All ETDs from UAB

Advisory Committee Chair

James S Hagood

Advisory Committee Members

Peter J Detloff

Joanne E Murphy-Ullrich

Thomas M Ryan

Tim M Townes

Document Type

Dissertation

Date of Award

2011

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Idiopathic Pulmonary Fibrosis (IPF) is characterized by chronic fibrosis in the lung interstitium of unknown etiology, and causes death within 2 to 4 years after diagnosis. There are no clinical interventions save for lung transplantation that give any survival benefit. Aberrant collagen deposition and aggregates of proliferating fibroblasts and myofibroblasts called fibroblastic foci (FF) are the classic features of IPF. Therefore, the cell type most often implicated as pathogenic in IPF is the fibroblast, especially its differentiated phenotype, the myofibroblast. Fibroblasts are a remarkably heterogeneous cell type. Expression of the cell surface glycoprotein Thy-1 (Thymocyte differentiation antigen 1) delineates a normal phenotype from a more fibrotic one. Thus, mechanisms that regulate Thy-1 expression could perhaps be exploited to abrogate phenotypes associated with disease. The initial goals of my work were to elucidate one such mechanism, namely Thy-1 shedding. To accomplish this, four specific aims were devised. They were to (1) determine the mechanism(s) by which pulmonary fibroblasts shed Thy-1 from their cell surface in response to pro-fibrotic stimuli. In conjunction with this, (2) characterize the C-terminus of Thy-1 released from fibroblasts and (3) identify residues of Thy-1 that modulate susceptibility to or are required for its shedding. Finally, (4) measure the concentrations of soluble Thy-1 and of identified sheddases in the bronchoalveolar lavage (BAL) fluid of patients with lung fibrosis and determine the degree to which they correlate. During the course of pursuing these aims it was determined that all the monoclonal antibodies to human THY1 available in the lab were incapable of recognizing delipidated THY1, e.g. soluble THY1. Taking this into consideration, we reevaluated the solubility of THY1 we reported detecting in the conditioned media (CM) of normal human lung fibroblast (NHLF) and found it to be entirely insoluble. From our and other's reporting there is a need to develop antibodies to THY1 that take into consideration the conformation of delipidated THY1. Additionally, the conformation of recombinant forms of THY1 used as controls in assays that rely on antibody recognition must be evaluated carefully. Beyond antibody recognition, the conformation of recombinant forms of THY1 used to elucidate its function should also be evaluated.

Share

COinS