Date of Award

8-17-2009

Document Type

Thesis

Degree Name

Medical Doctor (MD)

First Advisor

Lloyd Cantley

Second Advisor

Erica L. Herzog, MD, PhD

Third Advisor

Richard Bucala, MD, PhD

Abstract

Idiopathic Pulmonary Fibrosis (IPF) is a rapidly progressing interstitial lung disease (ILD) with an unclear etiology and poorly understood pathogenesis. IPF has no known curative treatment and a grim prognosis. One emerging area of interest in the field of fibrosing diseases is the role that bone marrow-derived cells, specifically fibrocytes, play in lung repair and remodeling. Derived from circulating CD14+ progenitors in the peripheral blood, these cells are identified by CD45 and pro-Collagen I co-expression. There exist scant data regarding fibrocytes derived from the peripheral blood in patients with pulmonary fibrosis. We hypothesized that (1) fibrocyte levels in the peripheral blood of patients with IPF would be greater than in patients with non-specific interstitial pneumonia (NSIP), the second most common type of ILD; (2) intrapulmonary fibrocyte levels would vary significantly between patients with IPF and non-IPF pathology; and (3) plasma from subjects with high fibrocyte levels would reveal a profile of immunomediators differing from that of patients with lower fibrocyte levels, regardless of disease state. Peripheral blood from patients with IPF (n=16), NSIP (n=7), and age-matched healthy controls (n=15) was collected, their peripheral blood mononuclear cells (PBMCs) isolated by Ficoll separation, and their plasma stored for proteomic analysis. PBMCs and lung tissue digests from patients with fibrotic lung disease (n=9) or normal controls (n=3) were then co-stained for intracellular pro-Collagen I and CD45. Plasma underwent multi-analyte ELISA for a battery of immunologic targets. We found that patients with IPF had elevated fibrocytes as percentage of PBMCs (2.74±2.12%) when compared to NSIP (0.88±0.43%) and healthy controls (0.94±0.84%), as measured by flow cytometry for CD45 and pro-Collagen I co-expression (p < 0.05 NSIP vs. IPF). Furthermore, within the IPF group, fibrocyte levels varied with disease severity (p < 0.05). Interestingly, while lung tissue from patients with active fibrosing pulmonary processes contained elevated fibrocyte levels (3.97±2.7%) when compared with normal human lung tissue (1.04±0.43%, p=0.11), there was no difference between IPF and non-IPF pathology. Lastly, plasma from patients with higher fibrocytes as percentages of PBMCs revealed distinct immune mediator patterns. Specifically, patients with elevated fibrocytes had higher plasma levels of IL-18, IL-8, IL-10, β2-microglobulin, ICAM-1, and TIMP-1 (p < 0.05). No previous study has compared fibrocyte levels in patients with IPF and NSIP. These data suggest there is a relationship between circulating fibrocytes and development of IPF. Moreover, patients with more severe IPF disease had higher levels of circulating fibrocytes, suggesting that they may be a biomarker of both disease state and severity. In addition, lungs of patients with both IPF and non-IPF fibrotic lung disease contained more fibrocytes than healthy lungs, suggesting a broader role for this cell type in the pathogenesis of ILD than previously hypothesized. Lastly, proteomic analysis suggests that elevated fibrocytes are associated with a distinct immunologic profile, but these results do not correlate with an expected dichotomy of profibrotic TH2 versus TH1 immune profiles. These findings advance our understanding of fibrocytes as they relate to fibrotic lung disease and suggest novel avenues for future investigation.

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