Optical Super-Resolution Imaging of β-Amyloid Aggregation In Vitro and In Vivo: Method and Techniques

2016 ◽  
pp. 125-141 ◽  
Author(s):  
Dorothea Pinotsi ◽  
Gabriele S. Kaminski Schierle ◽  
Clemens F. Kaminski
Keyword(s):  
Super Resolution ◽  
Amyloid Aggregation ◽  
Resolution Imaging ◽  
Β Amyloid

scholarly journals Differential FSH Glycosylation Modulates FSHR Oligomerization and Subsequent cAMP Signaling

2021 ◽  
Vol 12 ◽  
Author(s):  
Uchechukwu T. Agwuegbo ◽  
Emily Colley ◽  
Anthony P. Albert ◽  
Viktor Y. Butnev ◽  
George R. Bousfield ◽  
...  
Keyword(s):  
Super Resolution ◽  
Hek293 Cells ◽  
Camp Signaling ◽  
Camp Production ◽  
Functional Roles ◽  
Menopausal Women ◽  
Resolution Imaging ◽  
Post Menopausal

Follicle-stimulating hormone (FSH) and its target G protein-coupled receptor (FSHR) are essential for reproduction. Recent studies have established that the hypo-glycosylated pituitary FSH glycoform (FSH21/18), is more bioactive in vitro and in vivo than the fully-glycosylated variant (FSH24). FSH21/18 predominates in women of reproductive prime and FSH24 in peri-post-menopausal women, suggesting distinct functional roles of these FSH glycoforms. The aim of this study was to determine if differential FSH glycosylation modulated FSHR oligomerization and resulting impact on cAMP signaling. Using a modified super-resolution imaging technique (PD-PALM) to assess FSHR complexes in HEK293 cells expressing FSHR, we observed time and concentration-dependent modulation of FSHR oligomerization by FSH glycoforms. High eFSH and FSH21/18 concentrations rapidly dissociated FSHR oligomers into monomers, whereas FSH24 displayed slower kinetics. The FSHR β-arrestin biased agonist, truncated eLHβ (Δ121-149) combined with asparagine56-deglycosylated eLHα (dg-eLHt), increased FSHR homomerization. In contrast, low FSH21/18 and FSH24 concentrations promoted FSHR association into oligomers. Dissociation of FSHR oligomers correlated with time points where higher cAMP production was observed. Taken together, these data suggest that FSH glycosylation may modulate the kinetics and amplitude of cAMP production, in part, by forming distinct FSHR complexes, highlighting potential avenues for novel therapeutic targeting of the FSHR to improve IVF outcomes.

P1-174: Early detection of β-amyloid aggregation in in vivo and in vitro models of Alzheimer's disease

2010 ◽  
Vol 6 ◽  
pp. S224-S224 ◽  
Author(s):  
Louise A. Scrocchi ◽  
Elizabeth Karaskov ◽  
Vivian Lee ◽  
Hui Chen ◽  
Melissa Osborne ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Early Detection ◽  
In Vitro Models ◽  
Amyloid Aggregation ◽  
Β Amyloid

Programmable in vitro Co-Encapsidation of Guest Proteins Inside Protein Nanocages

2018 ◽  
Author(s):  
Noor H. Dashti ◽  
Rufika S. Abidin ◽  
Frank Sainsbury
Keyword(s):  
Self Assembly ◽  
Mammalian Cells ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Super Resolution ◽  
Cell Engineering ◽  
Particle Analysis ◽  
Protein Cages

Bioinspired self-sorting and self-assembling systems using engineered versions of natural protein cages have been developed for biocatalysis and therapeutic delivery. The packaging and intracellular delivery of guest proteins is of particular interest for both <i>in vitro</i> and <i>in vivo</i> cell engineering. However, there is a lack of platforms in bionanotechnology that combine programmable guest protein encapsidation with efficient intracellular uptake. We report a minimal peptide anchor for <i>in vivo</i> self-sorting of cargo-linked capsomeres of the Murine polyomavirus (MPyV) major coat protein that enables controlled encapsidation of guest proteins by <i>in vitro</i> self-assembly. Using Förster resonance energy transfer (FRET) we demonstrate the flexibility in this system to support co-encapsidation of multiple proteins. Complementing these ensemble measurements with single particle analysis by super-resolution microscopy shows that the stochastic nature of co-encapsidation is an overriding principle. This has implications for the design and deployment of both native and engineered self-sorting encapsulation systems and for the assembly of infectious virions. Taking advantage of the encoded affinity for sialic acids ubiquitously displayed on the surface of mammalian cells, we demonstrate the ability of self-assembled MPyV virus-like particles to mediate efficient delivery of guest proteins to the cytosol of primary human cells. This platform for programmable co-encapsidation and efficient cytosolic delivery of complementary biomolecules therefore has enormous potential in cell engineering.

scholarly journals Human pluripotent stem cell-derived kidney organoids for personalized congenital and idiopathic nephrotic syndrome modeling

2021 ◽  
Author(s):  
Jitske Jansen ◽  
Bartholomeus T van den Berge ◽  
Martijn van den Broek ◽  
Rutger J Maas ◽  
Brigith Willemsen ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Idiopathic Nephrotic Syndrome ◽  
Super Resolution ◽  
Induced Pluripotent Stem Cell ◽  
Glomerular Injury ◽  
Kidney Organoids

Nephrotic syndrome (NS) is characterized by severe proteinuria as a consequence of kidney glomerular injury due to podocyte damage. In vitro models mimicking in vivo podocyte characteristics are a prerequisite to resolve NS pathogenesis. Here, we report human induced pluripotent stem cell derived kidney organoids containing a podocyte population that heads towards adult podocytes and were superior compared to 2D counterparts, based on scRNA sequencing, super-resolution imaging and electron microscopy. In this study, these next-generation podocytes in kidney organoids enabled personalized idiopathic nephrotic syndrome modeling as shown by activated slit diaphragm signaling and podocyte injury following protamine sulfate treatment and exposure to NS plasma containing pathogenic permeability factors. Organoids cultured from cells of a patient with heterozygous NPHS2 mutations showed poor NPHS2 expression and aberrant NPHS1 localization, which was reversible after genetic correction. Repaired organoids displayed increased VEGFA pathway activity and transcription factor activity known to be essential for podocyte physiology, as shown by RNA sequencing. This study shows that organoids are the preferred model of choice to study idiopathic and congenital podocytopathies.

scholarly journals Design and Synthesis of New Benzo[d]oxazole-Based Derivatives and Their Neuroprotective Effects on β-Amyloid-Induced PC12 Cells

Molecules ◽  
2020 ◽  
Vol 25 (22) ◽  
pp. 5391
Author(s):  
Zheng Liu ◽  
Ming Bian ◽  
Qian-Qian Ma ◽  
Zhuo Zhang ◽  
Huan-Huan Du ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Glycogen Synthase ◽  
B Cell Lymphoma ◽  
Nuclear Factor Κb ◽  
In Vivo Studies ◽  
Neuroprotective Effects ◽  
Β Amyloid ◽  
Gsk 3Β

A series of novel synthetic substituted benzo[d]oxazole-based derivatives (5a–5v) exerted neuroprotective effects on β-amyloid (Aβ)-induced PC12 cells as a potential approach for the treatment of Alzheimer’s disease (AD). In vitro studies show that most of the synthesized compounds were potent in reducing the neurotoxicity of Aβ25-35-induced PC12 cells at 5 μg/mL. We found that compound 5c was non-neurotoxic at 30 μg/mL and significantly increased the viability of Aβ25-35-induced PC12 cells at 1.25, 2.5 and 5 μg/mL. Western blot analysis showed that compound 5c promoted the phosphorylation of Akt and glycogen synthase kinase (GSK-3β) and decreased the expression of nuclear factor-κB (NF-κB) in Aβ25-35-induced PC12 cells. In addition, our findings demonstrated that compound 5c protected PC12 cells from Aβ25-35-induced apoptosis and reduced the hyperphosphorylation of tau protein, and decreased the expression of receptor for AGE (RAGE), β-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1), inducible nitric oxide synthase (iNOS) and Bcl-2-associated X protein/B-cell lymphoma 2 (Bax/Bcl-2) via Akt/GSK-3β/NF-κB signaling pathway. In vivo studies suggest that compound 5c shows less toxicity than donepezil in the heart and nervous system of zebrafish.

scholarly journals Modulation of β-amyloid aggregation by graphene quantum dots

2019 ◽  
Vol 6 (6) ◽  
pp. 190271 ◽  
Author(s):  
Changliang Liu ◽  
Huan Huang ◽  
Lilusi Ma ◽  
Xiaocui Fang ◽  
Chen Wang ◽  
...  
Keyword(s):  
Quantum Dots ◽  
Structural Changes ◽  
Graphene Quantum Dots ◽  
Amyloid Peptide ◽  
Therapeutic Drug ◽  
Amyloid Aggregation ◽  
Β Amyloid ◽  
Regulatory Effects ◽  
Β Amyloid Peptide

Misfolding and abnormal aggregation of β-amyloid peptide is associated with the onset and progress of Alzheimer's disease (AD). Therefore, modulating β-amyloid aggregation is critical for the treatment of AD. Herein, we studied the regulatory effects and mechanism of graphene quantum dots (GQDs) on 1–42 β-amyloid (Aβ 1–42 ) aggregation. GQDs displayed significant regulatory effects on the aggregation of Aβ 1–42 peptide as detected by thioflavin T (ThT) assay. Then, the changes of confirmations and structures induced by GQDs on the Aβ 1–42 aggregation were monitored by circular dichroism (CD), dynamic light scattering (DLS) and transmission electron microscope (TEM). The in vitro cytotoxicity experiments further demonstrated the feasibility of GQDs on the regulation of Aβ 1–42 aggregation. Meanwhile, the structural changes of a Aβ 1–42 /GQDs mixture in different pH revealed that electrostatic interaction was the major driving force in the co-assembly process of Aβ 1–42 and GQDs. The proposed mechanism of the regulatory effects of GQDs on the Aβ 1–42 aggregation was also deduced reasonably. This work not only demonstrated the potential feasibility of GQDs as therapeutic drug for AD but also clarified the regulatory mechanism of GQDs on the Aβ 1–42 aggregation.

Inhibiting β-Amyloid-Associated Alzheimer’s Pathogenesis In Vitro and In Vivo by a Multifunctional Dimeric Bis(12)-hupyridone Derived from Its Natural Analogue

2014 ◽  
Vol 55 (4) ◽  
pp. 1014-1021 ◽  
Author(s):  
Shengquan Hu ◽  
Rui Wang ◽  
Wei Cui ◽  
Zaijun Zhang ◽  
Shinghung Mak ◽  
...  
Keyword(s):  
Natural Analogue ◽  
Β Amyloid

scholarly journals Olive-Oil-Derived Oleocanthal Enhances β-Amyloid Clearance as a Potential Neuroprotective Mechanism against Alzheimer’s Disease: In Vitro and in Vivo Studies

2013 ◽  
Vol 4 (6) ◽  
pp. 973-982 ◽  
Author(s):  
Alaa H. Abuznait ◽  
Hisham Qosa ◽  
Belnaser A. Busnena ◽  
Khalid A. El Sayed ◽  
Amal Kaddoumi
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Olive Oil ◽  
In Vivo Studies ◽  
Β Amyloid
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