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Updated: Dec 25, 2025
Reconstruct Human Retinoblastoma In Vitro
Published on: October 11, 2022
Role of RB1 in human embryonic stem cell-derived retinal organoids
Canbin Zheng1, Jay W Schneider2, Jenny Hsieh3
1Department of Molecular Biology and Hamon Center for Regenerative Science and Medicine, UT Southwestern Medical Center, Dallas, TX, 75390, USA; Department of Orthopedic and Microsurgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, GD, 510080, China.
Abstract:
Three-dimensional (3D) organoid models derived from human pluripotent stem cells provide a platform for studying human development and understanding disease mechanisms. Most studies that examine biallelic inactivation of the cell cycle regulator Retinoblastoma 1 (RB1) and the link to retinoblastoma is in mice, however, less is known regarding the pathophysiological role of RB1 during human retinal development. To study the role of RB1 in early human retinal development and tumor formation, we generated retinal organoids from CRISPR/Cas9-derived RB1-null human embryonic stem cells (hESCs). We showed that RB is abundantly expressed in retinal progenitor cells in retinal organoids and loss of RB1 promotes S-phase entry. Furthermore, loss of RB1 resulted in widespread apoptosis and reduced the number of photoreceptor, ganglion, and bipolar cells. Interestingly, RB1 mutation in retinal organoids did not result in retinoblastoma formation in vitro or in the vitreous body of NOD/SCID immunodeficient mice. Together, our work identifies a crucial function for RB1 in human retinal development and suggests that RB1 deletion alone is not sufficient for tumor development, at least in human retinal organoids.
Insights
Loss of the Retinoblastoma 1 (RB1) gene in human retinal organoids disrupts retinal development but does not cause retinoblastoma, indicating RB1 deletion alone is insufficient for tumor formation.
Area of Science:
- Developmental Biology
- Stem Cell Biology
- Ophthalmology
Background:
- Three-dimensional (3D) organoid models from human pluripotent stem cells are vital for studying human development and disease.
- The role of Retinoblastoma 1 (RB1) in human retinal development and retinoblastoma pathogenesis is not well understood.
- Most research on RB1 inactivation and retinoblastoma involves mouse models.
Purpose of the Study:
- To investigate the function of RB1 in early human retinal development.
- To explore the role of RB1 in retinoblastoma formation using human retinal organoids.
- To understand the pathophysiological consequences of RB1 loss in the human retina.
Main Methods:
- Generation of RB1-null human retinal organoids from human embryonic stem cells (hESCs) using CRISPR/Cas9 gene editing.
- Analysis of RB1 expression and its impact on cell cycle progression (S-phase entry) in retinal progenitor cells.
- Assessment of cell viability (apoptosis) and differentiation of retinal cell types (photoreceptor, ganglion, bipolar cells).
- In vitro and in vivo (NOD/SCID mice) evaluation of retinoblastoma formation potential.
Main Results:
- RB1 is highly expressed in retinal progenitor cells during human retinal development.
- Loss of RB1 function leads to increased S-phase entry and widespread apoptosis.
- RB1 deficiency reduces the number of key retinal cell types, including photoreceptors, ganglion cells, and bipolar cells.
- RB1 mutation in retinal organoids did not induce retinoblastoma formation in vitro or in vivo.
Conclusions:
- RB1 plays a critical role in normal human retinal development.
- RB1 deletion alone is insufficient to cause retinoblastoma in human retinal organoids.
- Further investigation is needed to identify additional factors contributing to retinoblastoma pathogenesis.
