Date of Award

4-18-2008

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

Colin K. Combs

Abstract

Inflammation-related neurotoxicity is evident in a number of different models neurodegenerative conditions, including Alzheimer's and Parkinson's diseases, Multiple Sclerosis, and HIV-associated dementia. Peroxisome proliferator-activated receptor gamma (PPARγ) agonists inhibit a number of select pro-inflammatory changes in models of CNS and systemic inflammation. Recent reports suggest that these anti-inflammatory effects are due to mechanisms other than canonical nuclear receptor-mediated transcriptional alteration. Results reported here demonstrate that rosiglitazone, a PPARγ-activating thiazolidinedione, decreases tumor necrosis factor α (TNFα) secretion in both primary microglia and the monocytic cell line, THP-1. Cells were pretreated with various PPARγ ligands, including rosiglitazone, prior to stimulation with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA) to stimulate cytokine production. Rosiglitazone significantly reduced tumor necrosis factor-alpha (TNFα) secretion in both primary microglia and THP-1 cells differentiated for 72 hours in the presence of PMA to induce a macrophage-like phenotype. No reduction in TNFα secretion was observed in undifferentiated THP-1 cells with rosiglitazone pre-treatment. Electrophoretic mobility shift assays (EMSA) revealed no significant difference in PPARγ activation between PMA-differentiated and undifferentiated THP-1 cells. When PMA-differentiated and undifferentiated THP-1 cells were treated with the irreversible PPARγ antagonist, GW 9662, a significant, dose-dependent decrease in TNFα secretion was observed. These results suggest that the anti-inflammatory benefit of PPARγ ligands occurs independently of classical PPARγ activation. PPARα activation elicits potent anti-inflammatory effects in some experimental models. When undifferentiated THP-1 cells were treated with the PPARα agonist WY14643, no decrease in TNFα secretion was observed. This result again suggests that these effects are independent of canonical pathway of PPAR activation and are not due to PPARα activation. When comparing the protein levels of the components of the PPARγ activation pathway, a significant increase in PPARγ and a significant decrease in RXRβ protein were observed in PMA-differentiated THP-1 cells compared to undifferentiated THP-1 cells. These results do not rule out the possibility that the increase in absolute PPARγ protein could be responsible for the increased effect seen with rosiglitazone in PMA-differentiated THP-1 cells. Ultimately, these findings demonstrate that PPARγ ligands are capable of inhibiting pro-inflammatory responses, even when operating outside the context of classical PPARγ activation.

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