Quantitative Assessment of Angiogenesis in Murine Antigen-Induced Arthritis by Intravital Fluorescence Microscopy

Abstract

Inhibition of angiogenesis might be a therapeutic approach to prevent joint destruction caused by the overgrowing synovial tissue during chronic joint inflammation. The aim of this study was to investigate angiogenesis in the knee joint of mice with antigen-induced arthritis (AIA) by means of intravital microscopy. In 14 mice (C57BL6/129Sv) intravital microscopic assessment was performed on day 8 after AIA induction in two groups (controls, AIA). Synovial tissue was investigated by intravital fluorescence microscopy using FITC-dextran (150 kD). Quantitative assessment of vessel density was performed according to the following categories: functional capillary density (FCD, vessels <10 µm in diameter), functional vessel density (FVD, vessels >10 µm) and FVD of vessels with angiogenic criteria (convoluted vessels, abrupt changes of diameter, vessels which are generated by sprouting and progressively pruned and remodelled). Microvessel count was performed using immunohistochemistry. There was no significant difference in FCD between the control group (337 ± 9 cm/cm²; mean ± SEM) and the AIA group (359 ± 13 cm/cm²). The density of vessels larger than 10 µm diameter was significantly increased in animals with AIA (135 ± 10 vs. 61 ± 5 cm/cm² in control). The density of blood vessels with angiogenic criteria was enhanced in arthritic animals (79 ± 17 vs. 12 ± 2 cm/cm² in control). There was a significant increase in the microvessel count in arthritic animals (297 ± 25 vs. 133 ± 16 mm^–2 in control). These findings demonstrate that angiogenesis in murine AIA can be assessed quantitatively using intravital microscopy. Further studies will address antiangiogenic strategies in AIA.