Abstract
Oxygen plays a key role in stem cell biology as a signaling molecule and as an indicator of cell energy metabolism. Quantification of cellular oxygen kinetics, i.e. the determination of specific oxygen uptake rates (sOURs), is routinely used to understand metabolic shifts. However current methods to determine sOUR in adherent cell cultures rely on cell sampling, which impacts on cellular phenotype. We present real‐time monitoring of cell growth from phase contrast microscopy images, and of respiration using optical sensors for dissolved oxygen. Time‐course data for bulk and peri‐cellular oxygen concentrations obtained for Chinese hamster ovary (CHO) and mouse embryonic stem cell (mESCs) cultures successfully demonstrated this non‐invasive and label‐free approach. Additionally, we confirmed non‐invasive detection of cellular responses to rapidly changing culture conditions by exposing the cells to mitochondrial inhibiting and uncoupling agents. For the CHO and mESCs, sOUR values between 8 and 60 amol cell−1 s−1, and 5 and 35 amol cell−1 s−1 were obtained, respectively. These values compare favorably with literature data. The capability to monitor oxygen tensions, cell growth, and sOUR, of adherent stem cell cultures, non‐invasively and in real time, will be of significant benefit for future studies in stem cell biology and stem cell‐based therapies.
Highlights
The survival, proliferation and phenotype of stem cells is determined by a dynamic and complex interaction between the cellular microenvironment and the cells [1]
The bespoke pressure-driven pumpwww.biotechnology-journal.com www.biotecvisions.com ing system consisted of a gas supply (21% O2, 5%CO2, N2; BOC, UK) controlled by a pressure regulator (ITV001-2BLQ, SMC Pneumatics Ltd, UK) and connected to a medium reservoir (DURAN® bottle with GL-45 cap, Schott AG, Germany)
The microfluidic cell culture device was placed on a motorized stage of an inverted fluorescence microscope for non-invasive monitoring
Summary
The survival, proliferation and phenotype of stem cells is determined by a dynamic and complex interaction between the cellular microenvironment and the cells [1]. Physico-chemical factors, such as pH, dissolved oxygen, hydrodynamic shear stress and temperature, and biochemical factors, for example the concentrations of nutrients, metabolites and signaling factors, contribute to specific cellular fates. Among these factors, dissolved oxygen (DO) is of particular importance. Analysis of the oxygen uptake of stem cells revealed shifts in their energy metabolism These shifts were correlated with the phenotypic changes observed in the study of inherited diseases [11], wound healing [12], and differentiation pathways [13,14,15,16,17]. The capacity to monitor the DO in a culture and to quantify the oxygen kinetics of the cells, such as the specific oxygen uptake rates (sOUR), is critical to the development of stem cell-based therapies
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