A Cleared View on Retinal Organoids

Abstract

Human induced pluripotent stem cell (hiPSC)-derived organoids mimicking tissues and organs in vitro have advanced medical research, as they opened up new possibilities for in-depth basic research on human organ development as well as providing a human in vitro model for personalized therapeutic approaches. hiPSC-derived retinal organoids have proven to be of great value for modeling the human retina featuring a very similar cellular composition, layering, and functionality. The technically challenging imaging of three-dimensional structures such as retinal organoids has, however, raised the need for robust whole-organoid imaging techniques. To improve imaging of retinal organoids we optimized a passive clearing technique (PACT), which enables high-resolution visualization of fragile intra-tissue structures. Using cleared retinal organoids, we could greatly enhance the antibody labeling efficiency and depth of imaging at high resolution, thereby improving the three-dimensional microscopy output. In that course, we were able to identify the spatial morphological shape and organization of, e.g., photoreceptor cells and bipolar cell layers. Moreover, we used the synaptic protein CtBP2/Ribeye to visualize the interconnection points of photoreceptor and bipolar cells forming the retinal-specific ribbon synapses.

Retinogenesis of the Human Fetal Retina: An Apical Polarity Perspective

“…Improved retinal organoid culturing methods using bioreactors or microfluidic organ-on-a-chip technology, which also allows for tight control of the physiological microenvironment, bring us a further step towards an in vivo like retina [167,215,326]. Furthermore, retinal organoid models derived from patients may be used in conjunction with two-photon imaging and light sheet microscopy as well as tissue clearing methods such as DISCO and PACT [47,169,[327][328][329]. This will allow unparalleled analysis of live and fixed cellular events including spatiotemporal process such as proliferation, differentiation and migration.…”

Dynamic full-field optical coherence tomography: 3D live-imaging of retinal organoids

“…These hiPSC-derived retinal organoids are routinely imaged with various techniques (see Supplementary Table 1 ). However, each of the existing methods presents major drawbacks, such as the need for fixation or mechanical sectioning, rendering the study of dynamic phenomena impossible; the need for labelling, requiring cumbersome and costly preparation; or a lack of functional contrast, indicating only cell presence and not cell health or behaviour 18 – 20 . Optical coherence tomography (OCT) is commonly used in biology and medicine to obtain 3D images of microstructures in tissue.…”

Tissue clearing technique: Recent progress and biomedical applications

“…They also exhibited that intraislet capillary density was positively related to the islet size while islet sphericity was negatively related to the islet size (Fowler et al, 2018). Cao interconnections of photoreceptor cells in three dimensions (Cora et al, 2019).…”