Menopause Effects on Vascular Function

Study Overview
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Brief Summary
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The purpose of the study is to identify the independent effect of estradiol (E2) on endothelin-1 (ET-1) mediated vasomotor function in women. The study is the first step in recognizing the impact of ovarian hormones on the mechanisms that regulate vascular function in women to provide a better understanding of the cardiovascular efficacy of hormone therapy.

Detailed Description
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Cardiovascular disease (CVD) is the leading cause of death in women (Roger, Go, Lloyd, Adams, Berry, Brown, et al, 2011). Functional changes in the microvasculature occur with aging and precede atherosclerosis, contributing to CVD (Seals, Jablonski, & Donato, 2011). Furthermore, because of the decline in ovarian hormones during menopause, age-related impairments in endothelial function are exacerbated in postmenopausal women (PMW). However, the safety and efficacy of currently available hormone-based therapies remains controversial (Devi, Sugiguchi, Pederson, Abrassart Glodowski, & Nachtigall, 2013: Miller, Black, Brinton, Budoff, Cedars, Hodis, et al, 2009). Endothelin-1 (ET-1) is a potent vasoconstrictor produced and released by endothelial cells and implicated in the development of atherosclerosis (Best, McKenna, Holmes, & Lerman, 1999; Donato, Gano, Eskurza, Silver, Gates, Jablonski, et al, 2009; Ihling, Szombathy, Bohrmann, Brockhaus, Schaefer, & Loeffler, 2009). ET-1 binds to two receptor subtypes, ET-A and ET-B (Yanagisawa, Kurihara, Kimura, Tomobe, Kobayashi, Mitsui, et al, 1988). While both receptors are located on vascular smooth muscle (VSM) and cause vasoconstriction, ET-B receptors are also located on the endothelium and cause vasodilation (Gomez-Sanchez, Cozza, Foecking, Chiou, & Ferris, 1990; Haynes, 1995; Ishikawa, Ihara, Noguchi, Mase, Mino Saeki, et al, 1994). In women, ET-1 preferentially binds to ET-B receptors compared to ET-A receptors, supporting findings of sex differences in ET-1 receptor responses and suggesting ET-B receptors are under hormonal control (Ergul, Shoemaker, Puett, & Tackett, 1998; Kellogg, Liu, & Pergola, 2001; Stauffer, Westby, Greiner, Van Guilder, & Desouza, 2010). In animal models, estradiol (E2) reduces ET-1 mediated vasoconstriction and increases ET-B receptor mRNA (Pederson, Nielsen, Mortensen, Nilas, & Ottesen, 2008). Thus, low levels of E2 in PMW may contribute to impaired vascular function through an ET-B receptor mechanism. However, the interaction between E2 and ET-1 receptor responses on regulating vascular function in women is currently unknown. The long-term goal of the laboratory is to understand the impact of ovarian hormones on the mechanisms that regulate vascular function in women to provide a better understanding of the cardiovascular efficacy of hormone therapy. The study is the first step in reaching our goal; the objective of the study is to identify the independent effect of E2 on ET-1 mediated vasomotor function in women. The investigators will measure blood flow responses to local heating in the cutaneous circulation during perfusion of ET-1 receptor antagonists via microdialysis, coupled with measures of intracellular protein and receptor expression on endothelial cells and skin punch biopsies (to assess VSM cells) collected from young and PMW while controlling ovarian hormone exposure. Young women will be tested after suppressing ovarian production of E2 and progesterone with a gonadotropin-releasing hormone antagonist (GnRHant), and again after E2 administration; PMW, who are not using hormone therapy, will be tested before and after E2 admin. The central hypothesis is that declines in E2 impair microvascular vasodilatory function due to cellular changes in ET-B receptor expression on endothelial and VSM cells, and that E2 administration reverses these responses.

Official Title
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Mechanisms of Vascular Dysfunction in Women: Role of Estradiol

Conditions
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Cardiovascular Risk Factor

Intervention / Treatment
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* Other: No to Low Endogenous Estrogen
* Other: Estradiol


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Young women between 18-35 years of age with regular menstrual cycles Postmenopausal women between 50-65 years of age and no more than 10 years past menopause Non-smoking BMI < 30 kg/m2 Free from known disease (heart disease, cancer, diabetes) Exclusion Criteria: Current use of hormone therapy or within the past year Women using Depo-provera or an intra-uterine device (IUD) Pregnant, are planning on becoming pregnant, or are breast- feeding. History of stable or unstable angina Diabetes Neurological disease Lung disease Kidney or liver disease Cancer Hysterectomy Peripheral vascular disease History of blood clots Heart disease Fibroids High blood pressure Stroke

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 65 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Basic Science
Allocation: Non-Randomized
Intervention Model: Parallel Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: No to Low Endogenous Estrogen<br>PMW and young women (YW) will receive medical study clearance after a detailed physical examination. YW will self-administer subcutaneous injections of the gonadotropin-releasing hormone (GnRH) antagonist, ganirelix acetate (Antagon, 0.25 mg/day in 0.5 ml of normal saline, Organon, Inc., West Orange, New Jersey,) daily to suppress endogenous ovarian hormone production (16, 17, 18). This will begin following a separate medical screening at Reproductive Associates of Delaware 48 hours prior to initiating the hormone intervention to rule out other contraindications prior to beginning the treatment. YW will begin using the antagonist on days 26-28 of their menstrual cycle, and continue daily for 10-12 days. The experimental protocol will be conducted in YW after 3-4 days of using the GnRH antagonist. PMW will complete the experimental protocol prior to use of the 17β-estradiol (E2, 0.1 mg/day patch, Vivelle dot; estradiol patch). | Other: No to Low Endogenous Estrogen<br>* Ganirelix acetate (Antagon) will be used to prevent endogenous production of ovarian hormones in young women. Ganirelix is derived from native GnRH, and acts by competitively blocking GnRH receptors on the pituitary and subsequent pathways. Thus, administration of the GnRH antagonist (GnRHant) suppresses steroidogenesis, leading to low or undetectable serum estrogen and progesterone concentrations, which occurs within two days of initiation of administration (Oberye, Mannaerts, Huisman & Timmer, 1999; Oberye, Mannaerts, Kleijn, & Timmer, 1999).<br>|
| Experimental: Estrogen Add-Back<br>Estradiol (E2, 0.1 mg/day patch, Vivelle dot; estradiol patch) will be administered for 7 days to both young and PMW. Young women will use the E2 over the last 7 days of Antagon administration. | Other: No to Low Endogenous Estrogen<br>* Ganirelix acetate (Antagon) will be used to prevent endogenous production of ovarian hormones in young women. Ganirelix is derived from native GnRH, and acts by competitively blocking GnRH receptors on the pituitary and subsequent pathways. Thus, administration of the GnRH antagonist (GnRHant) suppresses steroidogenesis, leading to low or undetectable serum estrogen and progesterone concentrations, which occurs within two days of initiation of administration (Oberye, Mannaerts, Huisman & Timmer, 1999; Oberye, Mannaerts, Kleijn, & Timmer, 1999).<br>Other: Estradiol<br>* Short term estradiol administration elicits changes in vascular function in women, and 0.1mg/day patch is the upper recommended limit for hormone therapy in women (Wenner, Taylor, & Stachenfeld, 2011; Moreau, Hildreth, Meditz, Deane & Kohrt, 2012).<br>* Other names: Vivelle dot;|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Vascular endothelial function | The capacity of the small and large blood vessels to dilate. | 3 years | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Endothelin receptor expression | Venous endothelial cells and skin punch biopsy will be collected and stained for ET-A and ET-B receptor expression | 3 years |