Date on Master's Thesis/Doctoral Dissertation

8-2014

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Microbiology and Immunology

Committee Chair

Suttles, Jill

Committee Member

Shirwan, Haval

Committee Member

Egilmez, Nejat

Committee Member

Chesney, Jason

Subject

Cytokines--Therapeutic use; Cancer--Treatment

Abstract

Tumor infiltrating and tumor associated myeloid cells (TIMs and TAMs) elaborate an array of factors that promote tumor growth and metastasis. IL-12, a potent inflammatory cytokine has been shown to induce regression of many cancers. We hypothesize that IL-12 augments the ability of TIMs and TAMs to respond to inflammatory stimuli providing a window in which these stimuli are more likely to promote tumor destruction. Related to this hypothesis, we asked two questions: (1) Does IL-12 directly change signaling events associated with inflammatory signal transduction? (2) Is IFN? required for the entirety of IL-12 induced enhancement of the response of TIMs to inflammatory stimuli? First, we looked broadly at the in vivo effects of microspheres containing IL-12 on the growth and metastasis of 4T1 tumors. We also did studies with tumor cell-dendritic cell fusion for future examination of the impact of IL-12 and myeloid cells on the efficacy of tumor cell-dendritic cell fusion vaccines in mice. Most of this dissertation focuses on in vitro work using TIMs and TAMs isolated from wild-type BALB/c or IFN? deficient mice bearing the 4T1 mammary carcinoma. TIMs and TAMs were pretreated in vitro with IL-12 followed by LPS. TNFa, IL-6, and IL-10 cytokine and mRNA levels were measured. We also examined the impact of IL-12 on the response 4T1 TIMs to tumor derived products. The phosphorylation of a number molecules involved in inflammatory signaling pathways, including MAPK proteins, were assessed by Western Blot. We found that treatment of TIMs with IL-12 followed by exposure to LPS enhances the amount of IL-6 and TNFa with a reciprocal decrease in IL-10. This observation is associated with increases in the phosphorylation of MAPKs. The presence of IFN? is only partially necessary for these effects. We observed that IL-12 only significantly impacted the production of IL-10 from 4T1 TAMs in response to LPS. IL-12 caused a significant increase in the amount of TNFa and IL-6 in response to tumor derived products without affecting IL-10. Our results provide additional insight into direct changes induced by IL-12 to the functional phenotype of TIMs and TAMs in response to inflammatory stimuli.

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