Introduction: A major barrier in tissue engineering is the impossibility of providing adequate oxygen to all cells within the engineered tissue before a full vascularization is achieved. To overcome this limitation, a variety of oxygen-producing particles have been developed for improving tissue survival. However, most of these particles are used in random mixtures with scaffolding materials, which usually leads to an uneven distribution of oxygen in bioengineered tissues. An ideal oxygen supply requires a precise spatial-temporal control of the oxygen-producing particles in scaffolds. Unfortunately, current oxygen delivering scaffold techniques are unable to perform as described and to precisely incorporate oxygen particles into the scaffolds. Cell inkjet printing is a novel tissue fabrication approach, in which a special inkjet printer can be programmed to deposit cells and/or biomaterials of various types and sizes in a very precise pattern. In this study we have applied the inkjet printing technology to allocate oxygen releasing materials to their designed positions for optimal cell viability and growth.

 

Methods: The controlled oxygen-releasing platform is fabricated by printing different patterns (“Black”, “White”, “Grey”, and “Dots” to represent different densities of the oxygen particles on the substrates) of encapsulated calcium peroxide (CPO) particles that were analyzed against C2C12 mouse myoblast cell line for cell viability. CPO has been found to release its oxygen over an extended timeframe. The effects of controlled oxygen-releasing particles on cell viability were analyzed using the cell morphology study, live/dead assay, and the MTS assay.

 

Results: These analyses showed the concentration of the oxygen-releasing particles in “Black” was toxic to the cells based on the decreasing trend in cell viability. The “White” did not have oxygen-releasing particles, which correlates to the decrease in cell viability over time due to oxygen deprivation. Both “Grey” and “Dots” showed a similar trend in absorbency, in which the absorbency was low at 24 hours, there was an increase in absorbency at 48 hours, and then an abrupt decrease at 72 hours. Both these results suggest that the amount of oxygen released was beneficial to the cells within the first 48 hours, yet may not have been sufficient to sustain cell viability after that time span. The cells treated in the printed “Dots” showed to have the most compatible treatment for an overall increase in cell viability.

 

 

Conclusion: The amount of oxygen released can be controlled to optimize the cell by bioprinting different densities of the oxygen releasing materials onto a substrate.