Oxidative Stress Under Ambient and Physiological Oxygen Tension in Tissue Culture

Abstract

Oxygen (O2) levels range from 2–9% in vivo. However, cell culture experiments are performed at atmospheric O2 levels (21%). Oxidative stress due to generation of reactive oxygen species (ROS) in cells cultured at higher than physiological levels is implicated in multitude of deleterious effects including DNA damage, genomic instability and senescence. In addition, oxidative stress activates redox sensitive transcription factors related to inflammatory signaling and apoptotic signaling. Furthermore, several chromatin-modifying enzymes are affected by ROS, potentially impacting epigenetic regulation of gene expression. While primary cells are cultured at lower O2 levels due to their inability to grow at higher O2, the immortalized cells, which display no such apparent growth difficulties, are typically cultured at 21% O2. This review will provide an overview of issues associated with increased oxygen levels in in vitro cell culture and point out the benefits of using lower levels of oxygen tension even for immortalized cells.

Extended hypoxia‐mediated H2S production provides for long‐term oxygen sensing

“…The physioxic Po 2 for cells used in this study is quite variable (Po 2 in parenthesis); HEK293 (50-70), BPASMC (42-100), stem (100 in peritoneum, 15 or lower for most cells), A549 (100-200), PTE (140), HTC116 (85 in crypt to <10 at villus tip). [51][52][53][54][55][56][57][58] In the present experiments, we clearly show that H 2 S production in a variety of cells varies inversely with Po 2 over a range of O 2 tensions from hyperoxic to physioxic to hypoxic. We also show that H 2 S production in HCT116 cells is less sensitive to Po 2 , which presumably reflects their relatively hypoxic environment in vivo.…”

“…Even normoxia (or more appropriately physioxia) will vary depending on the environment inhabited by cells in situ. The physioxic P o 2 for cells used in this study is quite variable (Po 2 in parenthesis); HEK293 (50‐70), BPASMC (42‐100), stem (100 in peritoneum, 15 or lower for most cells), A549 (100‐200), PTE (140), HTC116 (85 in crypt to <10 at villus tip) . In the present experiments, we clearly show that H 2 S production in a variety of cells varies inversely with Po 2 over a range of O 2 tensions from hyperoxic to physioxic to hypoxic.…”

Dynamic effects of genetic variation on gene expression revealed following hypoxic stress in cardiomyocytes

“…DNA methylation levels are also altered in response to other stressors such as bacterial infection ; however, the importance of timing is highlighted by the fact that, in this system, gene expression responses precede DNA methylation changes (Pacis et al, 2019). It is also important to note that our study considers baseline oxygen levels to be 10% oxygen, which is closer to physiological oxygen levels (5-13%) than atmospheric oxygen levels (21%; (Brahimi-Horn and Pouyssegur, 2007;Carreau et al, 2011;Jagannathan et al, 2016). Most studies define normoxia as 21% oxygen saturation, and while this likely leads to larger effect size differences in known hypoxia response genes following hypoxia, these comparisons may not give meaningful insight into the in vivo state.…”

Bioengineering a novel 3D in-vitro model to recreate physiological oxygen levels and tumor-immune interactions

“…Physiological oxygen levels in solid tumors are very heterogeneous ranging from 0.5 -4% oxygen saturation compared to 4 -14% in healthy tissues (59,(61)(62)(63). In addition, circulating and immune cells are exposed to a full dynamic range of blood physiological O2, in which the pO2 varies from 13.5% in alveoli and 9.5% in arterial blood to 6.5% at the venous end of circulation (64)(65)(66)(67)(68). Therefore, studies employing models incubated at atmospheric oxygen (21% O2) culture conditions are hyperoxic with respect to the tissue of origin (65,69).…”