Adipose angiogenesis: quantitative methods to study microvessel growth, regression and remodeling in vivo.

Genetic and diet-induced rodent obesity models provide outstanding opportunities to study the role of angiogenesis and vascular remodeling in modulation of adipogenesis and obesity. In this study, we describe methods to quantitatively study adipose angiogenesis and vascular remodeling on the basis of immunohistochemical analyses. Fresh white adipose tissue or brown adipose tissue are prepared for whole mount, cryosectioned and paraffin-embedded samples, followed by staining with specific markers such as platelet endothelial cell adhesion molecule-1 (PECAM-1)/CD31, CD34, isolectin B4 or alpha-smooth muscle actin. Adipocytes are visualized by staining lipid droplets with 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene-3-dodecanoic acid (BODIPY) 558/568 C(12). This protocol may take 2-5 d to obtain results. In the view of the crucial roles of vasculature in modulation of adipogenesis and obesity, this protocol is valuable for studying the molecular mechanisms of angiogenesis in obese adipose tissues and for assessing the anti-obesity activity of angiogenesis modulators.