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Effects of metformin on glucose induced staphylococcus aureus growth in airway epithelial co-culture

Effects of metformin on glucose induced Staphylococcus aureus I have always been interested in pursuing scientific research and looking into possible resolutions of people‟s maladies. With thanks to the BPS and from my mentors, Dr. Garnett and Dr. Baines I was given an opportunity to work for 6 weeks in the Basic Medical Labs of St. Georges‟, University of London. During these weeks I conducted an experiment into looking at how airway epithelial cells are affected by the biguanide drug, metformin. It is known that airway surface Liquid (ASL), found on the luminal side of epithelial airway cells has an exposure to bacteria but is usually kept sterile due to the antimicrobial agents. Maintaining a low glucose concentration in the ASL is vital in preventing bacterial growth, which a major cause of airway infection in diseases associated with airway inflammation and hyperglycemia. It had been suggested by Baines et al (2008) that „GLUT transporters and its subsequent metabolism in lung epithelial cells help to maintain low glucose concentrations in human ASL which is important for protecting the lung against infection.‟1 Metformin, known for using mechanisms to suppress gluconeogenesis via deploying GLUT transporters2, could potentially have an effect on the glucose concentration on the ASL by increasing GLUT-mediated glucose uptake into the airway epithelium and could thus prove important in managing conditions such as cystic fibrosis and hyperglycaemia. Staphyloccus aureus is a gram positive coccal bacterium, prevalent in approximately 20% of the human population. Using glucose as an substate for growth, it is known for its association with recurrent infections. With this in mind it can be seen as important in mediating the glucose concentration of the ASL to maintain its sterility and decrease the cases of infection. Using all this information I devised the hypothesis that „metformin could have a detrimental effect on the growth of Staphylococcus aureus in airway epithelial co-cultures.‟ To test this hypothesis I first investigated the effect of varying metformin concentrations (1mM, 100µM, 10µM, 1µM) on Staphylococcus growth. My results showed that there was no effect and as such I could proceed with the main experiment with knowledge that metformin does not directly affect the growth of the bacterium. This taught me the need to think ahead about all possible out comes of testing a drug and the use of appropriate controls. The main experiment composed of introducing Staphylococcus aureus at a concentration of 1x107 CFU/ml onto the apical surface of H441 airway epithelial cell monolayers grown on transwell supports. The H441 cells were pre-treated with metformin or the appropriate control. The basolateral glucose concentration of the Krebs ringer solution was varied (10mM, 20mM, 40mM and also 10mM bilateral). A 7 hour incubation time followed in a 5% CO2 incubator equilibrating the cells while at 37◦C. Lastly, a miles and misra count was done by plating dilutions of the bacteria-epithelia co-culture mix, preceding an overnight incubation. This gave me an estimate of Staph. aureus growth for each condition. I enjoyed the independence I was given to conduct the whole experiment from the raw materials and being able to work on the time constraints to get the most out of my experiment. After, collecting the results by miles and misra, I analysed my results to form figures which could be used for future publication. Analysis of the data showed no significant effect of metformin, as the results were very variable. Therefore further investigation may be necessary to test my hypothesis. If I was to continue this project further I would firstly, undergo more trials. In addition, I would use more concentrations of glucose and may also look at using other types of airway epithelial cells. Personally, I found my six week studentship as both interesting and informative. Working with Dr. Garnett and Dr. Baines has enlightened me of the work involved in research and I can say that I hope to use this as a platform to a future career. I like the idea of directing your own research and investigating whether the hypotheses are valid. After seeing how hard my supervisors worked I feel I could work in this environment where the work may be hard but in my eyes rewarding. Lastly, I would like to thank the BPS and my supervisors, Dr. Garnett and Dr. Baines, for the opportunity to gain this experience. Reference
1 Kalsi, K.K., Baker, E.H., Chung, Y.L., Mace, O.J., Philips, B.J. and Baines, D.L. (2008). Glucose homeostasis across
human airway epithelial cell monolayers: role of diffusion, transport and metabolism. Pflugers Arch. 457(5): 1061-1070.
2 Grisouard J, Timper K , Radimerski TM , Frey DM , Peterli R, Kola B , Korbonits M , Herrmann P , Krähenbühl
S,H Zulewski ,U Keller ,B Müller ,M Christ-Crain (2010) “Mechanisms of metformin action on glucose transport and
metabolism in human adipocytesPages 1736-1745
By Akin Falase



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