Unfortunately, these ideals are challenging to estimation and have to be established experimentally. potential outcomes, and alternative tradition strategies open to help circumvent this largely unrecognized issue currently. molecular air) through aqueous moderate and the need for effective air delivery systems to keep up cell viability. Following the cell tradition revolution in the 50s and 1940s, the data acquired by Krogh from cells explants was put on cells expanded in monolayer. In pivotal function by Dr. William McLimans in 1968, it had been noticed that the air consumption price (OCR) of Pinoresinol diglucoside cells in the bottom of the petri dish can simply surpass the diffusion price of air through the overlying culture medium. McLimans subsequently warned against growing cells at too high of density or at excessive medium depths due to significant risk of oxygen deficit or even anoxia at the cellular level [6,7]. A breadth of significant evidence now exists demonstrating that over-seeding cells can result in altered cell growth characteristics, aberrant signaling, and serious deficiencies in experimental validity [8C10]. The scientific community generally accepts that cell culture is not a perfect model system. However, there remains a significant amount of complacency with respect to the limitations of oxygen diffusion and the potential effects on cells. This review attempts to answer three fundamental questions: 1) How do the properties of oxygen diffusion and delivery differ between and environments? 2) What Pinoresinol diglucoside are the consequences of this altered oxygen availability on the experimental and translational validity of models? 3) What can scientists do to minimize fluctuations in oxygen concentration in their cell culture models? The review will conclude with basic recommendations to improve rigor and reproducibility of cell culture experiments, especially those focused on hypoxia and metabolism. ITGAV II. Theory of Oxygen Diffusion In the human body, tissue oxygenation is a tightly regulated process. Oxygenation of the blood is first controlled by respiratory rate and intrinsic mechanisms within the pulmonary circulation that maintain oxygen partial pressure in the arterial blood (PaO2). Physiologic PaO2 is maintained around 100 mmHg, which equates to 0.13 mmol of unbound oxygen per liter of blood at sea Pinoresinol diglucoside level (see subsequent sections for math). Hemoglobin increases oxygen capacity of blood an additional 60 times this amount, but it does not contribute to the partial pressure in its bound state [11]. At the level of the tissue, the local partial pressure of oxygen (PO2) is decreased to approximately 40 mmHg due to cellular oxygen consumption. This drop in oxygen partial pressure creates an oxygen gradient that pulls dissolved oxygen a short distance from the capillary to the respiring cells. The oxygen is quickly replaced by the vast hemoglobin stores that are sensitive to pH and other metabolic factors that fine tune the release of oxygen [11]. Hemoglobin therefore acts as a rheostat and buffer to maintain a constant rate of flow (otherwise known as flux) of Pinoresinol diglucoside oxygen to metabolically active tissues. In cell culture, these intricate regulatory mechanisms discussed above are stripped away. What is left are the raw physical laws that govern the properties of gasses and molecules in solution. To understand the extent to which oxygen delivery to cells is limited in the cell culture model, one must understand the nature of molecular oxygen at three levels: 1) 2) is no longer sufficient to maintain ATP production via oxidative phosphorylation [12]. However, the term hypoxia has been clouded by the discovery of the hypoxia-inducible factor (HIF), which is regulated by oxygen sensitive prolyl-hydroxylases (PHDs)[13]. This system (along with other oxygen-sensitive enzyme pathways) is adaptable to particular set-points that are tissue-dependent, and functions to sense changes in oxygen tension from the.