Quantitative Analysis of the Effect Antioxidants have on Macrophages' Destruction of Bacteria?

[deleted-305477]

I was thinking of doing a project along the lines of the "Effect Antioxidants have on Macrophages." I was wondering how I would quantitatively analyze how antioxidants effect the effieciency in which macrophages destroy bacteria. For example what would I grow the macrophages in? What would I add for the atioxidant variable? How would quantitatively analyze how macrophages destruction of bacteria is effected by antioxidants?

This would mean a lot if someone could help me

[SciB]

Hi Dave,

You are proposing a very ambitious project and one that would give some extremely interesting and timely data since antioxidant supplements are something many people take hoping they will prevent disease, but the results are still controversial.

I am assuming that you have access to a research lab where you can do tissue culture and perform these experiments under supervision. Culturing and working with mammalian cells like macrophages requires a BSL2 hood, inverted microscope, centrifuge and other equipment, culture media and expensive supplements. If you have all these resources and someone to teach you how to use the equipment, and at least a couple of months of pretty intensive labor to devote to this project you should be able to get some impressive results.

Macrophages are one of the innate immune systems first lines of defense against microbial invaders, but they also play a role in stimulating inflammation and the adaptive immune response that produces antibodies against foreign bodies like bacteria: https://en.wikipedia.org/wiki/Macrophage

One important thing to keep in mind about the human immune system and the macrophages in particular is that there has to be tight controls on their activity. Many diseases from arthritis to cancer are made worse by too much inflammation brought on by the activity of immune system cells such as macrophages. While their activity is mostly beneficial, they can overdo it and cause too much damage to the tissue. On the other hand, if their functions are weakened then microbes and cancer cells may be able to multiply and cause disease.

The usefulness of antioxidant supplementation in the diet is a very controversial subject: https://en.wikipedia.org/wiki/Antioxidant

In fact, several studies have linked antioxidant use to worsening of disease:

Bjelakovic G, Nikolova D, Gluud LL, Simonetti RG, Gluud C (14 March 2012). "Antioxidant supplements for prevention of mortality in healthy participants and patients with various diseases". The Cochrane Database of Systematic Reviews 3: CD007176. doi:10.1002/14651858.CD007176.pub2. PMID 22419320

Miller ER, Pastor-Barriuso R, Dalal D, Riemersma RA, Appel LJ, Guallar E (Jan 2005). "Meta-analysis: high-dosage vitamin E supplementation may increase all-cause mortality". Annals of Internal Medicine 142 (1): 37–46.

Seifried HE, McDonald SS, Anderson DE, Greenwald P, Milner JA (Aug 2003). "The antioxidant conundrum in cancer". Cancer Research 63 (15): 4295–8. PMID 12907593.

Which macrophages did you plan to use in your experiments? While you could isolate primary monocytes and culture them to differentiate into macrophages, I would recommend using a macrophage cell line such as the human U937 or the mouse RAW264.7. These are both available from the American Type Culture Collection (ATCC):
http://www.atcc.org/products/all/CRL-1593.2.aspx
http://www.atcc.org/products/all/TIB-71.aspx

Macrophages have many activities that can be measured quantitatively in the lab. What you choose is going to depend on what instruments and reagents are available to you. Macrophages will ingest bacteria and destroy them so that would be one effect that you could measure. They also produce many substances called cytokines that influence other cells of the immune system and these can be identified by a technique called immunofluorescence, but this requires that your lab have a fluorescence microscope.

I hope this information will help you decide how best to embark on this advanced project. Get back to us with a statement of what you think your hypothesis should be and more details on how you were thinking about doing the experiments to test it. We can guide you better once we know more about what time and resources you have available.

Good luck!

Sybee