A multifunctional bioreactor for three-dimensional cell (co)-culture.

Understanding how cells behave in three-dimensional environments is a central challenge in biotechnological research. Prior studies have demonstrated the importance of spatial cell organization for mimicking in vivo conditions. However, existing systems often fail to maintain physiological conditions over extended periods. Established knowledge shows that cell interactions and metabolic activity are influenced by culture geometry and matrix composition. The gap in current methods lies in the inability to separate co-cultured cell types while maintaining stable metabolic parameters. This uncertainty drove the need for a bioreactor that could support 3D cell culture with spatial separation. No prior work had resolved the combination of physiological oxygen tension and pH control in such systems. The development of a multifunctional bioreactor addresses this limitation.

The aim of this work is to develop a bioreactor that enables three-dimensional cell culture with spatial separation of co-cultured cell types. The specific problem addressed is the lack of a system that maintains physiological conditions while allowing independent culture environments. The motivation stems from the need to study cell behavior in more realistic settings. The study focuses on cardiomyocyte culture as a model system. The researchers propose that spatial separation can improve cell viability and metabolic stability. The system was designed to allow independent monitoring of oxygen and pH levels. The goal is to provide a platform for long-term cell culture experiments. The study tests whether this bioreactor can maintain stable conditions for two weeks.

The bioreactor was designed with spatially separated compartments for cell culture. Neonatal rat cardiomyocytes were isolated and seeded onto a biodegradable fibrin matrix. The cells were cultured in recirculating medium for 14 days. Metabolic parameters like glucose, lactate, and LDH were measured daily. Oxygen and carbon dioxide levels were monitored to assess aerobic metabolism. Cell viability was tested using LIVE/DEAD staining at multiple time points. Morphological changes were analyzed using hematoxylin and eosin staining. Immunostaining with MF-20 and DAPI was used to confirm cell identity and distribution.

The bioreactor maintained aerobic metabolism with low lactate and LDH release throughout the experiment. Oxygen tension remained high at 183.7 ± 18.4 mmHg. pH levels stayed near physiological values at 7.4 ± 0.02. pCO2 levels were stable at 43.1 ± 2.9 mmHg. Cell viability was sufficient with 82 ± 6.7% living cells after 14 days. No significant differences were observed between bioreactor compartments. The optimal seeding density was 2.0 x 10^7 cells per matrix. The system supported three-dimensional cell culture with spatial separation of co-cultured cells.

The authors propose that the bioreactor provides favorable conditions for three-dimensional cell culture. The system allows spatial separation of co-cultured cell groups. The results suggest that the bioreactor maintains physiological oxygen and pH levels. The system supports aerobic metabolism with minimal lactate and LDH release. The findings indicate that the bioreactor can maintain cell viability for at least 14 days. The study suggests that the system can be used for long-term cell culture experiments. The authors propose that the bioreactor is suitable for cardiomyocyte culture. The results suggest that the system can be adapted for other cell types.