Characterization of an Alternatively Spliced Platelet-Derived Growth Factor Variant During Myelin Synthesis
Ng, Benjamin
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https://hdl.handle.net/2142/84811
Description
Title
Characterization of an Alternatively Spliced Platelet-Derived Growth Factor Variant During Myelin Synthesis
Author(s)
Ng, Benjamin
Issue Date
2005
Doctoral Committee Chair(s)
Glaser, Michael
Department of Study
Biochemistry
Discipline
Biochemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Neuroscience
Language
eng
Abstract
An alternatively spliced platelet-derived growth factor (PDGF) A-chain isoform was detected in rat dorsal root ganglion (DRG) neuron/Schwann cell cocultures and shown to regulate myelin synthesis. mRNA transcripts of the long-form PDGF-A chain variant (LF-PDGF-A) were detected by RT-PCR in cocultures. Its expression level increased during active myelin synthesis in contrast to those of the short form variant (SF-PDGF-A). LF-PDGF-A mRNA transcripts were localized to the neuronal cell bodies in coculture with Schwann cells using in situ hybridization. LF-PDGF-A protein was detected along axon and Schwann cell interfaces both in coculture and in the developing sciatic nerve. An increase in the expression of the PDGF alpha-receptor was also seen during these time points in Schwann cells both in vitro and in vivo. The PDGF A-chain isoform genes were isolated from cocultures and expressed and purified from bacteria. An enhancement in MBP expression during myelin synthesis was seen in cocultures treated with purified PDGF A-chain. Relatively lower concentrations of LF-PDGF-A were able to evoke this response in comparison to SF-PDGF-A. A corresponding decrease in MBP expression was observed through the displacement of LF-PDGF-A from cellular surfaces by exogenous heparin. MBP expression levels were restored in heparin treated cocultures with the addition of exogenous LF-PDGF-A. In addition, a decrease in progesterone receptor expression and translocation was seen in DRG neurons by exogenous heparin treatment, suggesting the involvement of LF-PDGF-A in directing progesterone synthesis. Progesterone has previously been shown to enhance myelin synthesis in cocultures. Exogenously added LF-PDGF-A also increased the expression of 3beta-hydroxysteroid dehydrogenase in Schwann cells, an enzyme committed to the synthesis of progesterone. Through the use of multicompartmental (Campenot) chambers, LF-PDGF-A expression in DRG neurons and their localization on axons was confirmed. Myelin synthesis was enhanced when axons and Schwann cells were locally exposed to exogenous LF-PDGF-A. Progesterone translocation to the DRG cell body from Schwann cells through retrograde axonal transport was demonstrated with Campenot chambers. Lastly, progesterone precursors and metabolites were shown to be differentially synthesized in Schwann cells and DRG neurons during myelin synthesis.
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