July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Midkine-a deficiency causes cell cycle arrest and reactive gliosis in zebrafish Müller glia following photoreceptor cell death
Author Affiliations & Notes
  • Mikiko Nagashima
    Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, Michigan, United States
  • Travis D'Cruz
    Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, Michigan, United States
  • Doneen Hesse
    Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, Michigan, United States
  • Peter F Hitchcock
    Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, Michigan, United States
  • Footnotes
    Commercial Relationships   Mikiko Nagashima, None; Travis D'Cruz, None; Doneen Hesse, None; Peter Hitchcock, None
  • Footnotes
    Support  NIH Grant EY07060
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 3115. doi:
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      Mikiko Nagashima, Travis D'Cruz, Doneen Hesse, Peter F Hitchcock; Midkine-a deficiency causes cell cycle arrest and reactive gliosis in zebrafish Müller glia following photoreceptor cell death. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3115.

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

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Abstract

Purpose : Midkine (Mdk) is a cytokine/ growth factor with multiple roles in neural development, tissue repair and disease. During retinal development in zebrafish, Mdka, encoded by one of two paralogous midkine genes, functions to govern elements of the cell cycle. To explore the role of Mdka during regenerative neurogenesis, we generated mutant zebrafish using CRISPR-Cas9 gene editing and assayed photoreceptor regeneration following the selective death of photoreceptors.

Methods : Zebrafish lines, AB-wildtype and Tg(gfap:GFP)mi2002 were used in this study. CRISPR/Cas9 mediated midkine-a mutation was introduced in the AB-background. TUNEL assay was used to label dying cells following photolytic lesion. Immunocytochemistry was used to assay Müller glia dedifferentiation, progenitor proliferation, and photoreceptor regeneration. Quantitative PCR was used to quantify the expression of cell cycle regulators.

Results : In Mdka mutants, Müller glia, the intrinsic stem cell in the zebrafish retina, respond to the selective death of photoreceptors by partial dedifferentiation, characterized by increased expression of stem cell-associated genes, such as Rx1, Sox2, and ascl1a. These Müller glia then partially reenter the cell cycle, however, cell cycle transition from G1 to S phase is blocked, resulting in significant reduction of proliferation and photoreceptor regeneration. Importantly, Müller glia that fail to enter the cell cycle undergo reactive gliosis, a hallmark of the glial response following injury in the central nervous system in mammals. In addition, in Mdka mutants, microglia, the intrinsic macrophages of the central nervous system, respond abnormally to the apoptotic death of photoreceptors. Although photoreceptor apoptosis follows the time frame observed in wildtype animals, microglia in Mdka mutants fail to phagocytose dying photoreceptors, resulting in the persistence of apoptotic nuclei and an abnormal accumulation of reactive microglia among the dying photoreceptors.

Conclusions : These results demonstrate that Mdka functions as a key determinant of stem cell property of Müller glia during regenerative neurogenesis. Mdka also functions to orchestrate aspects of inflammatory events following neuronal death and mediates the interaction between microglia/macrophages and dying cells.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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