Modulation of cellular functions in retroorbital fibroblasts using antisense oligonucleotides targeting the c-myc protooncogene

A. E. Heufelder, R. S. Bahn

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

Purpose. To examine the signal transduction pathways involved in the activation of orbital fibroblast effector functions relevant to the pathogenesis of Graves' ophthalmopathy (GO). To determine, using antisense technology, whether the c-myc protooncogene is involved in cell proliferation and glycosaminoglycan (GAG) synthesis in cultured orbital fibroblasts (OF). Methods. The effects of a 16-met c-myc antisense phosphorothioate oligodeoxynucleotide (S-ODN) on OF monolayers derived from orbital connective tissue of patients with severe GO (n = 6) and healthy individuals (n = 3) were investigated. Quiescent OF monolayers were treated with serum or cytokines and were exposed to increasing concentrations of a c-myc antisense S-ODN and several control S-ODN. Cell proliferation was quantitated by direct cell counting and by immunocytochemistry for the nuclear Ki-67 antigen. Glycosaminoglycan synthesis was examined by [3H] GAG analysis. The effects of the c-myc antisense S-ODN and control S-ODN on c-myc mRNA and protein product levels were analyzed using reverse-transcriptase polymerase chain reaction, immunocytochemisty, and immunoblotting, respectively. Results. Transient suppression of c-myc mRNA and the c-myc protein product by a c-myc antisense S-ODN (2 to 8 μM) strongly inhibited cell proliferation and GAG synthesis in OF derived from patients with GO and healthy individuals. These effects occurred in a dose-dependent manner and were specific for the c-myc antisense S-ODN used. Cell morphology or viability were not affected. Conclusions. The c-myc protooncogene and its protein product are involved in the proliferative and metabolic activities of OF exposed to serum or cytokines in vitro. C myc appears to be an essential component of at least two OF cellular activities likely to contribute to the orbital tissue alterations in GO.

Original languageEnglish (US)
Pages (from-to)1420-1432
Number of pages13
JournalInvestigative Ophthalmology and Visual Science
Volume36
Issue number7
StatePublished - Jan 1 1995

Keywords

  • Graves' disease
  • antisense
  • c- myc
  • retroorbital fibroblasts
  • thyroid-related ophthalmopathy

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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