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Breathe Easy: We Fixed the Oxygen, The Rest Got Easier How Oxygen-Carrying Nanoparticles Alter Viability in 3D Culture

Year: 2025
Type: other
Abstract: Introduction Hypoxia remains a critical barrier to accurately modeling tumor biology and testing drug efficacy in three-dimensional (3D) cell culture [1]. Traditional approaches, including hemoglobin-based strategies, often struggle to deliver oxygen uniformly across spheroids larger than 250 µm [2]. To address this challenge, we developed poly(lactic-co-glycolic acid) (PLGA) nanoparticles loaded with perfluorocarbon (PFCE)—a stable, biocompatible oxygen carrier [3]. Here, we investigate the effects of PFCE-loaded nanoparticles on oxygenation in T47D breast cancer spheroids reaching up to 1000 µm in diameter, aiming to facilitate more robust long-term cultures and advance organ-on-a-chip methodologies [4]. Results and Discussion T47D spheroids were cultivated under controlled oxygen conditions and treated at varying concentrations of PFCE-loaded nanoparticles. Over 31 days, PFCE nanoparticles did not significantly affect spheroid growth rates (±\pm±5–10% difference from untreated controls), suggesting that the nanoparticles do not interfere with the overall expansion of the spheroid structure. However, viability assays revealed a notable benefit at lower PFCE doses, with up to ~15% higher ATP-based metabolic activity than control spheroids during the first two weeks of culture. Apoptosis assays further showed reduced cell death, particularly in peripheral and mid-layer regions of the spheroid. Beyond day 21, preliminary data indicate a subtle decline in viability, although ongoing experiments aim to clarify the long-term effects. Flow cytometric analysis of dissociated spheroids and targeted staining demonstrated a decrease in hypoxic regions when PFCE nanoparticles were present, indicating more efficient oxygen transport to the spheroid core. Three-photon imaging and high-field (14T) MRI provided additional insights into spheroid architecture, revealing minimal overlap between nanoparticles and residual hypoxic zones. While this imaging-based evidence is still being investigated, preliminary observations support our hypothesis that PFCE-loaded nanoparticles can facilitate improved oxygenation throughout larger 3D tumor models. Conclusion These results suggest that PLGA-PFCE nanoparticles can enhance oxygen delivery and bolster viability in large T47D spheroids without significantly altering growth rates. Improved oxygenation is pivotal for extending culture durations, refining tumor modeling, and advancing the design of organ-on-a-chip systems. Similar approaches may be adapted for tissue engineering or even the development of bioengineered donor organs, underscoring the far-reaching impact of nanoparticle-mediated oxygenation strategies. References 1. Wu, Q.; Liu, J.; Wang, X.; Feng, L.; Wu, J.; Zhu, X. 3D Tumor Spheroid as an In Vitro Tumor Model for Cancer Immunotherapy Evaluation. Tissue Eng. Part B Rev. 2022, 28(2),329–343. 2. Riedl, A.; Schleicher, M.; Huemer, F.; et al. Modeling the Tumor Microenvironment in 3D Culture: Challenges, Advances, and Opportunities for Nanoparticle-Mediated Drug Delivery. Acta Biomater. 2023, 156, 1–18. 3. Li, Y.; Lu, J.; Han, C.; Wang, C.; Wang, J.; Yang, H. Perfluorocarbon-Based Artificial Oxygen Carriers for Advanced Tumor Therapy. Adv. Healthcare Mater. 2021, 10(21), e2100820. 4. Zhu, L.; Ma, X.; Li, K.; et al. Engineering Perfluorocarbon-Loaded PLGA Nanocarriers for Enhanced Oxygen Delivery and Multimodal Imaging in 3D Tumor Models. ACS Appl. Bio Mater. 2023, 6(8), 2234–2247.
DOI: 10.5281/zenodo.17060561
Language: English
Subfield: Biomaterials
Open Access status: green
OpenAlex ID: W7080126254
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