April 2011
Volume 52, Issue 14
Free
ARVO Annual Meeting Abstract  |   April 2011
Staurosporine Induced "Differentiation" of the RGC-5 Cell Line Leads to Apoptosis and Cell Death at the Lowest Cell-Morphology-Changing Concentration
Author Affiliations & Notes
  • Maximilian Schultheiss
    Departement of Ophthalmology, University of Tuebingen, Tuebingen, Germany
  • Sven Schnichels
    Departement of Ophthalmology, University of Tuebingen, Tuebingen, Germany
  • Kapka Miteva
    Berlin-Brandenburg Center for Regenerative Therapies, Campus Virchow, Charité, University-Medicine Berlin, Berlin, Germany
  • Katrin Warstat
    Berlin-Brandenburg Center for Regenerative Therapies, Campus Virchow, Charité, University-Medicine Berlin, Berlin, Germany
  • Peter Szurman
    Departement of Ophthalmology, University of Tuebingen, Tuebingen, Germany
  • Martin S. Spitzer
    Departement of Ophthalmology, University of Tuebingen, Tuebingen, Germany
  • Sophie van Linthout
    Berlin-Brandenburg Center for Regenerative Therapies, Campus Virchow, Charité, University-Medicine Berlin, Berlin, Germany
    Department of Cardiology & Pneumology, Campus Benjamin Franklin, Charité, University- Medicine Berlin, Berlin, Germany
  • Footnotes
    Commercial Relationships  Maximilian Schultheiss, None; Sven Schnichels, None; Kapka Miteva, None; Katrin Warstat, None; Peter Szurman, None; Martin S. Spitzer, None; Sophie van Linthout, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 4608. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Maximilian Schultheiss, Sven Schnichels, Kapka Miteva, Katrin Warstat, Peter Szurman, Martin S. Spitzer, Sophie van Linthout; Staurosporine Induced "Differentiation" of the RGC-5 Cell Line Leads to Apoptosis and Cell Death at the Lowest Cell-Morphology-Changing Concentration. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4608.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose: : Supplementation of staurosporine is thé method of choice to differentiate the solely existing retinal ganglion cell (RGC)-5 cell line. This "differentiation" is claimed to be in the absence of apoptosis. In respect to the inconsistencies of RGC-5 and the known apoptosis-inducing effect of staurosporine, we investigated whether RGC-5 cells could be differentiated by staurosporine without the induction of apoptosis.

Methods: : 50-, 200-, 300- and 600 nM of staurosporine were supplemented on RGC-5 cells for 24 hours. Cell morphology and cell death, via propidium iodide staining, were investigated with phase contrast and fluorescence microscopy, respectively. Cell amount, cell proliferation and cell viability were analyzed by crystal violet staining, CFSE flow cytometry and MTS assay, respectively. Apoptosis was determined by analyzing caspase 3/7 activity, Annexin-V+/ 7AAD cells and the mRNA expression of the Bax/ Bcl-2 quotient via caspase 3/7 activity assay, flow cytometry and real-time PCR, respectively.

Results: : RGC-5 cells started to "differentiate" at 50 nM of staurosporine resulting in neurite formation. This was associated with apoptosis and cell death. Apoptosis was indicated by a 2.1-fold (p<0.0005) increase in caspase 3/7 activity, a 1.2-fold (p<0.05) induction of Annexin-V+/ 7AAD- cells, whereas cell death was indicated by a 11.7-fold (p<0.0005) increase in propidium iodide positive cells. Furthermore staurosporine led to a dose-dependent increase in apoptosis and reduction in cell viability, cell density and cell proliferation.

Conclusions: : The lowest staurosporine concentration changing RGC-5 cell`s morphology is accompanied with apoptosis and cell death. Therefore, its use as differentiation-agent is at least very doubtful.

Keywords: ganglion cells 
×
×

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.

×