Dissertation/Thesis Abstract

Hepatic stellate cell transdifferentiation: Pro-fibrogenic effectors
by Lakner, Ashley Marie, Ph.D., The University of North Carolina at Charlotte, 2011, 179; 3474322
Abstract (Summary)

Hepatic fibrosis is a significant cause of morbidity and mortality worldwide and can be described as exacerbated wound-healing marked by excessive deposition of extracellular matrix components, predominantly type I collagen. Chronic injury stimulates accumulation of scar matrix synthesized by hepatic stellate cells (HSCs) resulting in distortion of normal liver architecture, disruption of blood flow and organ dysfunction. In a normal healthy liver, HSCs are found in the quiescent state functioning to store vitamin A in the form of retinyl esters, regulate sinusoidal microcirculation, and maintain normal basement membrane conditions. Following exposure to fibrogenic stimuli, HSCs transdifferentiate to activated myofibroblast-like cells marked by increased proliferative and hypercontractile properties.

Currently there are no FDA-approved treatments for hepatic fibrosis. Outside of surgical resection, orthotopic transplantation is an option for certain disease states; however, the current list of patients in need of transplant far exceeds the annual donor rate. As HSCs are considered the primary effector cells of fibrosis, targeting various points of transdifferentiation and resolution to impede disease development/progression is of therapeutic interest. The process of HSC transdifferentiation is classically divided into initiation and perpetuation phases, each characterized by unique cellular events.

Examination of genetic programming in early HSC transdifferentiation revealed coordinate changes in gene expression by components of JAK/STAT signaling. Chemical inhibition of this signaling pathway blocked HSC activation morphologically and through interruption of genetic reprogramming denoted by significantly decreased pro-fibrotic gene expression. Post-transcriptional regulation of HSC transdifferentiation (initiation and perpetuation phases) is controlled, in part, by changes in global microRNA (miR) expression patterns. Specifically, miR 19b was significantly decreased in activated compared to quiescent HSCs. Overexpression of miR 19b in culture-activated HSCs significantly decreased pro-fibrotic transforming growth factor beta (TGFβ) signaling by direct inhibition of TGFβRII and subsequent decreases in collagen expression and markers of HSC activation. miR 19b expression was also significantly decreased in rodent models of fibrosis and in liver tissue from fibrotic patients, indicating miR 19b as a novel biomarker and possible therapeutic. Finally, examination of transdifferentiation resolution has indicated involvement of HSC apoptosis. However, newly transdifferentiated stellate cells display increased resistance to apoptotic stimuli. Aquaporins (AQP) are known regulators of programmed cell death in mediation of the apoptotic volume decrease necessary for downstream caspase and nuclease activation. Analyses showed AQP expression was significantly decreased in activated compared to quiescent HSCs. Quiescent HSCs expressing multiple AQP homologs responded to osmotic challenge, an effect abrogated by functional inhibition of AQPs. Similarly, quiescent HSCs were significantly more responsive to apoptotic stimulation, which was also abrogated by AQP inhibition. In vitro studies were confirmed in vivo utilizing liver tissue from a mild-fibrotic animal model. Dual fluorescent immunohistochemistry showed strong colocalization of aquaporins with quiescent, but not activated stellate cells.

HSC transdifferentiation is highly complex with multiple pro-fibrotic signaling networks conferring phenotype-specific genetic reprogramming observed in hepatic fibrosis. These studies provide insight into novel factors controlling initiation, perpetuation and resolution stages of HSC transdifferentiation as well as new avenues to be explored for therapeutic intervention.

Indexing (document details)
Advisor: Schrum, Laura W.
Commitee: Clemens, Mark G., McKillop, Iain H., Nelson, Daniel A., Rabinovich, Daniel
School: The University of North Carolina at Charlotte
Department: Biology (PhD)
School Location: United States -- North Carolina
Source: DAI-B 73/01, Dissertation Abstracts International
Subjects: Molecular biology, Cellular biology
Keywords: Activation, Fibrosis, Hepatic stellate cells, Liver disease
Publication Number: 3474322
ISBN: 9781124906140
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