April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Differentiation of Neuroblasts Into Photoreceptors in Co-Cultures of Muller Glial Cells and Neurons: Regulation of Apoptosis by Docosahexaenoic Acid and Sphingosine-1-Phosphate
Author Affiliations & Notes
  • L. E. Politi
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • M. V. Simon
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • P. De Genaro
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • C. E. Abrahan
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • E. B. De los Santos
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • N. P. Rotstein
    Neurobiology, Inst de Invest Bioquimicas, Bahia Blanca, Argentina
  • Footnotes
    Commercial Relationships  L.E. Politi, None; M.V. Simon, None; P. De Genaro, None; C.E. Abrahan, None; E.B. De los Santos, None; N.P. Rotstein, None.
  • Footnotes
    Support  ANPCYT PICT-2006-00711; CONICET PIP 6529, and UNS 24/146
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 5941. doi:
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      L. E. Politi, M. V. Simon, P. De Genaro, C. E. Abrahan, E. B. De los Santos, N. P. Rotstein; Differentiation of Neuroblasts Into Photoreceptors in Co-Cultures of Muller Glial Cells and Neurons: Regulation of Apoptosis by Docosahexaenoic Acid and Sphingosine-1-Phosphate. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5941.

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

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Abstract

Purpose: : Muller glial cells (MGCs) are stem cells that might be used to replace lost photoreceptors (PHRs) during neurodegenerative diseases of the retina such as retinitis pigmentosa. We have previously shown that MGCs promote generation and survival of round undifferentiated multipotent neuroblasts (NBs) in primary and secondary neuroglial cocultures. We now investigated whether MGCs promote NBs to differentiate into PHRs and if the PHR survival factors, DHA and sphingosine-1-phosphate (S1P) might prevent apoptosis in these neurons.

Methods: : Primary and secondary neuro-glia co-cultures from newborn rat retinas were supplemented with DHA or S1P. We measured BrdU uptake to evaluate proliferation, DAPI and TUNEL labeling to quantitate apoptosis, and the presence of nestin, and Crx and opsin, as stem cell, and PHR markers, respectively. PHR functionality was assessed by high affinity [3H]-Glutamate uptake and cGMP depletion in dark and light exposed cultures.

Results: : Nearly 80% and 10% of round cells occurring in secondary neuro-glial co-cultures expressed Crx and opsin respectively, and about 1% of these cells co-labeled with BrdU, suggesting that part of the NBs first divided in vitro and then differentiated into PHR-like cells. Interestingly, many of these cells displayed properties of functional PHRs, such as high affinity uptake of [3H]-Glutamate and light response, reducing cGMP levels after bright light illumination. Noteworthy, incubating the co-cultures with either DHA or S1P reduced the apoptosis of the differentiated PHR-like cells by half.

Conclusions: : Our results show that Muller stem cells help to generate or preserve a pool of undifferentiated NBs, which then differentiated and showed properties of mature, functional PHRs. Thus MGCs appear to be critical to give rise to new neurons, while DHA and S1P might provide useful tools to prevent the apoptosis of newly generated PHRs during regenerative processes of the retina.

Keywords: Muller cells • photoreceptors • regeneration 
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