Abstract
Oxygen is not only crucial for cell survival but also a determinant for cell fate and function. However, the supply of oxygen and other nutrients as well as the removal of toxic waste products often limit cell viability in 3-dimensional (3D) engineered tissues. The aim of this study was to determine the oxygen consumption characteristics of 3D constructs as a function of their cell density. The oxygen concentration was measured at the base of hepatocyte laden constructs and a tightly controlled experimental and analytical framework was used to reduce the system geometry to a single coordinate and enable the precise identification of initial and boundary conditions. Then dynamic process modeling was used to fit the measured oxygen vs. time profiles to a reaction and diffusion model. We show that oxygen consumption rates are well-described by Michaelis-Menten kinetics. However, the reaction parameters are not literature constants but depend on the cell density. Moreover, the average cellular oxygen consumption rate (or OCR) also varies with density. We discuss why the OCR of cells is often misinterpreted and erroneously reported, particularly in the case of 3D tissues and scaffolds.
Highlights
Engineered tissues have many applications, ranging from in-vitro models of human pathophysiology to platforms for drugs and treatment testing (Lee et al, 2005), to providing an alternative to the shortage of human donor organs for transplants (Mattei et al, 2017b, 2018)
The expected increase in Vmax with increasing cell density was concomitant with a decrease in sOCR and an increase in Km, both of which suggest the presence of cooperative behavior in the cell oxygen consumption characteristics, which are dependent on cell density
The capacity to monitor oxygen in a culture and to quantify the cell oxygen consumption kinetics is critical for the development of tissue engineered products
Summary
Engineered tissues have many applications, ranging from in-vitro models of human pathophysiology to platforms for drugs and treatment testing (Lee et al, 2005), to providing an alternative to the shortage of human donor organs for transplants (Mattei et al, 2017b, 2018). The supply of oxygen and other nutrients as well as the removal of toxic waste products, which typically occur via passive diffusion in in-vitro cellular constructs, often limit cell viability in thick (millimeter-sized) engineered tissues (Mattei et al, 2014). Oxygen is considered the limiting factor when culturing three-dimensional (3D) cell constructs in-vitro, especially in the case of non-porous systems where its mass transport relies only on gradient-driven passive diffusion. The reason for this is its poor solubility in culture media (typically ∼0.2 mM, when atmospheric oxygen is used), making it difficult to provide a sufficient
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