The Effect of a Novel Mica Nanoparticle, STB-MP, on an Alzheimer's Disease Patient-Induced PSC-Derived Cortical Brain Organoid Model

Abstract

Alzheimer's disease (AD) is one of the most well-known neurodegenerative diseases, with a substantial amount of advancements in the field of neuroscience and AD. Despite such progress, there has been no significant improvement in AD treatments. To improve in developing a research platform for AD treatment, AD patient-derived induced pluripotent stem cell (iPSC) was employed to generate cortical brain organoids, expressing AD phenotypes, with the accumulation of amyloid-beta (Aβ) and hyperphosphorylated tau (pTau). We have investigated the use of a medical grade mica nanoparticle, STB-MP, as a treatment to decrease the expression of AD's major hallmarks. STB-MP treatment did not inhibit the expression of pTau; however, accumulated Aβ plaques were diminished in STB-MP treated AD organoids. STB-MP seemed to activate the autophagy pathway, by mTOR inhibition, and also decreased γ-secretase activity by decreasing pro-inflammatory cytokine levels. To sum up, the development of AD brain organoids successfully mimics AD phenotype expressions, and thus it could be used as a screening platform for novel AD treatment assessments.

Keywords: autophagy; high throughput screening; inflammation; neurodegenerative disease modelling.

Figure 2

STB-MP nanoparticle characterization. (A) AFM image of STB-MP illustrating both the 2D and 3D shapes of the particles. (B) Toxicity testing of STB-MP in different cell lines. Serial dilutions from 1000 μg/mL to 1 μg/mL were used (n = 3). (C) Schematic image illustrating the STB-MP treatment schedule. (D) Representative images of immunostaining for the apoptotic marker c-Cas3 in WT and PS1 cortical organoids at Day 43 and Day 84 (n = 3). Scale bar 50 μm.

Figure A1

WT and PS1 cortical organoid neurogenesis. (A) Images of WT and PS1 cortical organoid neurogenesis, starting from iPSC to d84 matured organoid. Scale bar, 1 mm. (B) Area measured of WT and PS1 cortical organoid at different stages. n = 10, each data point represents a single organoid at a specific timepoint.

Figure A2

Cortical brain organoid size differences after STP-MP treatment. (A) Images of control group and STB-MP treated group of WT and PS1 cortical organoid. Scale bar, 1 cm. (B) Area measured of control and STB-MP treated organoid of WT and PS1 cortical organoid. N = 10, each data point represents a single organoid.

Figure 1

Generation and characterization of WT and PS1 hiPSC-derived cortical brain organoids. (A) Schematic image illustrating the timeline of hiPSC-derived cortical brain organoid development. (B,C) Representative images of immunostaining for cortical organoid markers Tuj1, SOX2, MAP2 and KI67 in sectioned WT and AD organoids at Day 43 and Day 84 (n = 3). Scale bar, 100 μm. (D,E) Representative images of immunostaining for AD phenotype-related markers Tau, pTau and Aβ in sectioned WT and AD organoids at Day 43 and Day 84 (n = 3). Scale bar, 50 μm. * p < 0.05, *** p <  0.001. The data are presented as the mean ± SD.

Figure 3

STB-MP nanoparticle diminishes the AD phenotype in PS1 cortical brain organoids. (A,B) Expression of the AD phenotype markers Tau, p-Tau and Aβ in WT and AD organoids at Day 98 was assessed by Western blotting analysis (n = 3, each data point represents 5 pooled organoids). (C) Lysates of WT and AD organoids were analyzed by ELISA at Day 98. The Aβ42/Aβ40 ratio in the RIPA fraction was measured by ELISA (n = 3, each data point represents 5 pooled organoids). * p  < 0.05. The data are presented as the mean ± SD.

Figure 4

Inflammation and autophagy as mechanisms of action. (A) Quantitative real-time RT–PCR analysis of both pro- and anti-inflammatory cytokine expression was performed on treated and untreated AD organoids at Day 98. (B–D) IFITM3 expression analysis of the treated and untreated AD organoids at Day 98 by quantitative real-time RT–PCR, immunostaining, and Western blot analysis. (E) Western blot analysis and quantification of autophagy-related marker expression in the treated and untreated AD organoids at Day 98. n = 3 or 4, each data point represents 5 pooled organoids, * p  < 0.05, ** p  < 0.01. The data are presented as the mean ± SD.