Scanning ion-conductance microscopy for studying β-amyloid aggregate formation on living cell surface

Abstract

Alzheimers disease (AD) is the most common form of dementia, a progressive neurological disorder characterized by short and long-term memory loss, including cognitive and functional impairment, which is refractory to current therapy. It is suggested that the aggregation of β-amyloid peptide on neuronal cell surface leads to various deviations of its vital function due to myriad pathways defined by internalization of calcium ions, apoptosis promotion, reduction of membrane potential, synaptic activity loss etc. These are associated with structural reorganizations and pathologies of the cell cytoskeleton mainly involving actin filaments and microtubules, and consequently alterations of cell mechanical properties. Thus, the effect of amyloid oligomers on cells Youngs modulus has been observed in a variety of studies. However, the precise connection between the formation of amyloid aggregates on cell membranes and their effects on local mechanical properties of living cells is still unresolved. In this work, we have used correlative scanning ion-conductance microscopy (SICM) to study cell topography, Youngs modulus mapping and confocal imaging of Aβ aggregates formation on living cell surfaces with subsequent assessment of the reactive oxygen species levels inside single cells using platinum nanoelectrodes. We showed that correlative SICM technique, in conjunction with topography mapping and confocal imaging, can be used for Patch-Clamp recordings from living cells with evidently formed FAM-labeled Aβ aggregates on its surface. As we demonstrated, SICM can be successfully applied to studying cytotoxicity mechanisms of Aβ aggregates on living cell surface.

The New Era of High-Throughput Nanoelectrochemistry

“…Their findings revealed that cell surface roughness is unevenly distributed, with the plasma membrane above the nucleus being the smoothest. 130 Leitao et al developed a high-speed SICM based on highbandwidth large-scale piezo actuator able to perform timeresolved, long-term topographical mapping of living eukaryotic cells. 131 They demonstrated continuous surface topography measurement of large areas (80 μm × 80 μm) with good spatial resolution (512 × 512 pixels) and temporal resolution (from 0.5 s/frame to 20 min/frame depending on scan area).…”

“…Gesper used SICM and fluorescence correlation spectroscopy to assess diffusion in cell membranes. Their findings revealed that cell surface roughness is unevenly distributed, with the plasma membrane above the nucleus being the smoothest …”

“…Their findings revealed that cell surface roughness is unevenly distributed, with the plasma membrane above the nucleus being the smoothest. 130 …”

Synthetic, Cell-Derived, Brain-Derived, and Recombinant β-Amyloid: Modelling Alzheimer’s Disease for Research and Drug Development

“…Such modifications constitute a long list including but not limited to: phosphorylation, isomerisation, citrullination, pyroglutamylation, and covalent cross-linking. Post-translational modifications affect the process of Aβ aggregation, change the structure and the biological effects of the oligomers [ 121 ], and influence Aβ hydrolysis by the angiotensin-converting enzyme thereby promoting its accumulation [ 122 ]. For example, it was shown that isomerisation of the aspartate residue in the seventh position accelerated the development of amyloidosis in vivo [ 67 ] and enhanced the toxic properties of the peptide [ 123 ].…”