TGF-beta signaling promotes tumor vasculature by enhancing the pericyte-endothelium association

Background: The breast cancer microenvironment promotes tumor vascularization through the complex interactions involving tumor-associated fibroblasts (TAFs). Emerging data indicate that TAFs increase production and signaling by TGF-beta cytokines, while the role of TGF-beta signaling in the regulation of tumor blood vessels is not fully understood. The current study presents evidence that TAFs enhance the organization of tumor blood capillaries, and TGF-beta signaling plays an important role in this response. Methods: Tumor vascularization was studied in xenograft models of breast carcinoma cells, alone and in combination with fibroblasts. TGF-beta signaling in breast cancer cells was modulated by expression of kinase-inactive TGFBR1-K232R (dnTGFBR1) or constitutive-active TGFBR1-T204D (caTGFBR1) receptor mutants. The architecture of tumor blood capillaries was assessed by immune-histochemical analysis of endothelium and pericytes. The role of TGF-beta-Smad signaling in fibronectin expression was examined using adenoviral transduction of signaling components. Results: Our studies revealed that TAFs significantly increase the lumen size of blood microvessels. Inactivation of TGF-beta signaling in tumor cells by dnTGFBR1 reduced the microvessel density and lumen sizes, decreasing tumor growth. In contrast, caTGFBR1-tumors exhibited greater vessel density and lumen sizes. Tumors with inactive dnTGFBR1 showed lower amounts of TAFs, while caTGFBR1 increased amounts of TAFs compared to the control. Inspection of pericytes and endothelial cells in tumor vasculature revealed that TAFs enhanced vessel coverage by pericytes, vascular cells supporting capillaries. This effect was impaired in dnTGFBR1-tumors, whereas active caTGFBR1 enhanced the association of pericytes with endothelium. Accordingly, dnTGFBR1-tumors exhibited the presence of hemorrhages, a sign of fragile blood vessels. Biochemical analysis showed that TGFBR1-SMAD signaling up-regulates fibronectin, a prominent regulator of endothelium-pericyte interactions. Conclusions: The current study indicates that tumor-fibroblast crosstalk enhances tumor vascularization by increasing the pericyte-endothelium association via a mechanism involving the TGF beta-fibronectin axis. The tumor-fibroblast model represents a useful system for dissecting the complex interactions governing tumor angiogenesis and developing new approaches to therapeutic targeting tumor vasculature.