Presentation Information
[P03-417]Sustained oxygen delivery using calcium peroxide–loaded gelatin microspheres enhances viability and function of hepatocyte spheroids under hypoxic conditions
○Hiroshi Mizumoto1, Tengchang Li2, Masamichi Kamihira1 (1. Department of Chemical Engineering , Faculty of Engineering, Kyushu University (Japan), 2. Graduate School of Systems Life Sciences, Kyushu University (Japan))
Keywords:
tissue engineering,biomaterial,oxygen
[Objective]
Insufficient oxygen delivery to the central regions of highly metabolically active and densely populated tissues, such as the liver, remains a major challenge in in vitro tissue construction, often leading to cellular necrosis. The development of materials capable of supplying oxygen transiently until vascular networks are established is therefore highly desirable in regenerative medicine and tissue engineering. In this study, calcium peroxide (CPO) was investigated as an oxygen source, and oxygen-generating gelatin microspheres (GMS) were fabricated by microencapsulating CPO within a gelatin matrix. The oxygen-release properties of these microspheres and their applicability in hepatocyte spheroid culture under hypoxic conditions were evaluated.
[Methods]
Oxygen-generating GMS were prepared using a water-in-oil (W/O) emulsion method, followed by glutaraldehyde crosslinking and freeze-drying to obtain powdered particles. Oxygen-release profiles were evaluated by monitoring dissolved oxygen concentration over time.
For three-dimensional liver tissue modeling, primary rat hepatocyte spheroids were embedded at high density in collagen gel (collagen gel-embedded culture, CG). Three experimental groups were prepared: CG supplemented with oxygen-generating GMS, CG supplemented with CPO powder, and CG without any oxygen source (control). Cultures were maintained under a 1% oxygen atmosphere, and cell viability, urea synthesis, and albumin secretion were assessed.
[Results]
In oxygen-release assays, GMS significantly suppressed the initial burst release of dissolved oxygen compared with unencapsulated CPO, extending the duration of oxygen release from approximately 70 h to 120 h.
In the three-dimensional culture study, the GMS-supplemented group exhibited the highest cell viability over 5 days, maintaining approximately 55% viability at the final time point. From day 3 onward, this group also demonstrated significantly higher urea synthesis and albumin secretion than the CPO powder group, indicating improved long-term functional maintenance.
[Discussion]
Although initial cell survival was comparable between groups, the GMS-supplemented group exhibited significantly higher metabolic activity than the CPO powder group over time. These findings suggest that the sustained and moderated oxygen release from GMS reduces reactive oxygen species (ROS)-induced oxidative stress and membrane damage associated with rapid oxygen release, thereby preserving cellular functionality.
[Conclusion]
A high-density three-dimensional liver tissue model incorporating oxygen-generating GMS enabled prolonged cell survival and maintenance of liver-specific functions under hypoxic conditions. These findings highlight the potential of integrating internal oxygen supply systems into tissue constructs as a promising strategy to overcome early-stage oxygen deficiency following transplantation.
Insufficient oxygen delivery to the central regions of highly metabolically active and densely populated tissues, such as the liver, remains a major challenge in in vitro tissue construction, often leading to cellular necrosis. The development of materials capable of supplying oxygen transiently until vascular networks are established is therefore highly desirable in regenerative medicine and tissue engineering. In this study, calcium peroxide (CPO) was investigated as an oxygen source, and oxygen-generating gelatin microspheres (GMS) were fabricated by microencapsulating CPO within a gelatin matrix. The oxygen-release properties of these microspheres and their applicability in hepatocyte spheroid culture under hypoxic conditions were evaluated.
[Methods]
Oxygen-generating GMS were prepared using a water-in-oil (W/O) emulsion method, followed by glutaraldehyde crosslinking and freeze-drying to obtain powdered particles. Oxygen-release profiles were evaluated by monitoring dissolved oxygen concentration over time.
For three-dimensional liver tissue modeling, primary rat hepatocyte spheroids were embedded at high density in collagen gel (collagen gel-embedded culture, CG). Three experimental groups were prepared: CG supplemented with oxygen-generating GMS, CG supplemented with CPO powder, and CG without any oxygen source (control). Cultures were maintained under a 1% oxygen atmosphere, and cell viability, urea synthesis, and albumin secretion were assessed.
[Results]
In oxygen-release assays, GMS significantly suppressed the initial burst release of dissolved oxygen compared with unencapsulated CPO, extending the duration of oxygen release from approximately 70 h to 120 h.
In the three-dimensional culture study, the GMS-supplemented group exhibited the highest cell viability over 5 days, maintaining approximately 55% viability at the final time point. From day 3 onward, this group also demonstrated significantly higher urea synthesis and albumin secretion than the CPO powder group, indicating improved long-term functional maintenance.
[Discussion]
Although initial cell survival was comparable between groups, the GMS-supplemented group exhibited significantly higher metabolic activity than the CPO powder group over time. These findings suggest that the sustained and moderated oxygen release from GMS reduces reactive oxygen species (ROS)-induced oxidative stress and membrane damage associated with rapid oxygen release, thereby preserving cellular functionality.
[Conclusion]
A high-density three-dimensional liver tissue model incorporating oxygen-generating GMS enabled prolonged cell survival and maintenance of liver-specific functions under hypoxic conditions. These findings highlight the potential of integrating internal oxygen supply systems into tissue constructs as a promising strategy to overcome early-stage oxygen deficiency following transplantation.
Comment
To browse or post comments, you must log in.Log in