Label-free imaging of amyloid plaques in Alzheimer’s disease with stimulated Raman scattering microscopy

Ji, Minhe; Arbel, Michal; Zhang, Lili; Freudiger, Christian W.; Hou, Steven S.; Lin, Dongdong; Yang, Xin; Bacskai, Brian J.; Xie, Xin

doi:10.1126/sciadv.aat7715

Cited by 146 publications

(147 citation statements)

References 52 publications

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“…We validated the Amyloid- specificity of the 1675 cm -1 peak by co-localization with fluorescent signals from Methoxy-04 (MX04), a molecule that binds to peptides in β-sheet conformation and is commonly used to stain Aβ plaques in Alzheimer's research (see Supplemental Figure S4). Our assignment also agrees with results from two previous studies on AD-affected tissues 6,7 , where similar structures exhibiting the same frequency-shift were observed and validated as Aβ plaques using fluorescent dye labeling with thioflavin S, another stain commonly used to visualize A The ratio of images at taken at 1675 cm −1 and 1665 cm −1 , shown in Figure 3 (A) (iii), therefore provides a clean and specific visualization of Aβ aggregates in plaques on an otherwise homogeneous background. We note that cell nuclei are visible with a very weak contrast in this image as well.…”

Section: Srs Allows For a Clean Label-free Visualization Of Aβ-peptisupporting

confidence: 92%

“…We first sought to identify a way of directly and specifically visualizing the aggregated Aβ content of the pathological structures. We focused on the Amide I (C=O stretch) vibration, which was shown to be particularly sensitive to protein secondary structure in previous spontaneous Raman measurements 23,24 and coherent Raman imaging studies 6,7 .…”

Section: Srs Allows For a Clean Label-free Visualization Of Aβ-peptimentioning

confidence: 99%

“…In recent years, CRS has emerged as a powerful approach for label-free, biochemical imaging of neuronal structures and for dynamic monitoring of functional parameters such as membrane potential 4 and neurotransmitter concentrations 5 . In addition, CRS has been used in fundamental research on neurodegenerative diseases, to characterize Alzheimer's pathology 6,7,8 , Parkinson's Disease 9 and multiple sclerosis 10 , or to study peripheral nerve degeneration in ALS 11 . Currently, there is a major push to develop label-free histopathological analyses based on CRS (sometimes integrated in a multi-modal approach with other nonlinear optical imaging techniques such as two-photon fluorescence or harmonic-generation microscopy) 12 , with major efforts in brain cancer 13,14,15 , other cancers 16,17,18 and a range of other diseases 19,20 .…”

Section: Introductionmentioning

confidence: 99%

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Label-free characterization of Amyloid-β-plaques and associated lipids in brain tissues using stimulated Raman scattering microscopy

Schweikhard¹,

Baral

Krishnamachari³

et al. 2019

Preprint

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The brains of patients with neurodegenerative diseases such as Alzheimer's Disease (AD) often exhibit pathological alterations that involve abnormal aggregations of proteins and lipids. Here, we demonstrate that high-resolution, label-free, chemically-specific imaging using Stimulated Raman Scattering Microscopy (SRS) provides novel insights into the biophysical properties and biochemical composition of such pathological structures. In brain slices of a mouse model of AD, SRS reveals large numbers of Amyloid-β plaques that commonly form a characteristic, three-dimensional core-shell structure, with a fibrillar proteinaceous core surrounded by a halo-like shell of lipid-rich deposits. SRS spectroscopic imaging allows for a clean, label-free visualization of the misfolded (β-sheet) Amyloid-β content in the plaque core. Surrounding lipid-rich deposits are found to contain comparatively high concentrations of membrane lipids (sphingomyelin, phosphatidylcholine), but lower levels of cholesterol than healthy white matter structures. Overall, the SRS spectra of plaque-associated lipids closely resemble those of nearby neurites, with the notable difference of a higher degree of lipid unsaturation compared to healthy brain structures. We hypothesize that plaque-associated lipid deposits may result from neuritic dystrophy associated with AD, and that the observed increased levels of unsaturation could help identify the kinds of pathological alterations taking place. Taken together, our results highlight the potential of Stimulated Raman Scattering microscopy to contribute to a deeper understanding of neurodegenerative diseases.

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Section: Srs Allows For a Clean Label-free Visualization Of Aβ-peptisupporting

confidence: 92%

Section: Srs Allows For a Clean Label-free Visualization Of Aβ-peptimentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Label-free characterization of Amyloid-β-plaques and associated lipids in brain tissues using stimulated Raman scattering microscopy

Schweikhard¹,

Baral

Krishnamachari³

et al. 2019

Preprint

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“…Thus, if several types of amyloid aggregates may exist simultaneously in the neuron, new approaches are required. Label‐free methods such as surface‐enhanced Raman spectroscopy have been used to study the structure of synthetic Aβ fibrils18 and amyloid plaques in brain tissue,19 however, when applied to a single cell, low signal to noise ratio, high autofluorescence, and irreversible photodamage20 make it challenging to reveal molecular structures at the subcellular level.…”

Section: Introductionmentioning

confidence: 99%

Super‐Resolution Infrared Imaging of Polymorphic Amyloid Aggregates Directly in Neurons

et al. 2020

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Loss of memory during Alzheimer's disease (AD), a fatal neurodegenerative disorder, is associated with neuronal loss and the aggregation of amyloid proteins into neurotoxic β‐sheet enriched structures. However, the mechanism of amyloid protein aggregation is still not well understood due to many challenges when studying the endogenous amyloid structures in neurons or in brain tissue. Available methods either require chemical processing of the sample or may affect the amyloid protein structure itself. Therefore, new approaches, which allow studying molecular structures directly in neurons, are urgently needed. A novel approach is tested, based on label‐free optical photothermal infrared super‐resolution microspectroscopy, to study AD‐related amyloid protein aggregation directly in the neuron at sub‐micrometer resolution. Using this approach, amyloid protein aggregates are detected at the subcellular level, along the neurites and strikingly, in dendritic spines, which has not been possible until now. Here, a polymorphic nature of amyloid structures that exist in AD transgenic neurons is reported. Based on the findings of this work, it is suggested that structural polymorphism of amyloid proteins that occur already in neurons may trigger different mechanisms of AD progression.

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“…We started with Amide I band since its Raman vibration was well documented to probe protein secondary structures in vibrational spectroscopy 16 . A recent report by label-free hSRS imaging of Amide I on amyloid plaques in brain tissues revealed a clear 12 cm -1 blue shift (resolvable by our spectral resolution ~12 cm -1 ), corresponding to cross-beta sheet structures 28 . Nonetheless, we observed no such spectroscopic difference from Amide I between mHtt aggregates and cellular background (Fig.…”

Section: Understand the Structures Of Mhtt Aggregates By Hyperspectramentioning

confidence: 49%

Live-Cell Imaging and Quantification of PolyQ Aggregates by Stimulated Raman Scattering of Selective Deuterium Labeling

Miao

2019

Preprint

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Huntington's disease, a major neurodegenerative disorder, involves deposition of aggregationprone proteins with long polyglutamine (polyQ) expansions. The ability to non-perturbatively visualize the formation of aggregates could offer new molecular insight for their pathologic roles. Here, we propose stimulated Raman scattering imaging of deuterium-labeled glutamine to investigate native polyQ aggregates in live cells with subcellular resolution. Through the enrichment of deuterated glutamine in the polyQ sequence of mutant Huntingtin (mHtt) proteins, we first achieved sensitive and specific SRS imaging of carbon-deuterium bonds (C-D) from aggregates without GFP labeling. These aggregates become 1.8-fold denser compared to those with GFP. Second, we performed ratiometric quantification, which revealed a dependence of protein compositions on aggregation sizes. Moreover, we calculated the absolute concentrations for sequestered mHtt and non-mHtt proteins within the same aggregates. Our method may readily reveal new features of polyQ aggregates and could be suited for in vivo investigations on multicellular organisms.

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Label-free imaging of amyloid plaques in Alzheimer’s disease with stimulated Raman scattering microscopy

Abstract: Misfolded proteins in amyloid plaques in transgenic Alzheimer’s disease mouse brains are visualized directly without labeling.

Cited by 146 publications

References 52 publications

Label-free characterization of Amyloid-β-plaques and associated lipids in brain tissues using stimulated Raman scattering microscopy

Label-free characterization of Amyloid-β-plaques and associated lipids in brain tissues using stimulated Raman scattering microscopy

Super‐Resolution Infrared Imaging of Polymorphic Amyloid Aggregates Directly in Neurons

Live-Cell Imaging and Quantification of PolyQ Aggregates by Stimulated Raman Scattering of Selective Deuterium Labeling

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