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Omar Delgado, Maura Crowley, Wei Zheng, Casey Lewis, Kathryn McAllister, Bruce D Jaffee, Sha-Mei Liao; Dynamic changes of M1 and M2 macrophages in a murine model of endotoxin-induced uveitis. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3093.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the roles of M1 and M2 macrophages in TLR4-mediated acute ocular inflammation using a murine model of endotoxin-induced uveitis (EIU). M1 macrophages promote inflammation, while M2 macrophages have anti-inflammatory and wound-healing properties. Both M1 and M2 macrophages have been implicated in the pathogenesis of age-related macular degeneration.
EIU was induced by an intraperitoneal injection of lipopolysaccharide (LPS), and tissues were collected at various time points. RNA samples were prepared for microarray (transcriptional profiling) and Taqman (gene expression) analysis and protein extracts were prepared for Western blot and multiplex cytokine assays (Aushon). To quantitate leukocyte infiltration, retinas were immunostained for Gr-1+ neutrophils and F4/80+ macrophages, while posterior eye cups (RPE-choroid-sclera) were stained for Iba1+ microglia cells.
Within 24 hours after an intraperitoneal injection of LPS, genes encoding M1 macrophage markers such as iNOS, CD40, IL-6, and IL-1b were up-regulated in eye tissues both the retina and in the RPE-choroid-sclera. In contrast, M2 macrophage markers Arg1, CD163, and CD209 were reduced during the first 24 hours as compared to PBS-treated controls. The M2 macrophage markers were elevated after 24 hours, and peaked between 3-5 days after LPS administration. Taqman analysis of several M1 and M2 genes correlated with the microarray results. In addition, the pro-inflammatory cytokine and chemokine proteins IL-6, Il-1b, IL-8, and MCP-1 were elevated at the same time as M1 gene induction (3-16 hours), followed by up-regulation of cell adhesion molecules, complement activation proteins, and Gr1+-neutrophil infiltrates in the retina (16 hours-2 days). The number of retinal F4/80+ macrophages and subretinal Iba1+ microglia cells increased in response to LPS and peaked around 3-5 days post LPS injection, correlating with the peak of M2 macrophage gene expression.
Intraperitoneal injection of LPS induces an acute inflammatory response in the eye by activating resident cells and M1 macrophages within the first 24 hours followed by M2 macrophages on day 3 to 5. These M1 macrophages are associated with inflammatory mediators and infiltrating neutrophils into the eye. This work identifies in vivo markers for M1 and M2 macrophages in the eye and demonstrates their role in the acute ocular inflammatory processes.
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