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J. A. Matsubara, G. Y. Tsai, H. Syed, J. Z. Cui, Z. Xia, J. H. McNeill; Early Expression of Inflammatory Markers in the Retina of STZ Rats. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1375. doi: https://doi.org/.
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Both an inflammatory response and oxidative stress have been implicated in the pathogenesis of diabetic retinopathy (DR). The purpose of this study is to investigate the relative expression of immunohistochemical markers of inflammation and oxidative stress in an animal model of diabetic retinopathy, and the influence of treatment by a free radical scavenger, N-acetylcysteine (NAC).
Eye tissues were obtained from 4 groups of animals: control rats (group C), rats undergoing 9 weeks of STZ induced diabetes (group D), STZ induced diabetic rats following 8 weeks of NAC treatment (group DT), or control rats following 8 weeks of NAC treatment (group CT). Immunohistochemical methods were used to identify the presence of vascular cells and growth factors, and markers of inflammation and oxidative stress in the retinal layers of paraffin embedded or frozen eye tissues. Primary antibodies against total macrophages (ED-1), microglia cells (Ox-42), pericytes (NG-2), endothelial cells (IB-4), heme oxidase 1 (HO-1) and vascular endothelial growth factor (VEGF) were used in combination with secondary antibodies tagged with Alexa-488 or biotin. A confocal laser scanning microscope (Zeiss-LSM 510 META) and a standard bright field light microscope were used to visualize immunohistocehmical labeling. Tonsil, spleen, and liver tissue were used as positive control tissues.
Expression of ED-1 was robust in the D group, and mild in the C, CT and DT groups, while expression of NG-2 was robust in C, CT and DT groups, and mild in D group. The intensity of IB-4 staining was higher in D and DT groups (194.3±45.7 and 180.7±32.8) compared to the C and CT groups (112.4±40.6 and 102.5±30.8). Ox-42 expression was higher in the D group (25.3±3.2) than in the DT (10.3±3.2), C (10.0±2.6), or CT group (9.7±1.0). Expression of VEGF and HO-1 was non-specific in all 4 groups
Preliminary data suggests that macrophage and microglia upregulation, along with pericyte loss and endothelial cell proliferation occur early in the pathogenesis of DR. These changes appear to be minimized by treatment with NAC. The results suggest that therapeutic applications focused on reducing free radicals may be helpful in minimizing the early events in diabetic retinopathy in the STZ model.
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