A Cyclic Undecamer Peptide Mimics a Turn in Folded Alzheimer Amyloid β and Elicits Antibodies against Oligomeric and Fibrillar Amyloid and Plaques

The 39- to 42-residue amyloid β (Aβ) peptide is deposited in extracellular fibrillar plaques in the brain of patients suffering from Alzheimer's Disease (AD). Vaccination with these peptides seems to be a promising approach to reduce the plaque load but results in a dominant antibody response directed against the N-terminus. Antibodies against the N-terminus will capture Aβ immediately after normal physiological processing of the amyloid precursor protein and therefore will also reduce the levels of non-misfolded Aβ, which might have a physiologically relevant function. Therefore, we have targeted an immune response on a conformational neo-epitope in misfolded amyloid that is formed in advance of Aβ-aggregation. A tetanus toxoid-conjugate of the 11-meric cyclic peptide Aβ(22–28)-YNGK′ elicited specific antibodies in Balb/c mice. These antibodies bound strongly to the homologous cyclic peptide-bovine serum albumin conjugate, but not to the homologous linear peptide-conjugate, as detected in vitro by enzyme-linked immunosorbent assay. The antibodies also bound—although more weakly—to Aβ(1–42) oligomers as well as fibrils in this assay. Finally, the antibodies recognized Aβ deposits in AD mouse and human brain tissue as established by immunohistological staining. We propose that the cyclic peptide conjugate might provide a lead towards a vaccine that could be administered before the onset of AD symptoms. Further investigation of this hypothesis requires immunization of transgenic AD model mice

Synthesis and apllications of multi 13C-labeled hormonal steroids.

Het Respiratory Distress Syndrome (RDS) is een van de belangrijkste oorzaken van de hoge mortaliteit van te vroeg geborenen. Er wordt aangenomen dat RDS veroorzaakt wordt door een niet goed functioneren van de bijnierschors. De beste methode om de werking van de bijnierschors te meten is het bepalen van de productie snelheid van cortisol, het belangrijkste bijnierschorshormoon. Daar het gebruik van radioaktief gemerkt cortisol, nodig voor de bepaling van de productie snelheid, ethisch niet verantwoord is, werd gezocht naar een mogelijk alternatief. In principe kan de productie snelheid van endogene stoffen bepaald worden door gebruik te maken van met stabiele isotopen gernerkte verbindingen. Het doel van dlt onderzoek was om steroid horrnonen te merken met vieer koolstof-dertien atomen. ... Zie: Samenvatting

Flow-chart for improved NOMV production.

<p>Process performance is evaluated with the NonaMen concept, a nonavalent PorA vaccine comprised of three trivalent RL production strains (Δ<i>rmpM</i>–Δ<i>lpxL1</i> mutant strains; Supplementary <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065157#pone.0065157.s002" target="_blank">Table S2</a>). Production of trivalent bulk NOMV is depicted in phases A to H. The transition from one phase to the next requires a specific unit operation (1 to 10). The seedlot (A) is first expanded in shake flask and bioreactor pre-cultures, then used to inoculate the production bioreactor. Harvested cells (B) are concentrated with microfiltration (C), and vesicle release is stimulated with a detergent-free buffer containing the chelating agent EDTA (D). Cells are discarded, and the crude NOMV are concentrated with ultrafiltration (E). Any residual DNA is digested with nuclease (F), and the extract is purified with gel filtration chromatography. The non-sterile bulk (G) is then sterilized by filtration to obtain the bulk NOMV (H). The trivalent product can be stored for at least one year before mixing as nonavalent vaccine and diluting to dose concentration (I).</p

Implications for vaccine development.

<p>A novel approach for the production of sOMV vaccine against N. meningitidis serogroup B is explored by utilizing the effect of cysteine depletion. (A) Biomass concentration (closed circles) is monitored in bioreactor cultivations (time points A to N). Time point G marks onset of stationary growth, caused by depletion of cysteine (open circles). (B) Yield of purified sOMV vaccine (black bars) is compared with eOMV reference vaccine, which uses detergent-free biomass extraction to improve yield (white bars). Several time points before (D, F) and after (I, K, M) cysteine depletion are included. After cysteine depletion, sOMV yield increases gradually to quantities that are comparable to the eOMV reference (no significant difference at time point M). Significant yield differences are indicated with asterikses (p<0.05). ‘NS’ indicates a non-significant difference.</p

Upstream process characteristics.

<p>A) The upstream process is performed at 40 L scale, which is representative for large-scale (800 L <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065157#pone.0065157-Baart1" target="_blank">[34]</a>). Biomass growth (black line) on chemically defined production medium is highly reproducible (regression R² = 0.977). Cultivations from RL production strain 1, 2, and 3 are indicated by squares, circles, and triangles (respectively). Duplicates for each strain are indicated with open and closed symbols, giving a total of 6 cultivations. B) Oxygen consumption is monitored continuously. Cultivations are aligned at the time-point of maximal oxygen consumption (t = 0), which represents onset of the stationary phase. Harvest point of the cultivations has previously been optimized at t = 3 hours <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065157#pone.0065157-vandeWaterbeemd2" target="_blank">[35]</a>.</p