Nobuo Jo1, Guey-Shuang Wu, Narsing A Rao. 1. Doheny Eye Institute and the Department of Ophthalmology, Keck School of Medicine of the University of Southern California, Los Angeles, California 90033, USA.
Abstract
PURPOSE: To evaluate chemokine expression at various retinal sites after ischemia-reperfusion injury, using reverse transcription-polymerase chain reaction (RT-PCR) analysis of selected tissue obtained by laser capture microdissection. METHODS: Retinal ischemia was produced in Lewis rats by increasing intraocular pressure for 75 minutes. At 3, 6, 12, and 24 hours after reperfusion, RT-PCR was used to measure the levels of monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, interleukin (IL)-8, and interferon-gamma-inducible 10-kDa protein (IP-10) mRNA expression in the ganglion cell layer (GCL), inner nuclear layer (INL), outer nuclear layer (ONL), and retinal vessels, after laser capture microdissection of these retinal layers. These chemokines were further localized by immunohistochemical methods, using antibodies specific to MCP-1 and MIP-1alpha. Leukocyte infiltration into the retina was detected with immunostaining for leukocyte common antigen. RESULTS: Ischemia-reperfusion induced expression of MCP-1, MIP-1alpha, and MIP-1beta mRNA in the retinal vessels 3 hours after reperfusion. Six hours after reperfusion, expression of these chemokines and IL-8 mRNA was seen in the GCL and INL. Twelve hours after reperfusion, IP-10 mRNA expression was seen in the GCL and INL. Immunoreactive MCP-1 and MIP-1alpha were detected in the GCL, INL, and the retinal vessels 24 hours after reperfusion. No chemokine mRNA expression or immunoreactivity was detected in the ONL at any time. Leukocyte infiltration was noted at 12 hours, increasing markedly 24 hours after reperfusion. CONCLUSIONS: Ischemia-reperfusion retinal injury results in generation of highly chemotactic agents, initially in the retinal vasculature, then in the other inner retinal layers. Such differential chemokine expression may play a role in leukocyte recruitment and selective leukocyte infiltration in the inner retina, leading to retinal damage primarily localized to the ganglion cells and other inner neuronal structures.
PURPOSE: To evaluate chemokine expression at various retinal sites after ischemia-reperfusion injury, using reverse transcription-polymerase chain reaction (RT-PCR) analysis of selected tissue obtained by laser capture microdissection. METHODS:Retinal ischemia was produced in Lewis rats by increasing intraocular pressure for 75 minutes. At 3, 6, 12, and 24 hours after reperfusion, RT-PCR was used to measure the levels of monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, interleukin (IL)-8, and interferon-gamma-inducible 10-kDa protein (IP-10) mRNA expression in the ganglion cell layer (GCL), inner nuclear layer (INL), outer nuclear layer (ONL), and retinal vessels, after laser capture microdissection of these retinal layers. These chemokines were further localized by immunohistochemical methods, using antibodies specific to MCP-1 and MIP-1alpha. Leukocyte infiltration into the retina was detected with immunostaining for leukocyte common antigen. RESULTS:Ischemia-reperfusion induced expression of MCP-1, MIP-1alpha, and MIP-1beta mRNA in the retinal vessels 3 hours after reperfusion. Six hours after reperfusion, expression of these chemokines and IL-8 mRNA was seen in the GCL and INL. Twelve hours after reperfusion, IP-10 mRNA expression was seen in the GCL and INL. Immunoreactive MCP-1 and MIP-1alpha were detected in the GCL, INL, and the retinal vessels 24 hours after reperfusion. No chemokine mRNA expression or immunoreactivity was detected in the ONL at any time. Leukocyte infiltration was noted at 12 hours, increasing markedly 24 hours after reperfusion. CONCLUSIONS:Ischemia-reperfusion retinal injury results in generation of highly chemotactic agents, initially in the retinal vasculature, then in the other inner retinal layers. Such differential chemokine expression may play a role in leukocyte recruitment and selective leukocyte infiltration in the inner retina, leading to retinal damage primarily localized to the ganglion cells and other inner neuronal structures.
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