April 2009
Volume 50, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2009
Proteomic Analysis of TNF- Signaling in Human Glaucoma
Author Affiliations & Notes
  • X. Yang
    Ophthalmology & Visual Sciences,
    University of Louisville, Louisville, Kentucky
  • C. Luo
    Ophthalmology & Visual Sciences,
    University of Louisville, Louisville, Kentucky
  • D. W. Powell
    Biochemistry & Molecular Biology,
    University of Louisville, Louisville, Kentucky
  • J. B. Klein
    Biochemistry & Molecular Biology,
    Medicine,
    University of Louisville, Louisville, Kentucky
  • M. H. Kuehn
    Ophthalmology & Visual Sciences, University of Iowa, Iowa City, Iowa
  • G. Tezel
    Ophthalmology & Visual Sciences,
    Anatomical Sciences & Neurobiology,
    University of Louisville, Louisville, Kentucky
  • Footnotes
    Commercial Relationships  X. Yang, None; C. Luo, None; D.W. Powell, None; J.B. Klein, None; M.H. Kuehn, None; G. Tezel, None.
  • Footnotes
    Support  NEI grants, 2R01EY013813, 1R01EY017131, and R24EY015636, and RPB
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 4043. doi:
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    • Get Citation

      X. Yang, C. Luo, D. W. Powell, J. B. Klein, M. H. Kuehn, G. Tezel; Proteomic Analysis of TNF- Signaling in Human Glaucoma. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4043.

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

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Abstract

Purpose: : Previous evidence supports the involvement of TNF- signaling in glaucomatous neurodegeneration. In respect to diverse bioactivities of TNF-, which can induce both cell death and survival signals, ongoing proteomic studies aim to further elucidate TNF- signaling in glaucoma to identify specific treatment targets for inhibition of cell death signaling and amplification of cell survival.

Methods: : Human retinal protein samples were obtained from ten donors with (n:5) or without (n:5) glaucoma. Protein expression was determined in tryptic digests of protein samples by 2-dimensional capillary liquid chromatography coupled with tandem mass spectrometry. BIGCAT statistical platforms were utilized to establish a protein abundance factor for comparison of protein enrichment within and between samples. Mass spectral data were also analyzed by bioinformatic tools using ARIADNE Pathway Studio and Ingenuity Pathways Analysis. For further validation of the proteomic findings, quantitative Western blot analysis was performed using specific antibodies. In addition, the extent and cellular localization of selected proteins were comparatively determined by immunohistochemical analysis of human donor eyes with (n:34) or without glaucoma (n:20).

Results: : Proteomic analysis of human retinal samples detected expression of multiple proteins involved in the TNF- signaling pathway and their differential regulation in human glaucoma. Proteins identified by mass spectrometry included a number of downstream adaptor/interacting proteins, such as TRAF-1, -2, -5, RIP, GRB2, SOS1, RAS, caspases, Bid, and different members of MAPKs and I-KKs. Bioinformatic analysis of the high-throughput comparative mass spectral data established extended maps of death-promoting and survival-promoting pathways of TNF- signaling, and functional networks revealed specific signaling molecules involved in diverse bioactivities. Western blot analysis and immunohistochemistry using specific antibodies to selected proteins validated protein expression and localization in retinal ganglion cells and glia.

Conclusions: : These findings provide new information improving the understanding of TNF- signaling in human glaucoma at the protein level. Ongoing studies should facilitate efforts searching neuroprotective treatment possibilities for glaucoma patients.

Keywords: proteomics • ganglion cells • apoptosis/cell death 
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