Date of Award

2019

Document Type

Dissertation - MUSC Only

Degree Name

Doctor of Philosophy (PhD)

Department

Microbiology and Immunology

College

College of Graduate Studies

First Advisor

Carl Atkinson

Second Advisor

Amanda C. LaRue

Third Advisor

Bei Liu

Fourth Advisor

Satish Nadig

Fifth Advisor

Demetri Spyropoulos

Sixth Advisor

Stephen Tomlinson

Abstract

The survival associated with double lung transplantation (LTx) is 7.1 years, however, closer inspection of recipient pre-LTx disease demonstrates that survival amongst COPD patients (related to cigarette smoke [CS] exposure) is amongst the worst at 5.5 years, as compared to 8.9 years in cystic fibrosis patients. A number of risk factors are thought to lead to this observed inferior outcome, chief amongst these, are pre-LTx diagnosis, recipient and donor age, and donor CS history. Indeed, CS exposure in either the recipients or donors has been linked with increased rates of severe primary graft dysfunction, acute rejection (AR), and ultimately earlier graft failure. The goal of these proposed studies is to demonstrate evidence that CS exposure, either donor or recipient, alters the local lung-specific immune environment in such a way as to predispose to exacerbated ischemia-reperfusion injury (IRI), with a resultant worsening of AR. We first examined the role of CS exposure on the lung microenvironment, and how this impacts post-LTx outcomes. We accomplished this goal by studying CS donor lungs, which allowed us to delineate lung-specific perturbations in the immune response to LTx exclusive from systemic CS-related immune derangements that may be present in CSexposed LTx recipients. Using the murine orthotopic left LTx model, a model that only a few research centers in the world employ, we were able to examine how donor CS exposure alters the complement cascade, and how these alterations could be therapeutically targeted to ameliorate AR. Importantly, we were able to demonstrate that systemic complement deficiency is inadequate to lessen AR. Instead, our results suggested that local complement production within the lung microenvironment appears to be far more important of a driver of post-LTx injury. Additionally, our data demonstrated that lung specific complement expression may also be correlated to peri-LTx primary graft dysfunction (PGD) in actual human studies. In the second aspect of our studies, we examined recipient CS exposure and how the associated immune-modulating effects of CS would impact LTx. We found in a series of experiments that in recipients with previous CS exposure, there was a pre-existing autoreactive antibody phenotype that not only exacerbated IRI but could actually drive IRI in immunodeficient transplant models. This autoreactive antibody response was mediated through a complement centric response, which we were then able to target to abrogate the injury phenotype. Using our novel C2-crry construct to target complement inhibition to post-ischemic neoepitopes on transplanted lungs, we showed that even in a model with a plethora of pre-existing autoreactive antibodies, as in our CS model, this type of therapeutic intervention could be effective. C2 is a so-called “natural” IgM antibody analog and targets phospholipid epitopes exposed through IRI. The ability of this targeting to effectively dampen the autoantibody driven IRI we demonstrated in CS-recipient LTx portends significant potential for the clinical translation of these findings. Finally, the impact of recipient CS exposure on the AR phenotype post-LTx. While our original intent was to explore the impact of pre-existing autoreactivity in the adaptive immune cell compartment, our findings led us to novel discoveries in T cell biology of LTx rejection. We utilized a systems biology approach in which data acquired from multi-gene array were analyzed using in silico predictive modeling to implicate the phosphoinositide 3-kinase (PI3K) pathway as a differential effector in CS LTx recipients. Not only were we able to demonstrate that PI3K inhibition could reverse the CS-associated rejection phenotype, but that it appeared to selectively be acting through CD8+ T cell specific mechanism. We found that the inhibition of PI3K in CD8+ T cells resulted in important functional and phenotypic differences, including the down regulation of IL-4, IL-6, and IL 17, all of which had been predicted in our in silico modeling. Our results also highlighted the importance of central memory CD8+ T cells in tolerance, a finding which had previously been published, but through PI3K inhibition, we demonstrated a viable therapeutic target that could be translated clinically to drive this tolerogenic shift in the adaptive immune response to LTx.

Rights

All rights reserved. Copyright is held by the author.

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