Small interfering RNA-mediated suppression of Ccl2 in Müller cells attenuates microglial recruitment and photoreceptor death following retinal degeneration

J Neuroinflammation. 2012 Sep 19;9:221. doi: 10.1186/1742-2094-9-221.

Abstract

Background: The recruitment and activation of inflammatory cells is thought to exacerbate photoreceptor death in retinal degenerative conditions such as age-related macular degeneration (AMD). We investigated the role of Müller cell-derived chemokine (C-C motif) ligand (Ccl)2 expression on monocyte/microglia infiltration and photoreceptor death in light-mediated retinal degeneration, using targeted small interfering (si)RNA.

Methods: Adult Sprague-Dawley rats were injected intravitreally with 1 μg of either Ccl2 siRNA or scrambled siRNA, and were then exposed to 1000 lux of light for a period of 24 hours. The mice were given an overdose of barbiturate, and the retinas harvested and evaluated for the effects of bright-light exposure. Ccl2 expression was assessed by quantitative PCR, immunohistochemistry, and in situ hybridization. Monocytes/microglia were counted on retinal cryostat sections immunolabeled with the markers ED1 and ionized calcium binding adaptor (IBA)1, and photoreceptor apoptosis was assessed using terminal dUTP nick end labeling.

Results: Intravitreal injection of Ccl2 siRNA significantly reduced the expression of Ccl2 following light damage to 29% compared with controls. In retinas injected with Ccl2 siRNA, in situ hybridization and immunohistochemistry on retinal cryostat sections showed a substantial decrease in Ccl2 within Müller cells. Cell counts showed significantly fewer ED1-positive and IBA1-positive cells in the retinal vasculature and outer nuclear layer of Ccl2 siRNA-injected retinas, compared with controls. Moreover, there was significantly less photoreceptor apoptosis in Ccl2 siRNA-injected retinas compared with controls.

Conclusions: Our data indicate that Ccl2 expression by Müller cells promotes the infiltration of monocytes/microglia, thereby contributing to the neuroinflammatory response and photoreceptor death following retinal injury. Modulation of exaggerated chemokine responses using siRNA may have value in reducing inflammation-mediated cell death in retinal degenerative disease such as AMD.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Analysis of Variance
  • Animals
  • Calcium-Binding Proteins / metabolism
  • Cell Death / drug effects*
  • Cell Death / radiation effects
  • Chemokine CCL2 / metabolism*
  • Disease Models, Animal
  • Ectodysplasins / metabolism
  • Gene Expression Regulation / drug effects
  • Gene Expression Regulation / radiation effects
  • In Situ Nick-End Labeling
  • In Vitro Techniques
  • Intravitreal Injections
  • Light / adverse effects
  • Mice
  • Microfilament Proteins / metabolism
  • Monocytes / drug effects
  • Monocytes / pathology
  • Monocytes / radiation effects
  • Neuroglia / drug effects*
  • Neuroglia / radiation effects
  • Photoreceptor Cells / drug effects
  • Photoreceptor Cells / pathology*
  • Photoreceptor Cells / radiation effects
  • RNA, Messenger / metabolism
  • RNA, Small Interfering / pharmacology*
  • RNA, Small Interfering / therapeutic use
  • Rats
  • Rats, Sprague-Dawley
  • Retinal Degeneration / drug therapy
  • Retinal Degeneration / etiology
  • Retinal Degeneration / pathology*

Substances

  • Aif1 protein, rat
  • Calcium-Binding Proteins
  • Chemokine CCL2
  • Ectodysplasins
  • Eda protein, mouse
  • Microfilament Proteins
  • RNA, Messenger
  • RNA, Small Interfering