All ETDs from UAB

Advisory Committee Chair

Charles O Elson

Advisory Committee Members

Robin G Lorenz

John F Kearney

Casey T Weaver

Lesley E Smythies

Document Type

Dissertation

Date of Award

2015

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

A dysregulated immune response to the microbiota is a defined characteristic of Crohn's Disease (CD), a type of Inflammatory Bowel Disease (IBD). Understanding how homeo-stasis is maintained in the intestine despite the immense concentration of bacteria present is an active area of research. Regulatory T cells (Tregs) play an important role in mucosal tolerance by controlling inflammation and modulating immune responses initiated by T effector cells (Teffs). Tregs also provide vital survival factors to IgA+ B cells in the intes-tine in order to support Immunoglobulin A (IgA) responses as part of a protective Treg-IgA pathway, which functions to maintain mutualism with the microbiota and promote intestinal health. The binding subunit of cholera toxin (CT), cholera toxin B (CTB), has been demonstrated to play a role in tolerance by inducing antigen-specific Foxp3+ Tregs in vitro and in vivo when conjugated to defined antigens. In order to study the regulation of the Treg-IgA pathway, we have generated a construct, CBirTox, composed of a por-tion of the C-terminus of the mucosal flagellin CBir1, an immunodominant microbiota antigen, genetically coupled to the A2 subunit of CT and expressed with CTB. Treatment of antigen-presenting cells (APCs) with CBirTox selectively induces Foxp3 expression in B6.CBir1 TCR Tg CD4+ T cells in vitro. The Foxp3 induction is dependent on basal TGF-ß levels, as neutralization of the cytokine abrogates Foxp3 expression, but acts in-dependently of retinoic acid (RA) signaling. APCs pulsed with CBirTox downregulate expression of phosphorylated p70S6 kinase, a protein directly downstream of mammalian target of rapamycin (mTOR), indicating CBirTox partially inhibits mTOR signaling. Ad-ditionally, CBirTox augments CD4+Foxp3+ Tregs in vivo. Furthermore, CBirTox-pulsed dendritic cells induce significant levels of IgA production from naïve CD43- splenic B cells in vitro, independent of additional stimulation. This induction is blunted by inhibi-tors to RA and TGF-ß receptor I kinase III. Collectively, our data demonstrate a role for CBirTox in inducing tolerance by augmenting the Treg-IgA pathway via induction of Foxp3+ Tregs and initiation of IgA responses from naïve B cells.

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