July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Customized rotating wall vessel bioreactors produce improved retinal organoids with reduced operational costs and less frequent experimental failure
Author Affiliations & Notes
  • Michael Phelan
    Bioengineering, Temple University, Philadelphia, Pennsylvania, United States
    Neurobiology Neurodegeneration & Repair Laboratory, National Eye Institute, Maryland, United States
  • Peter Lelkes
    Bioengineering, Temple University, Philadelphia, Pennsylvania, United States
  • Anand Swaroop
    Neurobiology Neurodegeneration & Repair Laboratory, National Eye Institute, Maryland, United States
  • Footnotes
    Commercial Relationships   Michael Phelan, First Author - Temple University (P); Peter Lelkes, Temple University (P); Anand Swaroop, None
  • Footnotes
    Support  NEI EY000450, EY000474, EY000546, NEI Intramural Research Program EY000450 and EY000546, NASA Grant # 80NSSC18K1480.
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 3316. doi:https://doi.org/
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      Michael Phelan, Peter Lelkes, Anand Swaroop; Customized rotating wall vessel bioreactors produce improved retinal organoids with reduced operational costs and less frequent experimental failure. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3316. doi: https://doi.org/.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Retinal organoids demonstrate genetic expression patterns similar to the developing retina, similar metabolic and stimuli reactivity, and even correct tissue lamination morphology of multiple cell phenotypes. We have previously demonstrated that retinal organoids grown in a rotating wall vessel (RWV) bioreactor exhibited significantly accelerated and improved differentiation. However, due to large media volumes and long protocols, traditional RWV systems can be expensive to operate for prolonged differentiation of human cell lines and can be prone to multiple failure modes. This study has built a next-generation bioreactor and tests the hypothesis that it can produce organoids at reduced volumes, with active prevention of significant failure modes of RWVs, and with additional control of key differentiation parameters.

Methods : This novel bioreactor was purpose-built with advanced laser cutting and 3D printing fabrication techniques. Embryoid bodies were generated from mouse embryonic stem cells and differentiated using the protocol described in our previous RWV studies. After key timepoints, organoids were collected and evaluated by PCR and IHC microscopy.

Results : The novel bioreactor incorporates an off-axis media exchange system enabling a reduction of active media volume from 100 ml to approximately 20 ml while decreasing fluid shear perturbations associated with media perfusion. This modification simultaneously removes nascent bubbles known to induce significant turbulence in RWV bioreactors. The system also incorporates a novel gas exchange column for optimized hypoxic cell culture. Organoid experiments using the novel system are still being optimized but preliminary engineering evaluations indicate the system can meet all necessary design criteria.

Conclusions : Our prototype bioreactor and its preliminary results support our hypothesis that customized bioreactor systems can generate organoids with improved expression and morphology using methodologies suitable for high-throughput studies.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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