Background Matrix metalloproteinases (MMPs) are implicated in the pathogenesis of varicose veins. We have shown that MMP-2 causes relaxation of venous segments and suggested a role of venous smooth muscle (VSM) hyperpolarization; however, the downstream mechanisms are unclear. We tested whether MMP-2 induced venous relaxation involves inhibition of the Ca2+ mobilization mechanisms of VSM contraction due to generation of Arg-Gly-Asp (RGD)-containing peptides. Methods Circular segments of inferior vena cava (IVC) were isolated from male Sprague-Dawley rats, suspended between two wires in a tissue bath, and isometric contraction was measured. Contraction data in mg/mg tissue were presented as means±SEM. Results In IVC incubated in normal Krebs (2.5 mM Ca2+), the α-adrenergic agonist phenylephrine (Phe, 10-5 M) caused initial peak (133.2±17.5) followed by a maintained contraction (73.4±11.6), that was inhibited by MMP-2 (1 μg/mL) to 32.4±12.8 in 30 min. The inhibitory effects of MMP-2 were reversible by washing the tissue with Krebs or in the presence of the MMP inhibitors TIMP-1 (1 μg/ml), Ro 28-2653 and BB-94 (10-6 M), and were not associated with changes in IVC structure, demonstrating specificity. Angiotensin II (AngII, 10-6 M) caused a monophasic contraction (114.2±12.2), that was also inhibited by MMP-2 (66.0±7.4), suggesting a post-receptor effect on the downstream mechanisms of VSM contraction. To test the role of Ca2+ release from the sarcoplasmic reticulum, IVC was incubated in Ca2+-free (2 mM EGTA) Krebs with or without MMP-2. In Ca2+-free Krebs, caffeine did not cause contraction, suggesting limited role of the Ca2+-induced Ca2+-release mechanism, and Phe and AngII caused a small contraction (7.2±1.7 and 14.9±2.8) that was slightly increased by MMP-2 (10.4±3.0 and 33.8±10.0), suggesting little effect on IP3-induced Ca2+ release. To test the role of Ca2+ entry through membrane channels, after eliciting a transient Phe contraction in nominally 0 Ca2+ Krebs, increasing concentrations of CaCl2 (0.1, 0.3, 0.6, 1, 2.5 mM) were added and the [Ca2+]e-contraction relationship was constructed. The [Ca2+]e-contraction relation was reduced in MMP-2 treated IVC, suggesting inhibition of Ca2+ entry. In IVC treated with MMP-2, the Ca2+ channel blocker diltiazem (10-5M) did not cause any further inhibition of Phe contraction, suggesting that Ca2+ entry is already inhibited by MMP-2. To test whether MMP-2 actions involve generation of RGD and modulation of integrin receptors, experiments where repeated in IVC segments saturated with RGD (10-5 M), or pretreated with the αvβ3 integrin blocker cyclo-RGD. RGD-peptide caused only small relaxation of Phe contracted IVC (6.4±3.4%), and addition of MMP-2 to RGD-treated IVC caused further relaxation (69.7±3.0%). Pretreatement of IVC with cyclo-RGD did not significantly affect MMP-2 induced relaxation (55.0±5.0%). Conclusions In rat IVC, MMP-2 attenuates [Ca2+]e-dependent VSM contraction, without affecting Ca2+ release from intracellular Ca2+ stores. MMP-2 induced VSM relax...

Cardiovascular disease (CVD) is more common in men and postmenopausal women than premenopausal women, suggesting vascular benefits of female sex hormones. Studies on the vasculature have identified estrogen receptors ERα, ERβ and a novel estrogen binding membrane protein GPR30, that mediate genomic and/or non-genomic effects. Estrogen promotes endothelium-dependent relaxation by inducing the production/activity of nitric oxide, prostacyclin, and hyperpolarizing factor, and inhibits the mechanisms of vascular smooth muscle contraction including [Ca2+]i, protein kinase C, Rho kinase and mitogen-activated protein kinase. Additional effects of estrogen on the cytoskeleton, matrix metalloproteinases and inflammatory factors contribute to vascular remodeling. However, the experimental evidence did not translate into vascular benefits of menopausal hormone therapy (MHT), and the HERS, HERS-II and WHI clinical trials demonstrated adverse cardiovascular events. The discrepancy has been partly related to delayed MHT and potential changes in the vascular ER amount, integrity, affinity, and downstream signaling pathways due to the subjects' age and preexisting CVD. The adverse vascular effects of MHT also highlighted the need of specific modulators of vascular sex hormone receptors. The effectiveness of MHT can be improved by delineating the differences in phramcokinetics and pharmacodynamics of natural, synthetic, and conjugated equine estrogens. Estriol, “hormone bioidenticals” and phytoestrogens are potential estradiol substitutes. The benefits of low dose MHT, and transdermal or vaginal estrogens over oral preparations are being evaluated. Specific ER modulators (SERMs) and ER agonists are being developed to maximize the effects on vascular ERs. Also, the effects of estrogen are being examined in the context of the whole body hormonal environment and the levels of progesterone and androgens. Thus, the experimental vascular benefits of estrogen can be translated to the outcome of MHT in postmenopausal CVD, as more specific modulators of sex hormone receptors become available and are used at the right dose, route of administration and timing, depending on the subject's age and preexisting cardiovascular condition.