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

Abstract

BackgroundThe 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.MethodsAdult 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.ResultsIntravitreal 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.ConclusionsOur 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.

Subretinal Infiltration of Monocyte Derived Cells and Complement Misregulation in Mice with AMD-Like Pathology

“…To further phenotype the subretinal cells, we immunostained them for markers of monocyte-derived cells including Iba1, CD11b, CD68 (ED-1), and the lectin GS-IB4 [6–7] in separate preparations. Iba1, CD11b, and CD68 antibodies labeled all of the subretinal cells in their respective preparations, while GS-IB4 labeled only a subset of them (Figure 2).…”

Cellular responses following retinal injuries and therapeutic approaches for neurodegenerative diseases

“…Functional interactions between microglia and Müller glial cells may be bidirectional and regulate photoreceptor cell survival during retinal degeneration (Harada et al, 2002). In this sense, chemokine ligand 2 expression by Müller cells seems to play a role in promoting the infiltration of monocytes/microglia and contribute to the neuroinflammatory response and photoreceptor death following retinal injury (Rutar et al, 2012). Still other researchers have observed that after photodegeneration, microglia fail to return to their original state, rather they continued to show some degree of activation (Santos et al, 2010).…”

Neuroprotective actions of progesterone in an in vivo model of retinitis pigmentosa

“…The recruitment of glia to damaged regions occurs in almost every pathological condition in the CNS and in a range of retinal pathologies, including age-related macular degeneration, RP, late-onset retinal degeneration, retinal detachment, glaucoma, diabetic retinopathy and in many experimental models of retinal degeneration [49][50][51][52][53][54][55][56][57]. The most well known aspect of the glial response in the retina is that Müller cells upregulate GFAP, a protein considered to be a marker for reactive gliosis, not only in the retina, but also across the entire CNS [58].…”