Abstract 2330: Murine interferon-α inhibits aortic endothelial outgrowth
Authors: Hennings Leah J. , Upreti Meenakshi , Jamshidi-Parsian A.J. , Smith Amanda , Cottler-Fox Michele , Koonce Nathan , Henry Ralph , Griffin Robert J. 
Published: August 2012

Abstract
Although Interferon-α exerts anti-angiogenic effects, it has proven clinically useful in only a handful of tumor types. IFNα has a high degree of species specificity, and this can complicate mechanistic studies in animal models. We reported that human IFNα causes premature senescence of endothelial cells through effects on interferon regulatory factor 1 (IRF-1). We hypothesize that IFNα may also lead to increased expression of anti-angiogenic factors. Treatment of human endothelial cells with IFNα at 0.5µg/ml or 1.0µg/ml led to cell death with increased expression of ANG-2 and fibulin-5 in surviving cells. To establish the activity of a murine IFNα synthesized by our group for expanded study with mouse models, 2H11 murine endothelial cells were treated with murine IFNα at 0.05µg/ml, 0.5µg/ml, or 1.0µg/ml. Growth was inhibited at 0.05µg/ml and not at the higher doses. The expression of IRF-1, a downstream target of IFNα, was also upregulated in these cells upon treatment with 0.05µg/ml IFNα. The purpose of this study was to characterize the angiogenic response to murine IFNα in the more physiologically relevant mouse aortic outgrowth assay. Aortas were removed from 4 week old C57B/6 mice immediately following euthanasia and sectioned in 1mm increments. Sections from a single mouse were used for each replicate, and treatments were replicated 5 times. Sections were washed in basal medium, plated on a thin layer of Matrigel (50µl) in a 96-well plate, covered with 50µl Matrigel and incubated at 37°C. 24 hours after plating, sections were treated with IFNα at 0.5µg/ml, 1.0µg/ml, and 10µg/ml, or EGM-2 only. Sections were imaged at days 5 and 7. Photographs were digitally analyzed) to determine area and density of outgrowth and maximum vessel length. H&E stained paraffin-embedded sections were examined to evaluate vessel morphology. All IFNα treatments significantly decreased the area and density of aortic outgrowth. The effect was most pronounced at lower doses. Successful outgrowth was achieved in a similar number of treated and untreated sections. Maximum vessel length was decreased at both time points Outgrowths were characterized by linear growth perpendicular to the surface of the aorta, and robust branching. Capillary-type structure with lumen was evident in H&E sections. Vessel morphology did not differ with treatment. These data suggest that our newly synthesized murine IFNα has an antiangiogenic effect on endothelial cells. These results agree with our preliminary data showing murine endothelial cells undergo growth inhibition upon treatment with IFNα. Interestingly the response is only to the lower doses of murine IFNα. Higher doses have no effect in 2H11 murine endothelial cells and reduced effect on aortic outgrowth. Further studies are underway in our laboratory to demonstrate the factors responsible for this effect.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2330. doi:1538-7445.AM2012-2330