Semester

Fall

Date of Graduation

2010

Document Type

Thesis

Degree Type

MS

College

Statler College of Engineering and Mineral Resources

Department

Chemical and Biomedical Engineering

Committee Chair

David Klinke.

Abstract

In order to design better therapeutic strategies, scientists work on understanding the interactions that take place at cell level and have an effect on the immune system. In order to identify a few possible inhibitors that may be up-regulated in tumor environment and that may affect T-cell action, a cellular assay was designed. An Interleukin-12 (IL-12)-responsive murine Th1 cell clone called 2D6 was used as cellular model to study the effects of transforming growth factor-beta (TGF-beta), insulin, and interleukin-6 (IL-6) on the phosphorylation of Signal transducer and activator of transcription 4 (STAT4), a key element in IL-12 signaling. Cells were IL-12-starved for 12 hours and then were treated for 15 minutes, 1 hour and 2 hours with one of these factors and/or IL-12 and the cell samples were analyzed by flow cytometry. In general, TGF-beta and insulin had a significant effect in the phosphorylation of STAT4 and cell viability particularly after 2 hours in cells incubated with these factors only. While more subtle, the effects of IL-6 also seemed to be have a stronger effect after 2 hours of treatment. The assay designed was able to provide with answers about inhibition of IL-12 signaling in a relatively restricted time frame, but more questions need to be answered to fully understand the effect of each factor, for which a new experimental approach may be needed.

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