P3-038: Atomic Force Microscopy to Study Molecular Mechanisms of Alzheimer's Disease

2016 ◽  
Vol 12 ◽  
pp. P832-P832
Author(s):  
Elizabeth Drolle ◽  
Francis Hane ◽  
Morgan Robinson ◽  
Jennifer Lou ◽  
Brenda Yasie Lee ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Atomic Force Microscopy ◽  
Molecular Mechanisms ◽  
Force Microscopy ◽  
Atomic Force

scholarly journals Gels of Amyloid Fibers

Biomolecules ◽  
2019 ◽  
Vol 9 (6) ◽  
pp. 210 ◽  
Author(s):  
Ruizhi Wang ◽  
Xiaojing Yang ◽  
Lingwen Cui ◽  
Hang Yin ◽  
Shaohua Xu
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Atomic Force Microscopy ◽  
Melting Point ◽  
Protein Concentration ◽  
Size Exclusion ◽  
Amyloid Fibers ◽  
Force Microscopy ◽  
Atomic Force ◽  
3D Network

Protein self-assembly and formation of amyloid fibers is an early event of numerous human diseases. Continuous aggregation of amyloid fibers in vitro produces biogels, which led us to suspect that amyloid plaques and neurofibrillary tangles in Alzheimer’s disease are of biogels in nature. We applied atomic force microscopy, size exclusion chromatography, and differential scanning calorimetry to elucidate the gel’s structure, kinetics of gel formation, and melting point. We found that (1) lysozyme gelation occurs when the protein concentration is above 5 mg/mL; (2) nonfibrous protein concentration decreases and plateaus after three days of gel synthesis reaction; (3) colloidal lysozyme aggregates are detectable by both atomic force microscopy (AFM) and fast protein liquid chromatography (FPLC); (4) the gels are a three-dimensional (3D) network crosslinked by fibers coiling around each other; (5) the gels have a high melting point at around around 110 °C, which is weakly dependent on protein concentration; (6) the gels are conductive under an electric field, and (7) they form faster in the presence than in the absence of salt in the reaction buffer. The potential role of the gels formed by amyloid fibers in amyloidosis, particularly in Alzheimer’s disease was thoroughly discussed, as gels with increased viscosity, are known to restrict bulk flow and then circulation of ions and molecules.

Imidazolium-Based Ionic Liquids Affect Morphology and Rigidity of Living Cells: an Atomic Force Microscopy Study

2018 ◽  
Author(s):  
Massimiliano Galluzzi ◽  
Carsten Schulte ◽  
Paolo Milani ◽  
Alessandro Podestà
Keyword(s):  
Atomic Force Microscopy ◽  
Ionic Liquids ◽  
Cell Membrane ◽  
Molecular Mechanisms ◽  
Single Cells ◽  
Metastatic Cancer ◽  
Toxic Effects ◽  
Force Microscopy ◽  
Atomic Force ◽  
The Impact

The study of the toxicity, biocompatibility, and environmental sustainability of room-temperature Ionic Liquids (ILs) is still in its infancy. Understanding the impact of ILs on living organisms, especially from the aquatic ecosystem, is urgent, since on one side large amounts of these substances are widely employed as solvents in industrial chemical processes, and on the other side evidences of toxic effects of ILs on microorganisms and single cells have been observed. To date, the toxicity of ILs have been investigated by means of macroscopic assays aimed at characterizing the effective concentrations (like the EC50) that cause the dead of a significant fraction of the population of microorganisms and cells. These studies allowed to identify the cell membrane as the first target of the IL interaction, whose effectiveness was correlated to the lipophilicity of the cation, i.e. to the length of the lateral alkyl chain. Our study aimed at characterizing the molecular mechanisms of the toxicity of ILs. To this purpose, we carried out a combined topographic and mechanical analysis by Atomic Force Microscopy of living breast metastatic cancer cells (MDA-MB-231) upon interaction with imidazolium-based ILs. We showed that ILs are able to induce modifications of the overall rigidity (effective Young modulus) and morphology of the cells. Our results demonstrate that ILs act on the physical properties of the cell membrane, and possibly induce cytoskeletal reorganization, already at concentrations below the EC50. These potentially toxic effects are stronger at higher IL concentrations, as well as with longer lateral chains in the cation.<br>

scholarly journals High speed atomic force microscopy to investigate the interactions between toxic Aβ1-42 peptides and model membranes in real time: impact of the membrane composition

Nanoscale ◽  
2019 ◽  
Vol 11 (15) ◽  
pp. 7229-7238 ◽  
Author(s):  
M. Ewald ◽  
S. Henry ◽  
E. Lambert ◽  
C. Feuillie ◽  
C. Bobo ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Atomic Force Microscopy ◽  
Real Time ◽  
High Speed ◽  
Model Membranes ◽  
Membrane Composition ◽  
Force Microscopy ◽  
Atomic Force ◽  
Disease Mechanisms ◽  
Toxic Peptides

For investigating Alzheimer's disease mechanisms, high-speed atomic force microscopy is a proper tool to monitor the interactions between toxic peptides and lipid model membranes.

scholarly journals High-speed atomic force microscopy highlights new molecular mechanism of daptomycin action

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Francesca Zuttion ◽  
Adai Colom ◽  
Stefan Matile ◽  
Denes Farago ◽  
Frédérique Pompeo ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
High Speed ◽  
Molecular Mechanisms ◽  
Molecular Level ◽  
Force Microscopy ◽  
Multi Scale ◽  
Atomic Force ◽  
Level Information ◽  
Bacterial Lipid ◽  
Membrane Deformations

AbstractThe increase in speed of the high-speed atomic force microscopy (HS-AFM) compared to that of the conventional AFM made possible the first-ever visualisation at the molecular-level of the activity of an antimicrobial peptide on a membrane. We investigated the medically prescribed but poorly understood lipopeptide Daptomycin under infection-like conditions (37 °C, bacterial lipid composition and antibiotic concentrations). We confirmed so far hypothetical models: Dap oligomerization and the existence of half pores. Moreover, we detected unknown molecular mechanisms: new mechanisms to form toroidal pores or to resist Dap action, and to unprecedently quantify the energy profile of interacting oligomers. Finally, the biological and medical relevance of the findings was ensured by a multi-scale multi-nativeness—from the molecule to the cell—correlation of molecular-level information from living bacteria (Bacillus subtilis strains) to liquid-suspended vesicles and supported-membranes using electron and optical microscopies and the lipid tension probe FliptR, where we found that the cells with a healthier state of their cell wall show smaller membrane deformations.

Imidazolium-Based Ionic Liquids Affect Morphology and Rigidity of Living Cells: an Atomic Force Microscopy Study

2018 ◽  
Author(s):  
Massimiliano Galluzzi ◽  
Carsten Schulte ◽  
Paolo Milani ◽  
Alessandro Podestà
Keyword(s):  
Atomic Force Microscopy ◽  
Ionic Liquids ◽  
Cell Membrane ◽  
Molecular Mechanisms ◽  
Single Cells ◽  
Metastatic Cancer ◽  
Toxic Effects ◽  
Force Microscopy ◽  
Atomic Force ◽  
The Impact

The study of the toxicity, biocompatibility, and environmental sustainability of room-temperature Ionic Liquids (ILs) is still in its infancy. Understanding the impact of ILs on living organisms, especially from the aquatic ecosystem, is urgent, since on one side large amounts of these substances are widely employed as solvents in industrial chemical processes, and on the other side evidences of toxic effects of ILs on microorganisms and single cells have been observed. To date, the toxicity of ILs have been investigated by means of macroscopic assays aimed at characterizing the effective concentrations (like the EC50) that cause the dead of a significant fraction of the population of microorganisms and cells. These studies allowed to identify the cell membrane as the first target of the IL interaction, whose effectiveness was correlated to the lipophilicity of the cation, i.e. to the length of the lateral alkyl chain. Our study aimed at characterizing the molecular mechanisms of the toxicity of ILs. To this purpose, we carried out a combined topographic and mechanical analysis by Atomic Force Microscopy of living breast metastatic cancer cells (MDA-MB-231) upon interaction with imidazolium-based ILs. We showed that ILs are able to induce modifications of the overall rigidity (effective Young modulus) and morphology of the cells. Our results demonstrate that ILs act on the physical properties of the cell membrane, and possibly induce cytoskeletal reorganization, already at concentrations below the EC50. These potentially toxic effects are stronger at higher IL concentrations, as well as with longer lateral chains in the cation.<br>

scholarly journals Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid beta 1-42

2019 ◽  
Vol 2 (2) ◽  
pp. 48 ◽  
Author(s):  
Priya Prakash ◽  
Travis C. Lantz ◽  
Krupal P. Jethava ◽  
Gaurav Chopra
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Amyloid Beta ◽  
High Performance ◽  
Force Microscopy ◽  
Atomic Force ◽  
Short Period ◽  
One Step

Amyloid plaques found in the brains of Alzheimer’s disease patients primarily consists of amyloid beta 1-42 (Aβ42). Commercially, Aβ42 is synthesized using high-throughput peptide synthesizers resulting in the presence of impurities and the racemization of amino acids that affects its aggregation properties. Furthermore, the repeated purchase of even a small quantity (~1 mg) of commercial Aβ42 can be expensive for academic researchers. Here, we describe a detailed methodology for robust expression of recombinant human Aβ(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli using standard molecular biology techniques with refined and rapid one-step analytical purification techniques. The peptide is isolated and purified from transformed cells using an optimized reverse-phase high-performance liquid chromatography (HPLC) protocol with commonly available C18 columns, yielding high amounts of peptide (~15–20 mg per 1 L culture) within a short period of time. The recombinant human Aβ(M1-42) forms characteristic aggregates similar to synthetic Aβ42 aggregates as verified by western blotting and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique produces pure recombinant human Aβ(M1-42) that may be used to synthesize chemical probes and in several downstream in vitro and in vivo assays to facilitate Alzheimer’s disease research.

scholarly journals Taking nanomedicine teaching into practice with atomic force microscopy and force spectroscopy

2015 ◽  
Vol 39 (4) ◽  
pp. 360-366 ◽  
Author(s):  
Filomena A. Carvalho ◽  
Teresa Freitas ◽  
Nuno C. Santos
Keyword(s):  
Atomic Force Microscopy ◽  
Single Molecule ◽  
Molecular Mechanisms ◽  
Force Spectroscopy ◽  
Force Microscopy ◽  
Atomic Force ◽  
Human Blood Cells ◽  
Hands On ◽  
Molecule Interactions ◽  
Near Future

Atomic force microscopy (AFM) is a useful and powerful tool to study molecular interactions applied to nanomedicine. The aim of the present study was to implement a hands-on atomic AFM course for graduated biosciences and medical students. The course comprises two distinct practical sessions, where students get in touch with the use of an atomic force microscope by performing AFM scanning images of human blood cells and force spectroscopy measurements of the fibrinogen-platelet interaction. Since the beginning of this course, in 2008, the overall rating by the students was 4.7 (out of 5), meaning a good to excellent evaluation. Students were very enthusiastic and produced high-quality AFM images and force spectroscopy data. The implementation of the hands-on AFM course was a success, giving to the students the opportunity of contact with a technique that has a wide variety of applications on the nanomedicine field. In the near future, nanomedicine will have remarkable implications in medicine regarding the definition, diagnosis, and treatment of different diseases. AFM enables students to observe single molecule interactions, enabling the understanding of molecular mechanisms of different physiological and pathological processes at the nanoscale level. Therefore, the introduction of nanomedicine courses in bioscience and medical school curricula is essential.

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