Connections in Behavioral Biology The Mechanisms of Drug-Induced Hepatitis Njoku Lab

Drug-Induced Hepatitis There are several types of hepatitis: Viral hepatitis, Alcoholic hepatitis, Autoimmune hepatitis, Iron overload, Fatty liver. Drug-Induced hepatitis or liver injury (DILI) is an idiosyncratic form of acute liver injury and accounts for about 10% of acute liver failure. It’s caused by taking certain medicines, sometimes for too long or simply too much, because the liver is involved in breaking the medicine down. Common medications include anti-seizure medications and some antibiotics Symptoms include nausea, jaundice, fatigue and usually stop after discontinuing the medication. Hepatitis usually affects males more than women, but drug-induced hepatitis disproportionately affects women.

The Role of Cytokines in dili Communication between immune cells by binding to receptors on cell surface. They can act alone, work together, or work against each other: “immunomodulating agents” There are several different types: interferons, interleukins. Different types of cytokines: interferons (proteins that stop viruses from replicating, activate NK), interleukins (first identified in 1970), tumor- necrosis factor Focused on IL-33 because it is associated with tissue disruption. A deficiency in can delay local inflammation and make repairmen of damaged tissue take longer IL-33 is well known for inducing IL-4 but KO IL-33 mice still have IL-4

Hypothesis Past studies have found that IL-10 is lower in males, while IL-6 is upregulated in females due to estrogen. This IL-6 induction causes lower Tregs counts in females and makes hepatitis worse in female mice. IL-33 differentially promotes Tregs in males over female mice.

Methodology -Isolated spleen cells from BALB/c mice, counted cells, and stained cells for a flow cytometry analysis: CFSE staining -Can be used to study different cell types or look for proteins on the cell surface based on light scatter and sensors are set to detect a specific wavelength -Signal intensity, based on the count of cells, is measured at different fluorescence intensities -Direct staining using CD3 and CD28 antibodies that increase cell proliferation

IL-33 KO Female and Male Left shift: increased proliferation Slightly more in females than males

Adding IL-33 The expected shift with CFSE did not occur. It is possible that there was cross- contamination between the plates or low cell viability in the initial cell populations. Will be redoing this experiment in the future. Possibilities: Shift to the left Shift to the right No change

Conclusion Past experiments have allowed me to set-up a system to analyze changes related to IL-33 and differences that may exist between sexes. Next steps: 1) Use more mice shift w/ il-33? 2) Measure changes in regulatory FoxP3+ Tregs 3) View changes in cytokine populations. For instance, we would expect IL-10 to be higher while IL1B should be lower since it has a pro-inflammatory function

Acknowledgements http://www.abcam.com/protocols/introduction-to-flow-cytometry https://emedicine.medscape.com/article/169814-overview https://www.nature.com/articles/nature10134 http://patients.gi.org/topics/medications-and-the-liver/ https://www.uptodate.com/contents/drug-induced-liver-injury https://study.com/academy/lesson/what-are-cytokines-definition-types-function.html