Evaluation of the Cryopreservation Potential of a Demineralized Bone Paper-Based Bone Organoid

Presenter: Daniel Nguyen

Faculty Sponsor: Jungwoo Lee

School: UMass Amherst

Research Area: Chemical and Biomolecular Engineering

Session: Poster Session 5, 3:15 PM - 4:00 PM, 165, D7

ABSTRACT

The ability to cryopreserve organoids would enhance reproducibility, scalability, and accessibility of complex tissue models. Cryopreserving organoid constructs remains challenging due to diffusion constraints and freeze-thaw-induced functional loss. Certain epithelial organoids, such as intestinal organoids, have been successfully cryopreserved; however, engineered bone models possess greater matrix density, thus demanding prolonged culture and intricate cell–matrix coordination to sustain remodeling activity. Our lab has developed a novel biomaterial, Demineralized Bone Paper (DBP), which preserves native collagen structure and supports cellular interactions for osteoblast-osteoclast bone remodeling. We hypothesized that DBP-based bone organoids can be cryopreserved using a standard DMSO-based slow-freezing protocol while retaining post-thaw viability and functional remodeling capacity. Post-thaw recovery was assessed by monitoring osteoblast viability and mineralization capacity. Mineral deposition was quantified by reductions in transmitted mean light intensity—reflecting progressive mineralization. Cell viability and cytoskeletal architecture were evaluated at the endpoint using DAPI, live/dead, and actin staining. Cryopreservation induced an initial ~30% decrease in osteoblast viability relative to continuously cultured, non-cryopreserved controls; however, it recovered to baseline levels by day 6. Osteoblasts continued proliferating through day 12, increasing by ~20% before stabilizing, mirroring control trends. Immediate post-cryopreservation morphology was elongated, which normalized within 3–4 days. To determine whether remodeling functionality is preserved beyond osteoblast recovery, bone marrow mononuclear cells were introduced to induce osteoclast differentiation and demonstrate the preservation of coordinated bone remodeling dynamics following cryopreservation. These findings demonstrate that the DBP scaffold can be cryopreserved while retaining osteogenic and remodeling functionality, establishing a practical framework for banking complex bone models.

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