Abstract
An organoid, a self-organizing organ-like tissue developed from stem cells, can exhibit a miniaturized three-dimensional (3D) structure and part of the physiological functions of the original organ. Due to the reproducibility of tissue complexity and ease of handling, organoids have replaced real organs and animals for a variety of uses, such as investigations of the mechanisms of organogenesis and disease onset, and screening of drug effects and/or toxicity. The recent advent of tissue clearing and 3D imaging techniques have great potential contributions to organoid studies by allowing the collection and analysis of 3D images of whole organoids with a reasonable throughput and thus can expand the means of examining the 3D architecture, cellular components, and variability among organoids. Genetic and histological cell-labeling methods, together with organoid clearing, also allow visualization of critical structures and cellular components within organoids. The collected 3D data may enable image analysis to quantitatively assess structures within organoids and sensitively/effectively detect abnormalities caused by perturbations. These capabilities of tissue/organoid clearing and 3D imaging techniques not only extend the utility of organoids in basic biology but can also be applied for quality control of clinical organoid production and large-scale drug screening.
Highlights
Directed differentiation of human pluripotent stem cells (PSCs) to generate target organ cells is currently the most promising method to create artificial organs for regenerative medicine
Organic solvent-based tissue clearing protocols originating from the Spalteholz reagent are represented by the benzyl alcohol/benzyl benzoate (BABB) method, 3D imaging of solventcleared organs (3DISCO), and the ethyl-cinnamate (ECi) method (Dent et al, 1989; Becker et al, 2012; Ertürk et al, 2012; Klingberg et al, 2017)
Regarding the use of organoids to recapitulate microcephaly caused by the Zika virus (ZIKV), forebrain organoid size and neuronal proliferation have been assessed (Dang et al, 2016; Qian et al, 2016)
Summary
Directed differentiation of human pluripotent stem cells (PSCs) to generate target organ cells is currently the most promising method to create artificial organs for regenerative medicine. Tissue clearing protocols for spheroids and organoids have been developed and even applied in recent studies (Costa et al, 2019). Organic solvent-based tissue clearing protocols originating from the Spalteholz reagent are represented by the benzyl alcohol/benzyl benzoate (BABB) method, 3D imaging of solventcleared organs (3DISCO), and the ethyl-cinnamate (ECi) method (Dent et al, 1989; Becker et al, 2012; Ertürk et al, 2012; Klingberg et al, 2017). Brismar and colleagues applied ExM to probe labeling and for high-resolution imaging of tumor cell spheroids, and found that as compared to simpler clearing protocols, ExM improved antibody penetration and image resolution in deeper regions (Edwards et al, 2020).
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