All ETDs from UAB

Advisory Committee Chair

Rosa Serra

Advisory Committee Members

Susan Bellis

Chenbei Chang

Amjad Javed

Joanne Murphy-Ullrich

Document Type

Dissertation

Date of Award

2018

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

THE MECHANISM OF TGF-β FUNCTION IN DEVELOPMENT OF THE AXIAL SKELETON GA I BAN CELL, MOLECULAR DEVELOPMENTAL BIOLOGY ABSTRACT Back pain is caused by degeneration of intervertebral discs (IVD). IVD consists of two compartments: the annulus fibrosus (AF) and the nucleus pulposus (NP). One strategy for treatment of IVD degeneration is stem cell therapy, which would require an in-depth knowledge of how the IVD develops. Currently, the molecular mechanism of IVD development are not well defined. AF and vertebral bodies (VB) are derived from the same embryonic progenitor, sclerotome. Specification of sclerotome is determined by complex interactions among various growth factors. Transforming growth factor beta (TGF-β) signaling plays an important role in development of fibrous tissues, including AF, tendon and ligament, whereas Bone Morphogenic Protein (BMP) regulates development of the VB. Cell fate decisions can be made directly, where a growth factor instructs progenitor cells to differentiate to a specific cell type (instructive signal). Alternatively, cell fate can be determined by inhibition of a particular cell type causing the cells to automatically switch to the other fate. A combination of these two processes is likely. We proposed that TGF-β provides an instructive signal to induce fibrous tissue differentiation in the sclerotome or/and inhibits chondrocyte cell fate by antagonizing BMP signaling, thus resulting in fibrous differentiation. How TGF-ß antagonizes BMP signaling and promotes fibrous differentiation are addressed in this thesis. First, we showed that TGF-β inhibits BMP signaling and chondrogenesis. BMP signaling was inhibited through a TGF-β-mediated increase in mRNA and protein levels of Noggin, a BMP antagonist. Inhibition of BMP signaling in itself was not sufficient to induce fibrous tissue differentiation, since Noggin and other BMP antagonists, Gremlin and DMH2, were not sufficient to induce expression of markers for fibrous differentiation. Since Noggin cooperated with TGF-β to induce fibrous tissue differentiation, we proposed that in addition to inhibiting BMP signaling, an instructive signal for fibrous tissue differentiation was also required. Next, the signaling pathway used by TGF-β to instruct fibrous differentiation was explored. We showed that the canonical TGF-β signaling pathway, which in involves Smad3, is required but not sufficient to induce expression of markers for fibrous differentiation suggesting a non-canonical signaling pathway is also required for TGF-ß to promote fibrous cell fate. We tested several known non-canonical signaling pathways and identified the Mapk ERK as being required for expression of fibrous markers in sclerotome. ERK was activated independently of Smad2/3 suggestion that these two signaling pathways do not act linearly but somehow cooperate to regulate cell fate decisions in the sclerotome. Future studies will further explore the instructive signals for fibrous tissue differentiation in the axial skeleton, including master transcriptional regulators of AF cell differentiation.

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