Media composition and O₂ levels determine effects of 17β-estradiol and selective estrogen receptor modulators on mitochondrial bioenergetics and cellular reactive oxygen species

Abstract

Estradiol (E2) and selective estrogen receptor modulators (SERMs) have broad-ranging cellular effects that include mitochondrial respiration and reactive oxygen species (ROS) metabolism. Many of these effects have been studied using cell culture models. Recent advances have revealed the extent to which cellular metabolism is affected by the culture environment. Cell culture media with metabolite composition similar to blood plasma (e.g. Plasmax, HPLM) alter cellular behaviours including responses to drugs. Similar effects have been observed with respect to O2 levels in cell culture. Given these observations, we set out to determine whether the effects of E2 and SERMs are also influenced by media composition and O2 level during cell culture experiments. We analyzed mitochondrial network characteristics, cellular oxygen consumption rates, and cellular H2O2 production in C2C12 myoblasts growing in physiologic (5%) or standard cell culture (18%) O2 and in physiologic (Plasmax) or standard cell culture (DMEM) media. The cell culture conditions affected all measured parameters under basal conditions and changed how cells responded to E2 or SERMs. These results indicate that the effects of E2 and SERMs on various aspects of cell physiology strongly depends on growth conditions, which in turn emphasizes the need to consider this carefully in cell culture experiments.

Physiological Media in Studies of Cell Metabolism

“…It is worth noting that different standard culture media were used for each of these lines, i.e., DMEM, DMEM-F12, and MEM. A similar increase in the respiratory activity of cells has been described by other researchers for other cell lines, i.e., breast cancer MCF7, adenocarcinoma prostate LNCaP, and osteosarcoma SaOS2 [ 42 , 43 ]. Moreover, in the latter work, an increase in respiratory activity is described in hypoxia conditions.…”

Role of Food Antioxidants in Modulating Gut Microbial Communities: Novel Understandings in Intestinal Oxidative Stress Damage and Their Impact on Host Health

“…This term needs to be clarified due to the connection between reactive oxygen species and free radicals is not entirely comprehensive and similar, but on the other hand ROS are more communal in the tendency of numerous disorders [48]. The ROS are generated through the continuous cell respiration process; therefore, the mitochondria might be considered as a primary source for ROS production in most of the organisms [49]. The ROS can be divided into free ROS (alkoxyl radical, peroxyl radical, hydroxyl radical, and superoxide) and non-free ROS (organic hydroperoxides, oxygen, chloramines, hydrogen peroxide, and hypochlorous acid) depending on the electron's status.…”

Oxygen toxicity: cellular mechanisms in normobaric hyperoxia

“…Thus, cells cultured under standard conditions are in fact experiencing hyperoxia. Many cellular processes measured in “physioxia” (2–6% O 2 ) are different compared to 18–19% O 2 , such as ROS production (Maddalena et al 2017 ), proliferation and senescence (Packer and Fuehr 1977 ; Busuttil et al 2003 ; Parrinello et al 2003 ), mitochondrial function (Moradi et al 2021b ), and response to drugs (Yan et al 2010 ; Fonseca et al 2018 ; Otto-Ślusarczyk et al 2021 ) and hormones (Moradi et al 2021a ). It is thus imperative to implement physiological O 2 conditions in cell culture, in order to improve the quality and validity of in vitro studies and avoid artificial outcomes.…”