1 inhibitors for AD therapy, as the complex biological function of BACE1 in the brain is becoming unraveled.Keywords: amyloid, dementia, secretase, aspartyl protease, neuregulin

  16. Nature of 54Mn binding protein in rat brain, liver and serum

    International Nuclear Information System (INIS)

    Studies were undertaken to investigate the 54Mn binding protein in rat serum, liver and brain. Equilibrium dialysis pattern in three biological media with 54Mn against distilld water showed a rectangular hyperbola after 24 hr dialysis. 54Mn in the bag was in the non-dialysable form, presumably bound to protein. Gel filtration studies with 54Mn bound protein showed that 54Mn had a specific carrier protein, which was identical for all the three biological media tested. The electrophorogram of concentrated 54Mn protein with Comassie blue stain for protein and PAS stain for glycoprotein showed a single band with an equal RF value clearly indicating the glycoprotein nature of 54Mn binding protein. Sulphydryl groups were absent and it had molecular weight of 6.3 x 104 dalton. (author)

  17. Cerebral Amyloid Angiopathy: Emerging Concepts

    Science.gov (United States)

    2015-01-01

    Cerebral amyloid angiopathy (CAA) involves cerebrovascular amyloid deposition and is classified into several types according to the amyloid protein involved. Of these, sporadic amyloid ?-protein (A?)-type CAA is most commonly found in older individuals and in patients with Alzheimer's disease (AD). Cerebrovascular A? deposits accompany functional and pathological changes in cerebral blood vessels (CAA-associated vasculopathies). CAA-associated vasculopathies lead to development of hemorrhagic lesions [lobar intracerebral macrohemorrhage, cortical microhemorrhage, and cortical superficial siderosis (cSS)/focal convexity subarachnoid hemorrhage (SAH)], ischemic lesions (cortical infarction and ischemic changes of the white matter), and encephalopathies that include subacute leukoencephalopathy caused by CAA-associated inflammation/angiitis. Thus, CAA is related to dementia, stroke, and encephalopathies. Recent advances in diagnostic procedures, particularly neuroimaging, have enabled us to establish a clinical diagnosis of CAA without brain biopsies. Sensitive magnetic resonance imaging (MRI) methods, such as gradient-echo T2* imaging and susceptibility-weighted imaging, are useful for detecting cortical microhemorrhages and cSS. Amyloid imaging with amyloid-binding positron emission tomography (PET) ligands, such as Pittsburgh Compound B, can detect CAA, although they cannot discriminate vascular from parenchymal amyloid deposits. In addition, cerebrospinal fluid markers may be useful, including levels of A?40 for CAA and anti-A? antibody for CAA-related inflammation. Moreover, cSS is closely associated with transient focal neurological episodes (TFNE). CAA-related inflammation/angiitis shares pathophysiology with amyloid-related imaging abnormalities (ARIA) induced by A? immunotherapies in AD patients. This article reviews CAA and CAA-related disorders with respect to their epidemiology, pathology, pathophysiology, clinical features, biomarkers, diagnosis, treatment, risk factors, and future perspectives. PMID:25692104

  18. In vivo radioimmunodetection of amyloid deposits in experimental amyloidosis

    International Nuclear Information System (INIS)

    Two rat IgG monoclonal antibodies (MAbs) to mouse AA protein have been used as reagents for an ELISA assay for SAA, for the demonstration of tissue deposits by the immunoperoxidase method, and for the definition of AA polymorphs in solubilized amyloidotic tissue specimens run onto two-dimensional (2D) gels and probed as Western Blots. Both MAbs localize to tissue deposits in vivo when labelled (MAb*) with I125 or I123 and injected into colchicine-pretreated amyloidotic mice, assessed by (a) whole-body autoradiography (WBAR) (b) external photoscanning (c) tissue autoradiography of perfused organs. Serum t 1/2 of MAb in amyloidotic animals was greatly accelerated compared to controls, and sustained concentration of label was found in spleen, liver, and kidney quantitated by organ counts up to 96 h post injection. MAb label copurified with amyloid fibrils up to 10-fold over saline-soluble proteins and residue following homogenization in distilled water and acid extraction; purified fibrils contained AA protein and MAb heavy and light chain visualized on 2D gels and by autoradiography. Animals were also injected with a mixture of the two MAbs, one labeled with Indium111-DTPA (126 ?Ci), the other with I125 (6 ?Ci) and serially scanned over a 48 h period. Fractionation of serum taken after injection showed both isotopes to be present as uncomplexed IgG in blood. WBAR was performed at 48 h and at a time at which I125/Iand at a time at which I125/In111 radioactivity was calculated to be 127/1; autoradiograms were similar and confirmed localization of both isotopes to tissue deposits in amyloidotic animals. These studies provide evidence that MAbs to amyloid subunit proteins may be useful reagents for the in vivo radioimmunodetection of some of the amyloid diseases

  19. Altered protein expression in serum from endometrial hyperplasia and carcinoma patients

    Directory of Open Access Journals (Sweden)

    Cong Qing

    2011-04-01

    Full Text Available Abstract Background Endometrial carcinoma is one of the most common gynecological malignancies in women. The diagnosis of the disease at early or premalignant stages is crucial for the patient's prognosis. To date, diagnosis and follow-up of endometrial carcinoma and hyperplasia require invasive procedures. Therefore, there is considerable demand for the identification of biomarkers to allow non-invasive detection of these conditions. Methods In this study, we performed a quantitative proteomics analysis on serum samples from simple endometrial hyperplasia, complex endometrial hyperplasia, atypical endometrial hyperplasia, and endometrial carcinoma patients, as well as healthy women. Serum samples were first depleted of high-abundance proteins, labeled with isobaric tags (iTRAQ™, and then analyzed via two-dimensional liquid chromatography and tandem mass spectrometry. Protein identification and quantitation information were acquired by comparing the mass spectrometry data against the International Protein Index Database using ProteinPilot software. Bioinformatics annotation of identified proteins was performed by searching against the PANTHER database. Results In total, 74 proteins were identified and quantified in serum samples from endometrial lesion patients and healthy women. Using a 1.6-fold change as the benchmark, 12 proteins showed significantly altered expression levels in at least one disease group compared with healthy women. Among them, 7 proteins were found, for the first time, to be differentially expressed in atypical endometrial hyperplasia. These proteins are orosomucoid 1, haptoglobin, SERPINC 1, alpha-1-antichymotrypsin, apolipoprotein A-IV, inter-alpha-trypsin inhibitor heavy chain H4, and histidine-rich glycoprotein. Conclusions The differentially expressed proteins we discovered in this study may serve as biomarkers in the diagnosis and follow-up of endometrial hyperplasia and endometrial carcinoma.

  20. Protein and fat mobilization and associations with serum ?-hydroxybutyrate concentrations in dairy cows.

    Science.gov (United States)

    van der Drift, S G A; Houweling, M; Schonewille, J T; Tielens, A G M; Jorritsma, R

    2012-09-01

    The objective of this study was to obtain information on variation between dairy cows in muscle and fat tissue mobilization around parturition and to study the association between protein and fat mobilization and serum ?-hydroxybutyrate (BHBA) concentrations (hyperketonemia) in this period. Thirty-four cows kept under similar conditions at a university dairy farm (no experimental treatments) were monitored from 4 wk before until 8 wk after calving. Mobilization of muscle protein was investigated by analysis of plasma 3-methylhistidine concentrations (3-MH, analyzed by a recently developed HPLC tandem mass spectrometry method) and ultrasound measurements of longissimus muscle thickness. Mobilization of fat tissue was monitored by serum nonesterified fatty acid (NEFA) concentrations and ultrasound measurements of backfat thickness. Large variation was observed between cows in onset and duration of periparturient protein and fat mobilization. Plasma 3-MH concentrations and muscle thickness profiles indicated that protein mobilization started, on average, before parturition and continued until approximately wk 4 of lactation. Serum NEFA concentrations and backfat thickness profiles showed that fat mobilization occurred from parturition until the end of the study. Thus, muscle protein mobilization occurred in advance of fat mobilization in most cows from this study. We hypothesized that this might be due to a prepartum amino acid deficiency in the absence of negative energy balance. The incidence of hyperketonemia in this study was 16/34 = 47%. With the exception of 3 cows defined as having severe hyperketonemia, cows with lower 3-MH concentrations had higher serum BHBA concentrations. A possible explanation for this observation might be that higher mobilization of protein around calving might restrict ketone body production due to the higher availability of glucogenic precursors in the period of most severe negative energy balance and highest fat mobilization. The validity of this hypothesis needs to be confirmed, but data from this study indicate that further research on the role of protein mobilization in the etiology of hyperketonemia in dairy cows is needed. PMID:22916895

  1. Comprehensive characterisation of pulmonary and serum surfactant protein D in COPD

    Directory of Open Access Journals (Sweden)

    Erpenbeck Veit J

    2011-03-01

    Full Text Available Abstract Background Pulmonary surfactant protein D (SP-D is considered as a candidate biomarker for the functional integrity of the lung and for disease progression, which can be detected in serum. The origin of SP-D in serum and how serum concentrations are related to pulmonary concentrations under inflammatory conditions is still unclear. Methods In a cross-sectional study comprising non-smokers (n = 10, young - (n = 10, elderly smokers (n = 20, and smokers with COPD (n = 20 we simultaneously analysed pulmonary and serum SP-D levels with regard to pulmonary function, exercise, repeatability and its quaternary structure by native gel electrophoresis. Statistical comparisons were conducted by ANOVA and post-hoc testing for multiple comparisons; repeatability was assessed by Bland-Altman analysis. Results In COPD, median (IQR pulmonary SP-D levels were lower (129(68 ng/ml compared to smokers (young: 299(190, elderly: 296(158 ng/ml; p Conclusions Pulmonary and serum SP-D levels are stable markers influenced by smoking and related to airflow obstruction and disease state. Smaller subunits of pulmonary SP-D and the rapid increase of serum SP-D levels in COPD due to exercise support the translocation hypothesis and its use as a COPD biomarker. Trial registration no interventional trial

  2. Evaluation of full-length, cleaved and nitrosylated serum surfactant protein D as biomarkers for COPD

    DEFF Research Database (Denmark)

    Duvoix, Annelyse; Miranda, Elena

    2011-01-01

    Chronic obstructive pulmonary disease (COPD) is a multicomponent condition that is characterized by partially reversible airflow obstruction. Serum surfactant protein D (SP-D) is synthesized by type II pneumocytes and Clara cells and participates in surfactant homeostasis and pulmonary host defense. Serum levels of SP-D are raised in individuals with COPD but there is no correlation between the serum level of SP-D and the severity of airflow obstruction. Serum SP-D is present in different forms that may have more utility as a biomarker for COPD. We report here the development of new monoclonal antibodies to full length and cleaved SP-D. We have assessed these and existing antibodies in 98 individuals with COPD recruited to the Evaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints (ECLIPSE) cohort. Our data show that neither monoclonal antibodies to full length nor cleaved SP-D provide additional information over that obtained with a polyclonal antibody. Moreover, levels of serum nitrosylated-SP-D did not correlate with serum level of SP-D or any clinical phenotype of COPD. The measurement of modified SP-D is of limited value in characterising individuals with COPD.

  3. Should we disclose amyloid imaging results to cognitively normal individuals?

    OpenAIRE

    Grill, Joshua D.; Johnson, David K.; Burns, Jeffrey M.

    2013-01-01

    Demonstration of brain accumulation of fibrillar amyloid beta protein via positron emission tomography (PET) with amyloid specific ligands may support the diagnosis of Alzheimer's disease (AD). There is increasing recognition of the potential use of amyloid imaging to detect in vivo the pathology of AD in individuals with no ostensible cognitive impairment. Research use of amyloid PET in cognitively normal patients will be key to pursuit of therapies able to delay cognitive impairment and dem...

  4. Towards a Pharmacophore for Amyloid

    Energy Technology Data Exchange (ETDEWEB)

    Landau, Meytal; Sawaya, Michael R.; Faull, Kym F.; Laganowsky, Arthur; Jiang, Lin; Sievers, Stuart A.; Liu, Jie; Barrio, Jorge R.; Eisenberg, David (UCLA)

    2011-09-16

    Diagnosing and treating Alzheimer's and other diseases associated with amyloid fibers remains a great challenge despite intensive research. To aid in this effort, we present atomic structures of fiber-forming segments of proteins involved in Alzheimer's disease in complex with small molecule binders, determined by X-ray microcrystallography. The fiber-like complexes consist of pairs of {beta}-sheets, with small molecules binding between the sheets, roughly parallel to the fiber axis. The structures suggest that apolar molecules drift along the fiber, consistent with the observation of nonspecific binding to a variety of amyloid proteins. In contrast, negatively charged orange-G binds specifically to lysine side chains of adjacent sheets. These structures provide molecular frameworks for the design of diagnostics and drugs for protein aggregation diseases. The devastating and incurable dementia known as Alzheimer's disease affects the thinking, memory, and behavior of dozens of millions of people worldwide. Although amyloid fibers and oligomers of two proteins, tau and amyloid-{beta}, have been identified in association with this disease, the development of diagnostics and therapeutics has proceeded to date in a near vacuum of information about their structures. Here we report the first atomic structures of small molecules bound to amyloid. These are of the dye orange-G, the natural compound curcumin, and the Alzheimer's diagnostic compound DDNP bound to amyloid-like segments of tau and amyloid-{beta}. The structures reveal the molecular framework of small-molecule binding, within cylindrical cavities running along the {beta}-spines of the fibers. Negatively charged orange-G wedges into a specific binding site between two sheets of the fiber, combining apolar binding with electrostatic interactions, whereas uncharged compounds slide along the cavity. We observed that different amyloid polymorphs bind different small molecules, revealing that a cocktail of compounds may be required for future amyloid therapies. The structures described here start to define the amyloid pharmacophore, opening the way to structure-based design of improved diagnostics and therapeutics.

  5. Administration of COG1410 reduces axonal amyloid precursor protein immunoreactivity and microglial activation after controlled cortical impact in mice.

    Science.gov (United States)

    Jiang, Yong; Brody, David L

    2012-09-01

    Traumatic axonal injury (TAI) accounts for at least 35% of the morbidity and mortality in traumatic brain injury (TBI) patients without space-occupying lesions. It is also believed to be a key determinant of adverse outcomes such as cognitive dysfunction across the spectrum of TBI severity. Previous studies have shown that COG1410, a synthetic peptide derived from the apolipoprotein E (apoE) receptor binding region, has anti-inflammatory effects after experimental TBI, with improvements in cognitive recovery. However, the effects of COG1410 on axonal injury following TBI are not known. The current study evaluated the effects of 1?mg/kg daily COG1410 versus saline administered intravenously starting 30?min after controlled cortical impact (CCI) injury on pericontusional TAI in young, wild-type C57BL6/J male mice. We found that COG1410 did not affect the number of amyloid precursor protein (APP)-immunoreactive axonal varicosities in the pericontusional corpus callosum and external capsule at 24?h, but reduced APP-immunoreactive varicosities by 31% at 3 days (p=0.0023), and 36% at 7 days (p=0.0009). COG1410 significantly reduced the number of Iba1-positive cells with activated microglial morphology at all three time points by 21-30%. There was no effect of COG1410 on pericontusional white matter volume or silver staining at any time point. This indicates a possible effect of COG1410 on delayed but not immediate TAI. Future studies are needed to investigate the underlying mechanisms, therapeutic time window, and physiological implications of this effect. PMID:22676717

  6. Ionic self-complementarity induces amyloid-like fibril formation in an isolated domain of a plant copper metallochaperone protein

    Directory of Open Access Journals (Sweden)

    Salom David

    2004-06-01

    Full Text Available Abstract Background Arabidopsis thaliana copper metallochaperone CCH is a functional homologue of yeast antioxidant ATX1, involved in cytosolic copper transport. In higher plants, CCH has to be transported to specialised cells through plasmodesmata, being the only metallochaperone reported to date that leaves the cell where it is synthesised. CCH has two different domains, the N-terminal domain conserved among other copper-metallochaperones and a C-terminal domain absent in all the identified non-plant metallochaperones. The aim of the present study was the biochemical and biophysical characterisation of the C-terminal domain of the copper metallochaperone CCH. Results The conformational behaviour of the isolated C-domain in solution is complex and implies the adoption of mixed conformations in different environments. The ionic self-complementary peptide KTEAETKTEAKVDAKADVE, derived from the C-domain of CCH, adopts and extended conformation in solution with a high content in ?-sheet structure that induces a pH-dependent fibril formation. Freeze drying electron microscopy studies revealed the existence of well ordered amyloid-like fibrils in preparations from both the C-domain and its derivative peptide. Conclusion A number of proteins related with copper homeostasis have a high tendency to form fibrils. The determinants for fibril formation, as well as the possible physiological role are not fully understood. Here we show that the plant exclusive C-domain of the copper metallochaperone CCH has conformational plasticity and forms fibrils at defined experimental conditions. The putative influence of these properties with plant copper delivery will be addressed in the future.

  7. Depolymerization of insulin amyloid fibrils by albumin-modified magnetic fluid

    International Nuclear Information System (INIS)

    Pathogenesis of amyloid-related diseases is associated with the presence of protein amyloid deposits. Insulin amyloids have been reported in a patient with diabetes undergoing treatment by injection of insulin and causes problems in the production and storage of this drug and in application of insulin pumps. We have studied the interference of insulin amyloid fibrils with a series of 18 albumin magnetic fluids (MFBSAs) consisting of magnetite nanoparticles modified by different amounts of bovine serum albumin (w/w BSA/Fe3O4 from 0.005 up to 15). We have found that MFBSAs are able to destroy amyloid fibrils in vitro. The extent of fibril depolymerization was affected by nanoparticle physical–chemical properties (hydrodynamic diameter, zeta potential and isoelectric point) determined by the BSA amount present in MFBSAs. The most effective were MFBSAs with lower BSA/Fe3O4 ratios (from 0.005 to 0.1) characteristic of about 90% depolymerizing activity. For the most active magnetic fluids (ratios 0.01 and 0.02) the DC50 values were determined in the range of low concentrations, indicating their ability to interfere with insulin fibrils at stoichiometric concentrations. We assume that the present findings represent a starting point for the application of the active MFBSAs as therapeutic agents targeting insulin amyloidosis. (paper)

  8. Using BAC transgenesis in zebrafish to identify regulatory sequences of the amyloid precursor protein gene in humans

    Directory of Open Access Journals (Sweden)

    Shakes Leighcraft A

    2012-09-01

    Full Text Available Abstract Background Non-coding DNA in and around the human Amyloid Precursor Protein (APP gene that is central to Alzheimer’s disease (AD shares little sequence similarity with that of appb in zebrafish. Identifying DNA domains regulating expression of the gene in such situations becomes a challenge. Taking advantage of the zebrafish system that allows rapid functional analyses of gene regulatory sequences, we previously showed that two discontinuous DNA domains in zebrafish appb are important for expression of the gene in neurons: an enhancer in intron 1 and sequences 28–31 kb upstream of the gene. Here we identify the putative transcription factor binding sites responsible for this distal cis-acting regulation, and use that information to identify a regulatory region of the human APP gene. Results Functional analyses of intron 1 enhancer mutations in enhancer-trap BACs expressed as transgenes in zebrafish identified putative binding sites of two known transcription factor proteins, E4BP4/ NFIL3 and Forkhead, to be required for expression of appb. A cluster of three E4BP4 sites at ?31?kb is also shown to be essential for neuron-specific expression, suggesting that the dependence of expression on upstream sequences is mediated by these E4BP4 sites. E4BP4/ NFIL3 and XFD1 sites in the intron enhancer and E4BP4/ NFIL3 sites at ?31?kb specifically and efficiently bind the corresponding zebrafish proteins in vitro. These sites are statistically over-represented in both the zebrafish appb and the human APP genes, although their locations are different. Remarkably, a cluster of four E4BP4 sites in intron 4 of human APP exists in actively transcribing chromatin in a human neuroblastoma cell-line, SHSY5Y, expressing APP as shown using chromatin immunoprecipitation (ChIP experiments. Thus although the two genes share little sequence conservation, they appear to share the same regulatory logic and are regulated by a similar set of transcription factors. Conclusion The results suggest that the clock-regulated and immune system modulator transcription factor E4BP4/ NFIL3 likely regulates the expression of both appb in zebrafish and APP in humans. It suggests potential human APP gene regulatory pathways, not on the basis of comparing DNA primary sequences with zebrafish appb but on the model of conservation of transcription factors.

  9. The OspE-Related Proteins Inhibit Complement Deposition and Enhance Serum Resistance of Borrelia burgdorferi, the Lyme Disease Spirochete ?

    OpenAIRE

    Kenedy, Melisha R; Akins, Darrin R

    2011-01-01

    Borrelia burgdorferi, the Lyme disease spirochete, binds the host complement inhibitors factor H (FH) and FH-like protein 1 (FHL-1). Binding of FH/FHL-1 by the B. burgdorferi proteins CspA and the OspE-related proteins is thought to enhance resistance to serum-mediated killing. While previous reports have shown that CspA confers serum resistance in B. burgdorferi, it is unclear whether the OspE-related proteins are relevant in B. burgdorferi serum resistance when OspE is expressed on the borr...

  10. Serum-dependent expression of promyelocytic leukemia protein suppresses propagation of influenza virus

    International Nuclear Information System (INIS)

    The rate of propagation of influenza virus in human adenocarcinoma Caco-2 cells was found to negatively correlate with the concentration of fetal bovine serum (FBS) in the culture medium. Virus replicated more rapidly at lower FBS concentrations (0 or 2%) than at higher concentrations (10 or 20%) during an early stage of infection. Basal and interferon (IFN)-induced levels of typical IFN-inducible anti-viral proteins, such as 2',5'-oligoadenylate synthetase, dsRNA-activated protein kinase and MxA, were unaffected by variation in FBS concentrations. But promyelocytic leukemia protein (PML) was expressed in a serum-dependent manner. In particular, the 65 to 70 kDa isoform of PML was markedly upregulated following the addition of serum. In contrast, other isoforms were induced by IFN treatment, and weakly induced by FBS concentrations. Immunofluorescence microscopy indicated that PML was mainly forme