A combinational treatment of carotenoids decreases Aβ secretion in human neurons via β-secretase inhibition

Sho, Misato; Ichiyanagi, Naoki; Imaizumi, Kent; Ishikawa, Mitsuru; Morimoto, Satoru; Watanabe, Hirotaka

doi:10.1016/j.neures.2019.10.006

Cited by 7 publications

(10 citation statements)

References 42 publications

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“…γ-Secretase is widely expressed in nearly all cell types of multicellular organisms. However, the expression of every subunit of γ-secretase in human neural development remains to be determined, whereas many studies have focused Aβ production in human iPSC-derived neurons (Yagi et al, 2011;Yahata et al, 2011;Israel et al, 2012;Sho et al, 2019). We first examined how γ-secretase subunits are expressed throughout human neural differentiation using the efficient and robust differentiation protocol of iPSC-derived cortical neurons (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Since then, substantial studies have reported many pathological AD phenotypes using iPSC-derived neural cells from AD patients, including FAD (Penney et al, 2020). Mutations in the PS gene expectedly led to a significant increase in the Aβ42/40 ratio and, in some cases, tau pathology in iPSC-derived neurons (Yagi et al, 2011;Woodruff et al, 2013;Imaizumi et al, 2015;Kondo et al, 2017;Sho et al, 2019;Ishikawa et al, 2020). Despite extensive investigations, however, PS physiological functions from the aspect of PS/γ-secretase heterogeneity remain to be investigated in human neural cells.…”

Section: Introductionmentioning

confidence: 99%

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Flexible and Accurate Substrate Processing with Distinct Presenilin/γ-Secretases in Human Cortical Neurons

Watanabe

Imaizumi

Cai

et al. 2021

eNeuro

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Mutations in the presenilin genes (PS1, PS2) have been linked to the majority of familial Alzheimer's disease (AD). Although great efforts have been made to investigate pathogenic PS mutations, which ultimately cause an increase in the toxic form of β-amyloid (Aβ), the intrinsic physiological functions of PS in human neurons remain to be determined. In this study, to investigate the physiological roles of PS in human neurons, we generated PS1 conditional knockout induced pluripotent stem cells (iPSCs), in which PS1 can be selectively abrogated under Cre transduction with or without additional PS2 knockout. We showed that iPSC-derived neural progenitor cells do not confer a maintenance ability in the absence of both PS1 and PS2, showing the essential role of PS in Notch signaling. We then generated PS-null human cortical neurons, where PS1 was intact until full neuronal differentiation occurred. Aβ40 production was reduced exclusively in human PS1/PS2-null neurons along with a concomitant accumulation of APP-CTFs, whereas Aβ42 was decreased in neurons devoid of PS2. Unlike previous studies in mice, in which APP cleavage is largely attributable to PS1, γ-secretase activity seemed to be comparable between PS1 and PS2. In contrast, cleavage of another substrate, N-cadherin, was impaired only in neurons devoid of PS1. Moreover, PS2/γ-secretase exists largely in late endosomes/lysosomes, as measured by specific antibody against the γ-secretase complex, in which Aβ42 species are supposedly produced. Using this novel stem cell-based platform, we assessed important physiological PS1/PS2 functions in mature human neurons, the dysfunction of which could underlie AD pathogenesis. Significance Statement Presenilins are crucial catalytic subunits of γ-secretase, an intramembranous protease complex, whose mutations underlie AD pathogenesis via the dysregulation of Aβ generation. The γ-secretase complex exhibits heterogeneity via the assembly of PS1 or PS2, but the correlation of γ-secretase heterogeneity with substrate processing remains to be established in human neurons. Here, using a novel iPSC-derived cellular model carrying PS1 and/or PS2 conditional knockout alleles, we uncovered the unique processing of three substrates, Notch, APP and N-cadherin, by PS1 or PS2 in human neural cell contexts. Furthermore, the intrinsic subcellular localization of γ-secretase depends on PS1 or PS2, leading to putative differences in the processing of substrates. This novel 3 platform will help ensure the correlation of γ-secretase/substrates in human neurons.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Flexible and Accurate Substrate Processing with Distinct Presenilin/γ-Secretases in Human Cortical Neurons

Watanabe

Imaizumi

Cai

et al. 2021

eNeuro

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“…These results were possibly significant for the analysis of pathophysiology in neuronal diseases. We used iPSCs from FAD patients [34,35], which were acquired after the disease phenotypes had become apparent in the patients. Ngn2+miRs promoted extracellular secretion of Aβ40 and Aβ42 from early stages of culture, resulting in clearly higher Aβ42/40 ratios in patients with PS1 and PS2 mutations.…”

Section: Discussionmentioning

confidence: 99%

“…Familial AD (FAD) is most frequently caused by mutations in the genes encoding the γ-secretase-component, one of the three components of the amyloid precursor protein (APP) processing pathway, which is encoded by the presenilin-1 (PS1) and presenilin-2 (PS2) genes, or by mutations in the APP gene itself, whereas a growing consensus suggests that sporadic AD (SAD) is more likely caused by impaired clearance of Aβ. We have already established iPS cells from patients with mutations in PS1 and PS2 and have observed a phenotypic expression of high extracellular release of Aβ42 [34,35]. In regard to the method of this study, we determined that overexpression of Bcl-xL would mask the consequences of the stress response to cells, which would not be appropriate for the evaluation of disease phenotypes.…”

Section: Early Expression Of Neuronal Phenotypes Of Alzheimer's Diseamentioning

confidence: 99%

“…(n = 5 independent experiments; mean ± SEM; # p < 0. First, as we have done in several previous reports [34,35], we quantified the amount of Aβ40 and Aβ42 in the cell culture supernatant by ELISA. Consequently, on day 20, the amounts of both Aβ species were increased with the addition of miRs, regardless of whether the cells were healthy or affected with AD, which was almost equivalent to the amount in the miRs-group on day 40.…”

Section: Early Expression Of Neuronal Phenotypes Of Alzheimer's Diseamentioning

confidence: 99%

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miRNA-Based Rapid Differentiation of Purified Neurons from hPSCs Advancestowards Quick Screening for Neuronal Disease Phenotypes In Vitro

Ishikawa

Aoyama

Shibata

et al. 2020

Cells

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Obtaining differentiated cells with high physiological functions by an efficient, but simple and rapid differentiation method is crucial for modeling neuronal diseases in vitro using human pluripotent stem cells (hPSCs). Currently, methods involving the transient expression of one or a couple of transcription factors have been established as techniques for inducing neuronal differentiation in a rapid, single step. It has also been reported that microRNAs can function as reprogramming effectors for directly reprogramming human dermal fibroblasts to neurons. In this study, we tested the effect of adding neuronal microRNAs, miRNA-9/9*, and miR-124 (miR-9/9*-124), for the neuronal induction method of hPSCs using Tet-On-driven expression of the Neurogenin2 gene (Ngn2), a proneural factor. While it has been established that Ngn2 can facilitate differentiation from pluripotent stem cells into neurons with high purity due to its neurogenic effect, a long or indefinite time is required for neuronal maturation with Ngn2 misexpression alone. With the present method, the cells maintained a high neuronal differentiation rate while exhibiting increased gene expression of neuronal maturation markers, spontaneous calcium oscillation, and high electrical activity with network bursts as assessed by a multipoint electrode system. Moreover, when applying this method to iPSCs from Alzheimer’s disease (AD) patients with presenilin-1 (PS1) or presenilin-2 (PS2) mutations, cellular phenotypes such as increased amount of extracellular secretion of amyloid β42, abnormal oxygen consumption, and increased reactive oxygen species in the cells were observed in a shorter culture period than those previously reported. Therefore, it is strongly anticipated that the induction method combining Ngn2 and miR-9/9*-124 will enable more rapid and simple screening for various types of neuronal disease phenotypes and promote drug discovery.

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Amyloid β (Aβ) ELISA of Human iPSC-Derived Neuronal Cultures

Watanabe

Murakami

2021

Methods in Molecular Biology

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A combinational treatment of carotenoids decreases Aβ secretion in human neurons via β-secretase inhibition

Cited by 7 publications

References 42 publications

Flexible and Accurate Substrate Processing with Distinct Presenilin/γ-Secretases in Human Cortical Neurons

Flexible and Accurate Substrate Processing with Distinct Presenilin/γ-Secretases in Human Cortical Neurons

miRNA-Based Rapid Differentiation of Purified Neurons from hPSCs Advancestowards Quick Screening for Neuronal Disease Phenotypes In Vitro

Amyloid β (Aβ) ELISA of Human iPSC-Derived Neuronal Cultures

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