Elizabeth Drolle

Effect of Surfaces on Amyloid Fibril Formation

Using atomic force microscopy (AFM) we investigated the interaction of amyloid beta (Aβ) (1–42) peptide with chemically modified surfaces in order to better understand the mechanism of amyloid toxicity, which involves interaction of amyloid with cell membrane surfaces. We compared the structure and density of Aβ fibrils on positively and negatively charged as well as hydrophobic chemically-modified surfaces at physiologically relevant conditions. We report that due to the complex distribution of charge and hydrophobicity amyloid oligomers bind to all types of surfaces investigated (CH3, COOH, and NH2) although the charge and hydrophobicity of surfaces affected the structure and size of amyloid deposits as well as surface coverage. Hydrophobic surfaces promote formation of spherical amorphous clusters, while charged surfaces promote protofibril formation. We used the nonlinear Poisson-Boltzmann equation (PBE) approach to analyze the electrostatic interactions of amyloid monomers and oligomers with modified surfaces to complement our AFM data.

Atomic force microscopy to study molecular mechanisms of amyloid fibril formation and toxicity in Alzheimer’s disease

Abstract Alzheimer’s disease (AD) is a devastating neurodegenerative disease characterized by dementia and memory loss for which no cure or effective prevention is currently available. Neurodegeneration in AD is linked to formation of amyloid plaques found in brain tissues of Alzheimer’s patients during post-mortem examination. Amyloid plaques are composed of amyloid fibrils and small oligomers – insoluble protein aggregates. Although amyloid plaques are found on the neuronal cell surfaces, the mechanism of amyloid toxicity is still not well understood. Currently, it is believed that the cytotoxicity is a result of the nonspecific interaction of small soluble amyloid oligomers (rather than longer fibrils) with the plasma membrane. In recent years, nanotechnology has contributed significantly to understanding the structure and function of lipid membranes and to the study of the molecular mechanisms of membrane-associated diseases. We review the current state of research, including applications of the latest nanotechnology approaches, on the interaction of lipid membranes with the amyloid-β (Aβ) peptide in relation to amyloid toxicity. We discuss the interactions of Aβ with model lipid membranes with a focus to demonstrate that composition, charge and phase of the lipid membrane, as well as lipid domains and rafts, affect the binding of Aβ to the membrane and contribute to toxicity. Understanding the role of the lipid membrane in AD at the nanoscale and molecular level will contribute to the understanding of the molecular mechanism of amyloid toxicity and may aid into the development of novel preventive strategies to combat AD.

Effect of melatonin and cholesterol on the structure of DOPC and DPPC membranes

The cell membrane plays an important role in the molecular mechanism of amyloid toxicity associated with Alzheimers disease. The membranes chemical composition and the incorporation of small molecules, such as melatonin and cholesterol, can alter its structure and physical properties, thereby affecting its interaction with amyloid peptides. Both melatonin and cholesterol have been recently linked to amyloid toxicity. Melatonin has been shown to have a protective role against amyloid toxicity. However, the underlying molecular mechanism of this protection is still not well understood, and cholesterols role remains controversial. We used small-angle neutron diffraction (SAND) from oriented lipid multi-layers, small-angle neutron scattering (SANS) from unilamellar vesicles experiments and Molecular Dynamics (MD) simulations to elucidate non-specific interactions of melatonin and cholesterol with 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) model membranes. We conclude that melatonin decreases the thickness of both model membranes by disordering the lipid hydrocarbon chains, thus increasing membrane fluidity. This result is in stark contrast to the much accepted ordering effect induced by cholesterol, which causes membranes to thicken.

Nanoscale electrostatic domains in cholesterol-laden lipid membranes create a target for amyloid binding.

Amyloid fibrils are associated with multiple neurodegenerative disorders, such as Alzheimers disease. Although biological membranes are involved in fibril plaque formation, the role of lipid membrane composition in fibril formation and toxicity is not well understood. We investigated the effect of cholesterol on the interaction of model lipid membranes with amyloid-β peptide (Aβ). With atomic force microscopy we demonstrated that binding of Aβ (1-42) to DOPC bilayer, enriched with 20% cholesterol, resulted in an intriguing formation of small nonuniform islands loaded with Aβ. We attribute this effect to the presence of nanoscale electrostatic domains induced by cholesterol in DOPC bilayers. Using frequency-modulated Kelvin probe force microscopy we were able to resolve these nanoscale electrostatic domains in DOPC monolayers. These findings directly affect our understanding of how the presence of cholesterol may induce targeted binding of amyloid deposits to biomembranes. We postulate that this nonhomogeneous electrostatic effect of cholesterol has a fundamental nature and may be present in other lipid membranes and monolayers.

Effect of cholesterol and amyloid-β peptide on structure and function of mixed-lipid films and pulmonary surfactant BLES: an atomic force microscopy study

Pulmonary surfactant forms a thin molecular film inside mammalian lung alveoli and lowers the surface tension of the air/fluid interface to reduce the work of breathing. Upon compression functional surfactant forms characteristic multilayer structures, which indicate surfactant surface activity. We showed that cholesterol adversely affects both structural and surface-active properties of BLES surfactant and DPPC/DOPG lipid films. Incorporation of small concentrations of fibril-forming peptide amyloid-β 1-40 helps to counteract the distractive effect of cholesterol by improving characteristic multilayer formation that occurs upon compression. In contrast to many negative effects of amyloid-forming peptides reported earlier, we report a positive effect of amyloid-β peptide on surfactant function, which may aid in the designing of novel surfactant formulations.

Changes in lipid membranes may trigger amyloid toxicity in Alzheimer’s disease

Amyloid-beta peptides (Aβ), implicated in Alzheimers disease (AD), interact with the cellular membrane and induce amyloid toxicity. The composition of cellular membranes changes in aging and AD. We designed multi-component lipid models to mimic healthy and diseased states of the neuronal membrane. Using atomic force microscopy (AFM), Kelvin probe force microscopy (KPFM) and black lipid membrane (BLM) techniques, we demonstrated that these model membranes differ in their nanoscale structure and physical properties, and interact differently with Aβ1-42. Based on our data, we propose a new hypothesis that changes in lipid membrane due to aging and AD may trigger amyloid toxicity through electrostatic mechanisms, similar to the accepted mechanism of antimicrobial peptide action. Understanding the role of the membrane changes as a key activating amyloid toxicity may aid in the development of a new avenue for the prevention and treatment of AD.

Atomic force microscopy and Langmuir–Blodgett monolayer technique to assess contact lens deposits and human meibum extracts

PURPOSE The purpose of this exploratory study was to investigate the differences in meibomian gland secretions, contact lens (CL) lipid extracts, and CL surface topography between participants with and without meibomian gland dysfunction (MGD). METHODS Meibum study: Meibum was collected from all participants and studied via Langmuir-Blodgett (LB) deposition with subsequent Atomic Force Microscopy (AFM) visualization and surface roughness analysis. CL Study: Participants with and without MGD wore both etafilcon A and balafilcon A CLs in two different phases. CL lipid deposits were extracted and analyzed using pressure-area isotherms with the LB trough and CL surface topographies and roughness values were visualized using AFM. RESULTS Meibum study: Non-MGD participant meibum samples showed larger, circular aggregates with lower surface roughness, whereas meibum samples from participants with MGD showed more lipid aggregates, greater size variability and higher surface roughness. CL Study: Worn CLs from participants with MGD had a few large tear film deposits with lower surface roughness, whereas non-MGD participant-worn lenses had many small lens deposits with higher surface roughness. Balafilcon A pore depths were shallower in MGD participant worn lenses when compared to non-MGD participant lenses. Isotherms of CL lipid extracts from MGD and non-MGD participants showed a seamless rise in surface pressure as area decreased; however, extracts from the two different lens materials produced different isotherms. CONCLUSIONS MGD and non-MGD participant-worn CL deposition were found to differ in type, amount, and pattern of lens deposits. Lipids from MGD participants deposited irregularly whereas lipids from non-MGD participants showed more uniformity.

Comparative study of lens solutions' ability to remove tear constituents.

Purpose The purpose of this study was to use atomic force microscopy to compare and characterize the cleaning abilities of a hydrogen peroxide–based system (HPS) and a polyhexamethylene biguanide–containing multipurpose solution (MPS) at removing in vitro deposited tear film constituents, as well as to determine deposition patterns on various silicone hydrogel contact lenses. Methods Silicone hydrogel materials—balafilcon A (BA), lotrafilcon B (LB), and senofilcon A (SA)—were incubated for 1 week in an artificial tear solution (ATS) containing representative lipids, proteins, and salts from the tear film. Atomic force microscopy was used to resolve each lens before and after being cleaned overnight in HPS or MPS. Atomic force microscopy was used again to resolve HPS/MPS-cleaned lenses, which were reincubated in fresh ATS for 1 week, before and after an overnight clean in their respective cleaning solution. Results Atomic force microscopy imaging was able to characterize lens deposits with high resolution. Lenses incubated in ATS revealed distinct differences in their deposition pattern across lens materials. The surface of BA contained about 20-nm-high deposits, whereas deposit heights up to 150 nm completely occluded the surface of SA. Lotrafilcon B lenses revealed clusters of deposits up to 90 nm. The use of either lens solution left trace amounts of tear film constituents, although components from the MPS were seen adsorbed onto the surface after cleaning. Surface roughness (Ra) measurements revealed a significant difference between ATS-incubated and HPS/MPS-cleaned SA and LB lenses (p < 0.05). Ra between first incubated and HPS/MPS-cleaned reincubated SA and LB was also significant (p < 0.05). Conclusions Unique variations in ATS deposition patterns were seen between lenses with atomic force microscopy. The application of both HPS and MPS removed most visible surface deposits.