May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Regulation of Gliosis in the DBA/2J Mouse Model of Glaucoma by NF-B and STAT3 Pathways
Author Affiliations & Notes
  • C. Lupien
    Neurological Surgery, University of Washington, Seattle, Washington
  • D. Inman
    Neurological Surgery, University of Washington, Seattle, Washington
  • P. J. Horner
    Neurological Surgery, University of Washington, Seattle, Washington
  • Footnotes
    Commercial Relationships  C. Lupien, None; D. Inman, None; P.J. Horner, None.
  • Footnotes
    Support  Glaucoma Research Foundation, Catalyst for a cure
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 5489. doi:https://doi.org/
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      C. Lupien, D. Inman, P. J. Horner; Regulation of Gliosis in the DBA/2J Mouse Model of Glaucoma by NF-B and STAT3 Pathways. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5489. doi: https://doi.org/.

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

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Abstract

Purpose: : Despite intense study, the mechanism of neurodegeneration in glaucoma is not understood. Neurodegenerative disease induces changes in glia comprising the condition known as reactive gliosis. Gliosis, the proliferation and hypertrophy of glial cells such as astrocytes and Müller cells, occurs early in the chronic mouse model of glaucoma (DBA/2J (D2)). There are numerous signaling pathways involved in gliosis development. Retinal gliosis is potentially triggered by several cytokines, which are activators of NF-ΚB and STAT3. The NF-ΚB pathway has been implicated in gliosis by its transcriptional control of cell adhesion molecules. Also, recent observations strongly implicate a link between activation of STAT3 and gliosis, as defined by enhanced expression of GFAP. As of yet, the positive or negative impact of gliosis on the progression of glaucoma has not been studied. The goal of this study is to determine the role of both pathways in gliosis by tempering the glial response in D2 mouse.

Methods: : To clarify the relative contribution of astrocytes and Müller cells in both pathways we used an in vitro paradigm. We cultivated astrocytes (P1 day) and Müller cells (P7 day) for 4 days before the addition of cytokines (CNTF, IL-6 or LIF) for the activation of the pathways. Once we determined that the retinal glial cells could be activated by different cytokines, cultured glial cells were exposed to a range of inhibitors (AG-490, PTP-2, JSI-124 or UDCA). Immunohistochemistry and Western blot analysis were performed using GFAP antibody as a marker of gliosis and the active form (phosphorylated)-NF-kB (pNF-ΚB) and -STAT3 (pSTAT3) antibodies both before and after inhibitor treatment.

Results: : By immunohistochemistry we have determined that NF-kB and STAT3 pathways are activated in D2 retinal glial cells. The addition of 10ng of CNTF over 24h induces the activation of STAT3 and GFAP expression in astrocytes and Müller cells. Also addition of 50ng of IL-6 or LIF gives us similar results. The addition of UDCA (IΚB inhibitor) to Müller cells reduced the GFAP protein expression and prevents translocation of pNF-ΚB to the nucleus. We also obtained similar results with the addition of PTP-2 (CNTF inhibitor), preceded by the addition of 10ng of CNTF. The GFAP expression and pSTAT3 translocation are reduced.

Conclusions: : By this study we have demonstrated that it is possible to modulate the NF-ΚB and STAT3 pathways in order to temper gliosis in an in vitro mouse model of glaucoma.

Keywords: retinal glia • signal transduction 
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