Dioxin Receptor Deficiency Impairs Angiogenesis by a Mechanism Involving VEGF-A Depletion in the Endothelium and Transforming Growth Factor-β Overexpression in the Stroma*

Angiogenesis has key roles in development and in the progression of human diseases such as cancer. Consequently, identifying the novel markers and regulators of angiogenesis is a critical task. The dioxin receptor (AhR) contributes to vascular homeostasis and to the endothelial response to toxins, although the mechanisms involved are largely uncharacterized. Here, we show that AhR-null mice (AhR−/−) have impaired angiogenesis in vivo that compromises tumor xenograft growth. Aortic rings emigration experiments and RNA interference indicated that AhR−/− endothelial cells failed to branch and to form tube-like structures. Such a phenotype was found to be vascular endothelial growth factor (VEGF)-dependent, as AhR−/− aortic endothelial cells (MAECs) secreted lower amounts of active VEGF-A and their treatment with VEGF-A rescued angiogenesis in culture and in vivo. Further, the addition of anti-VEGF antibody to AhR+/+ MAECs reduced angiogenesis. Treatment under hypoxic conditions with 2-methoxyestradiol suggested that HIF-1α modulates endothelial VEGF expression in an AhR-dependent manner. Importantly, AhR-null stromal myofibroblasts produced increased transforming growth factor-β (TGFβ) activity, which inhibited angiogenesis in human endothelial cells (HMECs) and AhR−/− mice, whereas the co-culture of HMECs with AhR−/− myofibroblasts or with their conditioned medium inhibited branching, which was restored by an anti-TGFβ antibody. Moreover, VEGF and TGFβ activities cooperated in modulating angiogenesis, as the addition of TGFβ to AhR−/− MAECs further reduced their low basal VEGF-A activity. Thus, AhR modulates angiogenesis through a mechanism requiring VEGF activation in the endothelium and TGFβ inactivation in the stroma. These data highlight the role of AhR in cardiovascular homeostasis and suggest that this receptor can be a novel regulator of angiogenesis during tumor development.