T-cadherin regulates vascular smooth muscle cell contractile signalling and promotes acquisition of the de-differentiated phenotype

Cadherins are a superfamily of intercellular adhesion molecules essential for structural maintenance of tissue cohesion, precise primary tissue segregation and regulation of regeneration processes in adult. Cadherins are widely expressed in the vasculature. Adherens junctions and desmosomes, where cadherins are the intercellular adhesion transmembrane linkers, have been demonstrated in large and small arteries in vivo and their participation in correct organization of vascular smooth muscle architecture is doubtless. However, knowledge on precise functional roles for cadherin in healthy or diseased vascular smooth muscle is limited.
T-cadherin is an atypical cadherin highly expressed on endothelial and smooth muscle layers of the vasculature. Dynamic T-cadherin expression on vascular smooth muscle in vivo has been reported in number of vascular pathologies including two major vasoproliferative disorders – atherosclerosis and restenosis. Functions and molecular mechanisms regulated by this molecule in the smooth muscle cell component of the vasculature are unknown. The primary functions of vascular smooth muscle cells (VSMC) are contraction and regulation of blood vessel tone. However, VSMC possess inherent plasticity: they can switch from mature contractile phenotype to a de-differentiated proliferative and synthetic phenotype in response to vascular injury, or local environmental cues signalling. Studies in this dissertation are aimed at establishing cellular functions for T-cadherin in VSMC contraction and phenotype plasticity and identifying mediating molecular mechanisms.
First, we found that T-cadherin modulates non-metabolic insulin signalling via Akt/mTOR, which in turn leads to alterations in VSMC contractile competence and increased matrix remodelling. T-cadherin overexpressing cells exhibited elevated constitutive levels of phosphorylated Aktser473, GSK3βser9, S6RPser235/236 and IRS-1ser636/639. Contractile machinery was constitutively altered in a manner indicative of reduced intrinsic contractile competence, namely decreased phosphorylation of MYPT1thr696 or MYPT1thr853 and MLC20thr18/ser19, reduced RhoA activity and increased iNOS expression. T-cadherin overexpressing VSMC-populated collagen lattices exhibited greater compaction which was due to increased collagen fibril packing/reorganization. These cells also exhibited a state of insulin insensitivity as evidenced by attenuation of the ability of insulin to stimulate Akt/mTOR axis signalling, phosphorylation of MLC20 and MYPT1, compaction of free-floating lattices and collagen fibril reorganization in unreleased lattices.
Second, T-cadherin upregulation on VSMC, a phenomenon observed in VSMC-driven vascular pathologies (atherosclerosis and restenosis) promotes VSMC phenotype transition. T-cadherin upregulation in VSMC caused loss of spindle morphology, reduced/disorganized stress fibre formation, decay of SMC-differentiation marker proteins, increased levels of β-catenin and cyclin D1, and migro-proliferative behaviour. Genetic T-cadherin ablation, on the other hand, enforced differentiated phenotype. T-cadherin hyperactivates Akt axis signalling and inactivates classical downstream effector GSK3β. Ectopic adenoviral-mediated co-expression of constitutively active GSK3β restored morphological, molecular, and functional characteristics of differentiated VSMC in T-cadherin overexpressing cells, suggesting that GSK3β inactivation is essential for T-cadherin induced VSMC de-differentiation.
The studies have revealed novel cadherin-based modalities to regulate VSMC sensitivity to insulin and phenotype plasticity, which is achieved via Akt/mTOR axis hyperactivation and altered downstream effector signalling.