Amyloid-beta Fibrillogenesis Seeded by Interface-Induced Peptide Misfolding and Self-Assembly
2010; 98 (10): 2299-2308
Modular injectable matrices based on alginate solution/microsphere mixtures that gel in situ and co-deliver immunomodulatory factors
2009; 5 (4): 969-982
The amphipathicity of the natively unstructured amyloid-beta (Abeta40) peptide may play an important role in its aggregation into beta-sheet rich fibrils, which is linked to the pathogenesis of Alzheimer's disease. Using the air/subphase interface as a model interface, we characterized Abeta's surface activity and its conformation, assembly, and morphology at the interface. Abeta readily adsorbed to the air/subphase interface to form a 20 A thick film and showed a critical micelle concentration of approximately 120 nM. Abeta adsorbed at the air/subphase exhibited in-plane ordering that gave rise to Bragg peaks in grazing-incidence x-ray diffraction measurements. Analysis of the peaks showed that the air/subphase interface induced Abeta to fold into a beta-sheet conformation and to self-assemble into approximately 100 A-sized ordered clusters. The formation of these clusters at the air/subphase interface was not affected by pH, salts, or the presence of sucrose or urea, which are known to stabilize or denature native proteins, suggesting that interface-driven Abeta misfolding and assembly are strongly favored. Furthermore, Abeta at the interface seeded the growth of fibrils in the bulk with a distinct morphology compared to those formed by homogeneous nucleation. Our results indicate that interface-induced Abeta misfolding may serve as a heterogeneous, nucleation-controlled aggregation mechanism for Abeta fibrillogenesis in vivo.
View details for DOI 10.1016/j.bpj.2010.01.056
View details for Web of Science ID 000277858400032
View details for PubMedID 20483339
Injectable dendritic cell-carrying alginate gels for immunization and immunotherapy
2008; 29 (27): 3671-3682
Biocompatible polymer solutions that can crosslink in situ following injection to form stable hydrogels are of interest as depots for sustained delivery of therapeutic factors or cells, and as scaffolds for regenerative medicine. Here, injectable self-gelling alginate formulations obtained by mixing alginate microspheres (as calcium reservoirs) with soluble alginate solutions were characterized for potential use in immunotherapy. Rapid redistribution of calcium ions from microspheres into the surrounding alginate solution led to crosslinking and formation of stable hydrogels. The mechanical properties of the resulting gels correlated with the concentration of calcium-reservoir microspheres added to the solution. Soluble factors such as the cytokine interleukin-2 were readily incorporated into self-gelling alginate matrices by simply mixing them with the formulation prior to gelation. Using alginate microspheres as modular components, strategies for binding immunostimulatory CpG oligonucleotides onto the surface of microspheres were also demonstrated. When injected subcutaneously in the flanks of mice, self-gelling alginate formed soft macroporous gels supporting cellular infiltration and allowing ready access to microspheres carrying therapeutic factors embedded in the matrix. This in situ gelling formulation may thus be useful for stimulating immune cells at desired locales, such as solid tumors or infection sites, as well as for other soft tissue regeneration applications.
View details for DOI 10.1016/j.actbio.2008.11.019
View details for Web of Science ID 000266017200003
View details for PubMedID 19117820
Lipid membrane templates the ordering and induces the fibrillogenesis of Alzheimer's disease amyloid-beta peptide
PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS
2008; 72 (1): 1-24
Dendritic cell vaccines, in which antigen-loaded dendritic cells (DCs) are injected directly into patients to trigger immune responses, are in development as a treatment for cancer and some infectious diseases. In this study, we tested the concept of delivering DCs in an injectable hydrogel matrix, with the aim of harboring dendritic cells for prolonged time periods at a defined site and trapping/concentrating factors secreted by DCs to establish an inflammatory milieu in situ. To achieve these goals, a self-gelling formulation of alginate was developed, obtained by mixing calcium-loaded alginate microspheres with soluble alginate solution and dendritic cells, a formulation that rapidly gelled in vivo. When injected subcutaneously in mice, these alginate 'vaccination nodes' containing activated DCs attracted both host dendritic cells and a large number of T cells to the injection sites over a week in vivo, while some of the inoculated DCs trafficked to the draining lymph nodes. Using an adoptive transfer model to track a defined population of T cells responding to immunization with antigen-loaded DCs, we show that DC/alginate immunization led to recruitment of activated antigen-specific T cells to the alginate matrix, in a manner dependent on the presence of the DCs. This gel/DC immunization system may thus be of interest for immunotherapy to direct the accumulation of immune cells at solid tumors or infection sites in the presence of supporting factors co-delivered by the hydrogel matrix.
View details for DOI 10.1016/j.biomaterials.2008.05.033
View details for Web of Science ID 000258439300007
View details for PubMedID 18565578
The lipid membrane has been shown to mediate the fibrillogenesis and toxicity of Alzheimer's disease (AD) amyloid-beta (Abeta) peptide. Electrostatic interactions between Abeta40 and the phospholipid headgroup have been found to control the association and insertion of monomeric Abeta into lipid monolayers, where Abeta exhibited enhanced interactions with charged lipids compared with zwitterionic lipids. To elucidate the molecular-scale structural details of Abeta-membrane association, we have used complementary X-ray and neutron scattering techniques (grazing-incidence X-ray diffraction, X-ray reflectivity, and neutron reflectivity) in this study to investigate in situ the association of Abeta with lipid monolayers composed of either the anionic lipid 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG), the zwitterionic lipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), or the cationic lipid 1,2-dipalmitoyl 3-trimethylammonium propane (DPTAP) at the air-buffer interface. We found that the anionic lipid DPPG uniquely induced crystalline ordering of Abeta at the membrane surface that closely mimicked the beta-sheet structure in fibrils, revealing an intriguing templated ordering effect of DPPG on Abeta. Furthermore, incubating Abeta with lipid vesicles containing the anionic lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (POPG) induced the formation of amyloid fibrils, confirming that the templated ordering of Abeta at the membrane surface seeded fibril formation. This study provides a detailed molecular-scale characterization of the early structural fluctuation and assembly events that may trigger the misfolding and aggregation of Abeta in vivo. Our results implicate that the adsorption of Abeta to anionic lipids, which could become exposed to the outer membrane leaflet by cell injury, may serve as an in vivo mechanism of templated-aggregation and drive the pathogenesis of AD.
View details for DOI 10.1002/prot.21887
View details for Web of Science ID 000256609800001
View details for PubMedID 18186465