School: The International School
City: Bellevue, WA
Grades taught: 7-8

Principle Investigator: Nephi Stella, Ph.D.
Department: Pharmacology
Institution: University of Washington

Project Description
The current line of research in Dr. Stella’s laboratory is to study the underlying causes and mechanisms linked to neuroinflammation caused by the autoimmune disorder Multiple sclerosis with the ultimate goal to generate specific drugs that may reduce this inflammation. Specifically the lab is studying the different molecular mechanisms underlying microglial cell activation, with a focus on the cannabinoid signaling pathway. They are interested in how cannabinoids, the active agents found in Cannabis sativa, affect the activity of microglial cells. Microglial are the immune cells of the brain. In a healthy brain they are not activated. As a result of an infection or damage to the brain tissue, microglial cells undergo a morphological change and migrate towards the site of infection where they attack invaders and clean up dead and dying cells. The lab has several research projects running that are trying to determine how the activity of microglial cells is modulated by endogenous cannabinoids (produced by our bodies) and exogenous cannabinoids found in the marijuana plant. So far two cannabinoid receptors have been identified (CB1 and CB2) but only CB2 has been found on microglial cells.

It is well known that that LPS/IFNg, LPS a cell surface component of bacteria and IFNg an immune signaling molecule, activates microglial cells.  It has been found that the addition of various cannabinoids along with LPS/IFNg does not affect microglial cell activation. The goal of my project was to find a substance that activates microglial cells and interacts in some capacity with one or more cannabinoids. In order to determine if a substance activates microglial cells I did three different tests. The first test, WST-1, tests for a molecule produced when WST-1 is cleaved by ATP producing enzymes in the mitochondria. The more active the microglial cells, the more WST-1 will be cleaved. The second test detected for the presence of Nitric Oxide (NO), a toxin produced by microglial cells when activated. These two tests were performed by applying the cells with a common inflammatory agents and different cannabinoids. The last test I used to measure microglial cell activation was the migration assay. This tested a cannabinoid’s ability to attract microglia cells when treated with inflammatory agents. This was performed by placing the drug and potential attractant in a bottom well, then placing the cells above separated by filter paper. The cells that migrated to the bottom side of the filter were stained and counted.

I tested three different substances and unfortunately did not discover one that had the ability to activate the cells like LPS/IFN or that changed the pattern of activation in the presence of cannabinoids. Despite these results this summer experience has been very beneficial for me. I have learned firsthand what it is like to be a scientist and how the process of science works. I believe this experience will help me teach the scientific method more accurately and allow me to bridge the gap between science in the classroom and real science.

Acknowledgements
Many thanks to Dr. Stella and Allyn Franklin for their time, knowledge and patience.