Redox status in mammalian cells and stem cells during culture in vitro: Critical roles of Nrf2 and cystine transporter activity in the maintenance of redox balance

Abstract

Culturing cells and tissues in vitro has provided valuable insights into the molecular mechanisms regulating redox signaling in cells with implications for medicine. However, standard culture techniques maintain mammalian cells in vitro under an artificial physicochemical environment such as ambient air and 5% CO2. Oxidative stress is caused by the rapid oxidation of cysteine to cystine in culture media catalyzed by transition metals, leading to diminished intracellular cysteine and glutathione (GSH) pools. Some cells, such as fibroblasts and macrophages, express cystine transport activity, designated as system xc-, which enables cells to maintain these pools to counteract oxidative stress. Additionally, many cells have the ability to activate the redox sensitive transcription factor Nrf2, a master regulator of cellular defenses against oxidative stress, and to upregulate xCT, the subunit of the xc- transport system leading to increases in cellular GSH. In contrast, some cells, including lymphoid cells, embryonic stem cells and iPS cells, express relatively low levels of xCT and cannot maintain cellular cysteine and GSH pools. Thus, fibroblasts have been used as feeder cells for the latter cell types based on their ability to supply cysteine. Other key Nrf2 regulated gene products include heme oxygenase 1, peroxiredoxin 1 and sequestosome1. In macrophages, oxidized LDL activates Nrf2 and upregulates the scavenger receptor CD36 forming a positive feedback loop to facilitate removal of the oxidant from the vascular microenvironment. This review describes cell type specific responses to oxygen derived stress, and the key roles that activation of Nrf2 and membrane transport of cystine and cysteine play in the maintenance and proliferation of mammalian cells in culture.

Characterization of the growth modulatory activities of osteoblast conditioned media on cord blood progenitor cells

“…myeloid cells and CFC myeloid progenitors in OCM cultures (Pineault et al 2011;Metcalf 2008;Broudy 1997). It is also possible that OCM contains growth nutrients that have been partially metabolized by M-OST (Ishii and Mann 2014).…”

Transcription Factor NRF2 as a Therapeutic Target for Chronic Diseases: A Systems Medicine Approach

“…Systems Medicine Approach to NRF2 in Chronic Diseases lipoprotein Ishii and Mann, 2014). Similarly, the gene encoding the proinflammatory cytokine, IL-17D, contains AREs, and this NRF2-T helper (Th)17 axis seems to confer protection against tumorigenesis and viral infections (Saddawi-Konefka et al, 2016).…”

Remote Limb Ischaemic Postconditioning Protects Against Myocardial Ischaemia/Reperfusion Injury in Mice: Activation of JAK/STAT3-Mediated Nrf2-Antioxidant Signalling

“…Under physiological conditions, nuclear factor erythroid 2-related factor 2 (Nrf2) localizes in the cytoplasm and binds to its inhibitor Kelch-like ECH-associated protein 1 (KEEP1) [23]. Upon exposure of cells to oxidative stress or electrophilic compounds, Nrf2 is activated and translocates into the nucleus, where it binds to antioxidant response elements to promote a series of antioxidant genes, including heme oxygenase (HO-1) and superoxide dismutase (SOD) [24]. Therefore, Nrf2 is a key component of cellular redox homeostasis in the attenuation of oxidative stress-associated pathological processes and plays an important role in cardiovascular diseases [25].…”