June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Characterization of a new retinal cell line (MU-PH1) expressing stem cell markers from adult mouse
Author Affiliations & Notes
  • Nicolas Cuenca
    Physiology, Genetics and Microbiology, University of Alicante SPAIN, Alicante, Spain
  • Violeta Gomez-Vicente
    Physiology, Genetics and Microbiology, University of Alicante SPAIN, Alicante, Spain
  • Ana Flores
    Microbiology and Ecology, University of Valencia, Valencia, Spain
  • Pedro Lax
    Physiology, Genetics and Microbiology, University of Alicante SPAIN, Alicante, Spain
  • Celia Murciano
    Microbiology and Ecology, University of Valencia, Valencia, Spain
  • Alberto Yañez
    Microbiology and Ecology, University of Valencia, Valencia, Spain
  • Maria Luisa Gil
    Microbiology and Ecology, University of Valencia, Valencia, Spain
  • Daniel Gonzalbo
    Microbiology and Ecology, University of Valencia, Valencia, Spain
  • Victoria Maneu
    Optics, Pharmacology and Anatomy, University of Alicante, Alicante, Spain
    Teófilo Hernando Institute of I+D of drugs, University Autonoma of Madrid, Madrid, Spain
  • Footnotes
    Commercial Relationships Nicolas Cuenca, Universidad de Alicante (P); Violeta Gomez-Vicente, Universidad de Alicante (P); Ana Flores, Universitat de Valencia (P); Pedro Lax, None; Celia Murciano, Universidad de Valencia (P); Alberto Yañez, Universidad de Valencia (P); Maria Luisa Gil, Universitat de Valencia (P); Daniel Gonzalbo, Universitat de Valencia (P); Victoria Maneu, Universidad de Alicante (P)
  • Footnotes
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Investigative Ophthalmology & Visual Science June 2013, Vol.54, 2222. doi:
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      Nicolas Cuenca, Violeta Gomez-Vicente, Ana Flores, Pedro Lax, Celia Murciano, Alberto Yañez, Maria Luisa Gil, Daniel Gonzalbo, Victoria Maneu; Characterization of a new retinal cell line (MU-PH1) expressing stem cell markers from adult mouse. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2222.

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

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Abstract

Purpose: To characterize a novel cell line, derived from retinal progenitors, which could serve as an in vitro model for the study of photoreceptor degenerative mechanisms and to evaluate the efficacy of neuroprotective compounds.

Methods: A Müller-derived cell line (MU-PH1) was isolated from adult C57BL/6 mouse retina. RT-PCR, immunoblot and immunofluorescence analyses were performed to characterize these cells. Calcium imaging allowed the determination of light-responsiveness. To establish a model for the screening of potential neuroprotective drugs, apoptosis was induced with cytotoxic agents and measured using XTT and crystal violet viability assays.

Results: We report the isolation and characterization of a novel, spontaneously immortalized, Müller-derived cell line (MU-PH1). These cells demonstrated a glial-like morphology and were able to form neurospheres under the appropriate culture conditions. RT-PCR, immunoblot and immunofluorescence techniques showed that MU-PH1 cells expressed the Müller cell markers vimentin, S-100 and glutamine synthetase, and several neural and stem cell markers (nestin, Abcg2, alpha-tubulin, beta-III-tubulin and Ascl1). The purity of the culture was confirmed, as the expression of other non-neuronal populations’ markers such as CRALBP (retinal pigment epithelial cells), GFAP (astrocytes) or CD31 (endothelium) was undetectable by RT-PCR. The presence of CD11b immunopositive cells (microglia) was also excluded by immunomagnetic purification. Interestingly, MU-PH1 cells expressed the photoreceptor markers rhodopsin, recoverin, transducin, melanopsin and blue and red/green opsins. Stimulation with 480 nm light evoked slow and fast transient calcium responses in most of the cells tested. MU-PH1s were sensitive to oxidative stress induced by sodium nitroprusside or oligomycin/rotenone and cell death was prevented by anti-apoptotic and antioxidant compounds such as tauroursodeoxycholate and N-acetylcysteine.

Conclusions: Availability of this mouse cell line could be of great interest as a photoreceptor model and will facilitate studies aimed to a better understanding of retinal Müller cell biology, as well as a convenient model for the screening and development of new therapeutic drugs.

Keywords: 694 retinal culture • 603 Muller cells • 503 drug toxicity/drug effects  
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