Tumor growth is angiogenesis dependent. We hypothesized that nonneoplastic tissue growth also depends on neovascularization. We chose adipose tissue as an experimental system because of its remodeling capacity. Mice from different obesity models received anti-angiogenic agents. Treatment resulted in dose-dependent, reversible weight reduction and adipose tissue loss. Marked vascular remodeling was evident in adipose tissue sections, which revealed decreased endothelial proliferation and increased apoptosis in treated mice compared with controls. Continuous treatment maintained mice near normal body weights for age without adverse effects. Metabolic adaptations in food intake, metabolic rate, and energy substrate utilization were associated with antiangiogenic weight loss. We conclude that adipose tissue mass is sensitive to angiogenesis inhibitors and can be regulated by its vasculature. Substantial evidence has established that tumor growth is angiogenesis-dependent (1). Neovascularization promotes tumor growth (2), whereas angiogenesis inhibition prevents it, and can regress the lesions (3). This raises the question of whether the endothelium functions similarly in nonneoplastic tissue growth (4).The challenges in addressing this question are that most adult tissues normally do not grow, their mass is stable, and the supporting vasculature is quiescent (5). Exceptionally, adipose tissue can grow and regress throughout adulthood. It is highly vascularized and has angiogenic properties (6, 7). An extensive capillary network surrounds each adipocyte, and there are few other resident cell types (8). Therefore, adipose tissue is uniquely suited to study the role of angiogenesis in nonneoplastic adult tissue growth.The potential for adipose tissue to grow and regress is substantial. One would expect the vasculature to have a commensurate capacity for remodeling. We hypothesized that adipose tissue growth is angiogenesis dependent and, therefore, may be inhibited by anti-angiogenesis agents. This would suggest that adipose tissue mass may be regulated via the vascular endothelium.We primarily used ob͞ob mice because they rapidly accumulate adipose tissue (9). This strain develops spontaneous obesity because of a lack of functional leptin, a protein secreted by adipocytes that acts on the hypothalamus to regulate appetite and metabolism (9-11). Without leptin, animals eat excessively, expend less energy, and become morbidly obese. Leptin replacement induces weight loss in ob͞ob mice by specifically reducing adipose tissue (9), unlike diet restriction or appetite suppression (fenfluramine), which decreases both lean and fat mass (12, 13).We treated ob͞ob mice with various angiogenesis inhibitors. We primarily used TNP-470 (14) because of availability. TNP-470 inhibits endothelial cell proliferation in vitro (15) and angiogenesis in vivo (16). At significantly higher concentrations, TNP-470 can also suppress nonendothelial cell proliferation (14,17). Angiostatin (kringle 1-4 domains of plasminogen) (18) and endostatin (...

Tumor angiogenesis is necessary for solid tumor progression and metastasis. Tumor blood vessels have been shown to differ from normal counterparts, for example, by changes in morphology. An important concept in tumor angiogenesis is that tumor endothelial cells are assumed to be genetically normal, although these endothelial cells are structurally and functionally abnormal. However, we hypothesized that given the phenotypic differences between tumor and normal blood vessels, there may be genotypic alterations as well. Mouse endothelial cells were isolated from two different human tumor xenografts, melanoma and liposarcoma, and from two normal endothelial cell counterparts, skin and adipose. Tumor-associated endothelial cells expressed typical endothelial cell markers, such as CD31. They had relatively large, heterogeneous nuclei. Unexpectedly, tumor endothelial cells were cytogenetically abnormal. Fluorescence in situ hybridization (FISH) analysis showed that freshly isolated uncultured tumor endothelial cells were aneuploid and had abnormal multiple centrosomes. The degree of aneuploidy was exacerbated by passage in culture. Multicolor FISH indicated that the structural chromosomal aberrations in tumor endothelial cells were heterogeneous, indicating that the cytogenetic alterations were not clonal. There was no evidence of human tumor-derived chromosomal material in the mouse tumor endothelial cells. In marked contrast, freshly isolated normal skin and adipose endothelial cells were diploid, had normal centrosomes, and remained cytogenetically stable in culture even up to 20 passages. FISH analysis of tumor sections also showed endothelial cell aneuploidy. We conclude that tumor endothelial cells can acquire cytogenetic abnormalities while in the tumor microenvironment.

Dysfunctional endothelium contributes to more disease than any other tissue in the body. Small interfering RNAs (siRNAs) have the potential to help study and treat endothelial cells in vivo by durably silencing multiple genes simultaneously, but efficient siRNA delivery has so far remained challenging. Here we show that polymeric nanoparticles made of low molecular weight polyamines and lipids can deliver siRNA to endothelial cells with high efficiency, thereby facilitating the simultaneous silencing of multiple endothelial genes in vivo. Unlike lipid or lipid-like nanoparticles, this formulation does not significantly reduce gene expression in hepatocytes or immune cells even at the dosage necessary for endothelial gene silencing. It mediates the most durable non-liver silencing reported to date, and facilitates the delivery of siRNAs that modify endothelial function in mouse models of vascular permeability, emphysema, primary tumour growth, and metastasis. We believe these nanoparticles improve the ability to study endothelial gene function in vivo, and may be used to treat diseases caused by vascular dysfunction.

Epoxyeicosatrienoic acids (EETs) are small molecules produced by cytochrome P450 epoxygenases. They are lipid mediators that act as autocrine or paracrine factors to regulate inflammation and vascular tone. As a result, drugs that raise EET levels are in clinical trials for the treatment of hypertension and many other diseases. However, despite their pleiotropic effects on cells, little is known about the role of these epoxyeicosanoids in cancer. Here, using genetic and pharmacological manipulation of endogenous EET levels, we demonstrate that EETs are critical for primary tumor growth and metastasis in a variety of mouse models of cancer. Remarkably, we found that EETs stimulated extensive multiorgan metastasis and escape from tumor dormancy in several tumor models. This systemic metastasis was not caused by excessive primary tumor growth but depended on endothelium-derived EETs at the site of metastasis. Administration of synthetic EETs recapitulated these results, while EET antagonists suppressed tumor growth and metastasis, demonstrating in vivo that pharmacological modulation of EETs can affect cancer growth. Furthermore, inhibitors of soluble epoxide hydrolase (sEH), the enzyme that metabolizes EETs, elevated endogenous EET levels and promoted primary tumor growth and metastasis. Thus, our data indicate a central role for EETs in tumorigenesis, offering a mechanistic link between lipid signaling and cancer and emphasizing the critical importance of considering possible effects of EET-modulating drugs on cancer.