By generating and analyzing a T-cadherin-deficient mouse model of mammary cancer, we have revealed an unprecedented role for T-cadherin in tumor angiogenesis. T-cadherin is expressed on epithelial, myoepithelial and some endothelial cells in virgin mouse mammary tissue, and down-regulated from epithelial cells during development of luminal epithelial tumors. Thus, T-cadherin becomes progressively confined to the vasculature during tumorigenesis. The restricted vascular expression of T-cadherin in the MMTV-PyV-mT model replicates our observations in MMTV-Neu induced mouse tumors (19
) and is similar to the situation in human cancers, including those of the breast (8
). As T-cadherin expression is progressively lost from epithelial cells during mammary tumorigenesis, the T-cadherin-deficient PyV-mT model can only assess a limited range of changes and leaves open the role of T-cadherin in the tumor cells. We report that genetic inactivation of T-cadherin does not accelerate tumorigenesis or result in formation of spontaneous tumors as might be expected if T-cadherin acted as a bona fide tumor suppressor. Rather, the loss of T-cadherin delays MMTV-PyV-mT tumor formation, retards growth and increases metastases. We attribute this phenotype to a major function of T-cadherin in tumor angiogenesis. The T-cadherin−/−
MMTVPyV-mT tumors are significantly less vascularized than those from corresponding wild type mice. The null mice develop no life-threatening vascular defects during embryogenesis, but show specific impairments in pathological neovascularization of mammary tumors and hypoxia-induced blood vessel remodeling of the retina. Reduced endothelial cell density and increased hypoxia in T-cadherin-deficient MMTV-PyV-mT tumors together with the observation that T-cadherin in the host microenvironment is needed for supporting growth of MMTV-PyV-mT tumors after transplantation, suggest an important role for T-cadherin in pathological neovascularization. We correlate the association of adiponectin with the tumor vasculature with T-cadherin's pro-angiogenic role and suggest a link between T-cadherin's vascular expression, adiponectin binding capability and tumor neovascularization.
An unexpected outcome of the current study was that T-cadherin−/−
MMTV-PyV-mT tumors attain a more malignant pathology and metastatic potential even though overall tumor growth is limited. The metastatic rate of MMTV-PyV-mT mouse tumors is known to be influenced by the genetic background (25
), and is low in the C57Bl/6 strain used for the current study (See Supplemental Table 2
). As T-cadherin is progressively lost from the ductal epithelium during normal tumorigenesis, how can the ablation of T-cadherin affect tumor cell invasion and metastasis? Two explanations are possible. First, the increased metastatic potential results from the disruption of T-cadherin-mediated adhesive interactions that prevents straying of neoplastic epithelial cells. This model assumes that low T-cadherin levels normally present on PyV-mT tumor cells are sufficient to restrain cells to the primary tumor mass and inhibit metastatic spreading. This suggestion would need to be tested in a gain-of-function genetic model. Alternatively, the limited blood supply of T-cadherin-deficient tumors increases hypoxia and changes in the tumor pathology (32
). Hypoxia is well known to be associated with a poor clinical outcome of invasive human breast carcinoma (34
), and T-cadherin-deficient tumors present with reduced blood vessel density, enhanced apoptosis, and enlarged hypoxic and necrotic regions. The metastases in T-cadherin−/−
MMTV-PyV-mT mice are not associated with endothelial cells and thus may represent section for an epithelial-meschenymal type transition.
T-cadherin has at least two activities that may influence angiogenesis. First, T-cadherin confers homophilic binding between cells (7
), and this engagement is reported to decrease adhesion, enhance migration and induce proliferation of endothelial cells (16
). Moreover, ectopic T-cadherin expression in the capillary microenvironment in vivo
repulses blood vessels and stops their growth (35
). Thus, inactivation of T-cadherin in vivo
might be expected to increase vascularization. However, the data presented here from the mouse null model do not support a restrictive role for T-cadherin in blood vessel growth in vivo
. Rather, the T-cadherin null mice show limited angiogenic responses suggesting a function for T-cadherin in supporting angiogenesis. Our work thus leaves open if the repulsive, homophilic binding function of T-cadherin plays into the complex interactions during angiogenesis in vivo
. Secondly, T-cadherin is a binding protein for the hexameric and HMW forms of adiponectin (21
), the predominant active forms in serum (36
). We find that adiponectin is sequestered to the vasculature in a T-cadherin-dependent manner and levels are dramatically increased in serum. Thus, T-cadherin may serve as a major adiponectin repository. The functions of adiponectin in the vasculature remain controversial, both positive and negative actions on blood vessel growth are reported (30
). Linking T-cadherin and adioponectin functions at a mechanistic level is thus a primary research task. Because of its membrane attachment through a glycosylphosphatidyl inositol moiety, T-cadherin alone is insufficient to act as a receptor transducing signals elicited by adiponectin binding. Thus, we favour a model in which T-cadherin signals through associated molecules that perhaps provide a link with adiponectin- or other vascular receptors. One conceivable role for T-cadherin as an adiponectin binding protein includes sequestering adiponectin-associated growth factors, such as PDGF-BB, bFGF, HB-EGF and Trombospondin-1 (37
). These factors exert important roles in establishing functional and stable vascular networks (39
), and the T-cadherin-dependent accumulation of adiponectin may regulate their availability upon signals eliciting vascular responses. Irrespective of the mechanism, the work reported here establishes that T-cadherin regulates retinal and tumor angiogenesis and is responsible for sequestering adiponectin to the vasculature. These studies thus open new avenues for unravelling the molecular complexity of the vascular response under challenging physiological conditions.