Background: Postmenopausal women taking hormone replacement therapy (HRT) require secondary intervention after vascular reconstruction more frequently than women not taking HRT, often due to increased development of intimal hyperplasia (IH). Matrix metalloproteinases (MMPs) play a role in IH by degradation and remodeling of components of the vascular basement membrane. The MMP pathway is regulated by a balance between MMPs, membrane-type MMPs (MT-MMPs), and tissue inhibitor of MMPs (TIMPs). We have recently provided evidence for unbalanced regulation of the MT1-MMP/MMP-2 pathway in vascular smooth muscle cells (VSMCs) exposed to hormones in vitro. Herein we study the role of HRT in the development of IH in a postmenopausal rodent model of vascular injury and in the modulation of this MMP regulatory pathway in vivo.Methods: Female rats were aged to 12 months. Animals were ovariectomized (OVX) and 4 weeks later hormones or placebo was delivered via a 90-day slow-release pellet. After 6 weeks of HRT each rat underwent balloon angioplasty of the left common carotid artery. At 14 days postinjury tissue samples were collected and stained with trichrome elastin and for isoform-specific MMPs.Results: After vascular injury, the intima:media (I:M) ratio was decreased in OVX rats receiving placebos as compared with non-OVX controls (P < 0.05). In OVX animals receiving HRT, estrogen with and without progesterone and progesterone alone slightly increased I:M ratio compared with placebo, although no significant difference was found in any HRT group. Injury-induced intimal expression of MMP-2 and -9 was decreased in OVX placebo animals compared with non-OVX controls (P < 0.05). MMP-2 and -9 levels were subsequently increased by each type of hormone therapy compared with placebo, with a significant increase in MMP-9 in response to estrogen with and without progesterone (P < 0.05). Conversely, TIMP-2 was decreased by estrogen compared with placebo (P < 0.05). There was no effect on intimal MT1-MMP in any group.Conclusions: In this study we detected a statistically significant decrease in IH as a result of OVX. Subsequent HRT exposure resulted in increased I:M ratios compared with OVX animals given placebo, although significance was not reached with the doses given. Long-term exogenous exposure may have a more deleterious effect compared with acute exposure and should be examined further. We also demonstrated a significant reduction in MMP-2 and -9 and TIMP-2 in response to OVX. Subsequent hormone exposure resulted in the upregulation of MMP-2 and -9 without a counterregulatory increase in TIMP, indicating that HRT modulates the MMP regulatory pathway in vivo. The data suggest that the lack of hormones after OVX protects against pathologic remodeling in our aged model of disease and that exposure to both natural and exogenous hormones could be a negative risk factor resulting in an exaggerated vascular response to injury. Future studies should focus on in vivo manipulation of unbalanced MMP regulation for prevention of IH in response to HRT and in general. Furthermore, the age-associated difference in response to the presence of natural hormones in young vs aged models should be investigated.

Postmenopausal women taking hormone replacement therapy (HRT) have increased intimal hyperplasia (IH) following vascular intervention. Matrix metalloproteinases (MMPs) play a major role in IH development, and we have shown that hormone exposure results in unbalanced MMP regulation in vascular smooth muscle cells in vitro. Previously we presented data from a small pilot study suggesting a role for HRT in the development of IH via MMP modulation in vivo, using a postmenopausal rodent model of vascular injury. Here we further investigated the role of HRT as a modulator of MMPs and IH in a larger follow-up study. Female rats were aged to 12 months and ovariectomized (OVX). Four weeks later estrogen (E), progesterone (P), estrogen + progesterone (E/P), or placebo (Plac) were delivered via 90-day slow-release pellets. Following 6 weeks of HRT, each rat underwent balloon angioplasty of the left common carotid artery. At 14 days postinjury, tissue samples were collected and stained with Trichrome elastin for intima:media (I:M) measurement and stained for various MMP isoforms. Following vascular injury I:M was decreased in OVX rats compared to non-OVX controls (Fig 1, n = 5-6, P = NS). In OVX animals, HRT exposure did not significantly increase the I:M ratios. (Fig 1, n = 5-6, P = NS). Injury-induced expression of MMP-2 and −9 was significantly decreased in OVX animals compared to non-OVX controls (Table 1; n = 5-6). MMP-2 and −9 levels were subsequently increased by each type of hormone therapy compared to placebo, with a significant increase in MMP-9 in response to estrogen with and without progesterone (Table; n = 5-6). Conversely, TIMP-2 was significantly decreased by estrogen compared to placebo (Table; n = 5-6). There was no effect on intimal MT1-MMP in any group.Table 1% intima stainedNonOVX COVX - PlacOVX - EOVX - POVX - EPMMP-237.8 ± 10.015.4 ± 3.7⁎P < 0.05 vs. NonOVX C26.5 ± 3.738.8 ± 7.638.5 ± 7.6MMP-94.3 ± 0.12.7 ± 0.4⁎P < 0.05 vs. NonOVX C4.7 ± 0.6#P < 0.05 vs. OVX-Plac6.0 ± 1.37.1 ± 1.0#P < 0.05 vs. OVX-PlacTIMP-250.9 ± 8.922.2 ± 3.4⁎P < 0.05 vs. NonOVX C8.7 ± 1.4#P < 0.05 vs. OVX-Plac24.8 ± 10.128.2 ± 9.4 P < 0.05 vs. NonOVX C# P < 0.05 vs. OVX-Plac Open table in a new tab Here we were not able to demonstrate a statistically significant decrease in IH as a result of ovariectomy. Furthermore, HRT at the doses given did not remarkably increase IH. Here we also demonstrate a significant reduction in MMP-2, −9, and TIMP-2 in response to ovariectomy. Subsequent hormone exposure results in the upregulation of MMP-2 and −9 without a counter-regulatory increase in TIMP. We have previously shown elevation in MT1-MMP to occur during the initial phases of IH development; therefore, examination earlier than 14 days postinjury is needed to determine the effect of HRT on this MMP regulatory isoform. Future studies should investigate in vivo manipulation of this unbalanced MMP regulation for prevention of IH in response to HRT exposure.