The review article “Frequently asked questions in hypoxia research” by Wenger et al. covers several issues every researcher performing In vivo or In vitro hypoxia-based research should think about before performing their studies.

This article is meant to help those in the early stages of attempting hypoxic studies. It should also be useful to those who are well versed in the subject as it prompts reflection on important questions pertaining to their current research. For those not already doing so, the article also provides a variety of reasons why one may want to consider controlling oxygen along side CO2, temperature and humidity within their incubator and/or workstation in future studies.

Key Points

  1. Partial pressure of oxygen as a measurement vs. percentage of oxygen
    • The authors discuss some of the issues with using percentage, for example, that it does not account for altitude or differences of other pressures (like humidity) and thus is a less than perfect metric for measuring the amount of oxygen. On the other hand, using mmHg or any other pressure measurement for oxygen will account for all of these variations and, thus, provides a far more accurate measure.
  1. Oxygen Concentrations and hypoxia in cell culture dishes
    • The take away point is cells near the surface “feel” oxygen, or the lack thereof, differently than cells deeper into the media. One must be mindful that diffusion of oxygen into or out of media is a process that takes time. It is best, if you have the means, to test pO2 of cells at different depths within the media.
    • Hypoxia needs to be well controlled as any major fluctuations will re-oxygenate cells quickly. Hypoxia chambers need to have ways to limit oxygen entry into a system (for example – a negative pressure entry flap system) and correct for any fluctuations quickly.
  1. The partial pressure of tissues and organs
    • This of course is highly variable from tissue to tissue and organ to organ. What is important to note is some organs have pO2 gradients (like liver and kidney), some are cyclic (like heart), and some are constant (lungs; high, intervertebral discs; low). It is important for researchers to know what the correct pO2 is for their organ/tissue/cell of interest and try and ensure the environment they are in is as close to “normal” as possible.
    • The survival of tissues and organs relies on an adequate oxygen supply. The measurement of tissue oxygen tension provides a direct measure of the balance between oxygen supply (by the blood) and metabolic oxygen consumption (by the tissue). These levels can be measured by optical fluorescence technology intended for the quantitative measurement of oxygen partial pressure.
  1. Normoxia, Hyperoxia and Hypoxia
    • Normoxia, when referring to a specific tissue should be defined as its normal oxygen range, and hypoxia should refer to any amount below the physiological range.
    • Although incubators not controlling for O2 are usually referred to as normoxic, they should actually be referred to as hyperoxic because no tissue in the body is ever exposed to atmospheric oxygen levels.

For more details on any of the points above you can read “Frequently asked questions in hypoxia research” here!

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Keywords: Hypoxia incubator, cell culture, partial pressure, oxygen sensor, tissue oxygen, cell environment, in vitro