June 2015
Volume 56, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2015
Involvement of REST-Sox9-miR-124 axis in neuronal conversion of the mammalian Müller Glia
Author Affiliations & Notes
  • Li Zheng
    Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, NE
  • Qulsum Mir
    Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, NE
  • Iqbal Ahmad
    Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, NE
  • Footnotes
    Commercial Relationships Li Zheng, None; Qulsum Mir, None; Iqbal Ahmad, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2015, Vol.56, 412. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to Subscribers Only
      Sign In or Create an Account ×
    • Get Citation

      Li Zheng, Qulsum Mir, Iqbal Ahmad; Involvement of REST-Sox9-miR-124 axis in neuronal conversion of the mammalian Müller Glia. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):412.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose: Recent observations that mammalian Müller glia (MG) possess neural stem cell (NSC) properties posit these cells as a potential target for regenerating diseased or injured photoreceptors and retinal ganglion cells (Ahmad et al., 2011, IOVS, 52(8):5758-64). However, the injury/disease-activated mammalian MG proliferate but convert into neurons rather infrequently (Del Debbio et al., 2010, Plos One, 5(8):e12425). To address this barrier to a practical therapeutic regeneration, we are examining the molecular axes that regulate differentiation of cells along the neuronal lineage. Here, we have examined the role of REST-Sox9-miR-124 axis in neuronal conversion of MG.

Methods: Rat MG were enriched using the hypoxia-based method, as previously described (Del Debbio et al., 2010). Enriched MG were cultured in the presence of FGF2 and Noggin to generate neurospheres. All experiments were carried out in the secondary neurospheres to ensure the enrichment of MG stem cells. MG neurospheres were transduced with miR-124 retrovirus/empty retrovirus or RESTsiRNA lentivirus/empty lentivirus or miR-124 retrovirus and REST siRNA lentivirus. Following transduction, neurospheres were plated on poly-D-Lysin plus laminin coated glass coverslips and cultured in the presence of conditioned medium obtained from embryonic day 18 (E18)/postnatal day 1 (PN1) rat retinal cells for eight days followed by qPCR and immunocytochemical analyses.

Results: The morphology of cells in both the gain-of function (GOF) of miR-124 and loss-of function (LOF) of REST was altered from controls. Cells in miR-124 GOF+REST LOF group had bipolar morphology, with few cells displaying neuronal morphology, compared to controls. qPCR analyses of cells in miR-124 GOF/REST LOF groups displayed higher levels of transcripts corresponding to proneural gene Mash1 and early neuronal marker gene βIII-tubulin, compared to controls. Levels of Mash1 and βIII-tubulin transcripts were significantly higher in cells in miR-124 GOF+REST LOF group, compared to those in miR-124 GOF/REST LOF groups. In all three GOF/LOF groups, levels of transcripts corresponding to Sox9, a MG regulator, were decreased with synergistic effects in miR-124 GOF+REST LOF group.

Conclusions: Our initial findings suggest that REST-Sox9-miR-124 axis is involved in MG regulation and a target for increasing the efficiency of neuronal conversion of MG.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×