Date of Award

12-15-2016

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

Jian-Dong Li

Second Advisor

Zhi-Ren Liu

Third Advisor

Sang-Moo Kang

Abstract

Otitis media (OM) is the most common childhood bacterial infection, and leading cause of conductive hearing loss. Nontypeable Haemophilus influenzae (NTHi) is a major bacterial pathogen causing OM. During infection, epithelial cells act as the first line of defense by secreting numerous pro-inflammatory mediators including C-X-C motif chemokine ligand 5 (CXCL5) and mucin 5AC (MUC5AC). While appropriate inflammatory responses are critical for the containment and removal of the invading pathogen, excess inflammation can lead to tissue damage, impaired mucociliary clearance and be detrimental to the host. Therefore, inflammatory responses must be tightly regulated. Current therapies for OM are ineffective due to the emergence of antibiotic-resistant NTHi strains and risk of side effects with prolonged use of immunosuppressant drugs. Therefore, therapeutic strategies that increase the levels of endogenous negative regulators of inflammation while leaving the positive pathways intact are gaining prominence. Thus, understanding the underlying molecular mechanisms regulating inflammation is critical for developing effective therapeutic strategies.

Despite the importance of CXCL5 chemokine in mediating inflammation, the signaling cascade mediating its up-regulation in OM remains largely unknown. Here we show that NTHi up-regulates CXCL5 expression by activating IKKβ-IκBα and p38 MAPK pathways via NF-κB nuclear translocation-dependent and -independent mechanism in middle ear epithelial cells. We also show that MKP-1 is a negative regulator of NTHi-induced CXCL5 expression. We further demonstrated the translational significance of these findings by reporting for the first time that curcumin, derived from Curcuma longa plant suppressed CXCL5 expression by direct inhibition of IKKβ phosphorylation, and inhibition of p38 MAPK via induction of negative regulator, MKP-1. Next, we show that curcumin also suppressed NTHi-induced MUC5AC expression via up-regulation of MKP-1, demonstrating for the first time the efficacy and pleiotropic anti-inflammatory action of curcumin to suppress NTHi-induced inflammatory responses in OM model. Finally, we demonstrate for the first time that MKP-1 protein undergoes lysine 63 (K63)-linked polyubiquitination, suggesting the importance of post-translational modification on MKP-1 activity. We also show that curcumin enhanced K63-linked polyubiquitination of MKP-1. Since K63-linked polyubiquitination mediates non-degradative molecular functions such as protein-protein interactions, protein trafficking and regulation of signal transduction events, our findings suggest a new mechanism of action of curcumin on MKP-1. Taken together our study demonstrates the therapeutic potential of curcumin in treating NTHi-induced OM by modulating the expression and activity of negative regulator MKP-1.

Available for download on Wednesday, November 21, 2018

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