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Impact of micronised progesterone and medroxyprogesterone acetate in combination with transdermal oestradiol on cardiovascular markers in women diagnosed with premature ovarian insufficiency or an early menopause: a randomised pilot trial

  • Author Footnotes
    # INSTITUTION WHERE THE WORK WAS UNDERTAKEN: King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS
    Monica Mittal
    Correspondence
    Corresponding author. Imperial College Healthcare NHS Trust, St Mary's and Hammersmith Hospitals, Praed Street, Paddington, London W2 1NY, UK
    Footnotes
    # INSTITUTION WHERE THE WORK WAS UNDERTAKEN: King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS
    Affiliations
    Imperial College Healthcare NHS Trust, St Mary's and Hammersmith Hospitals, Department of Obstetrics and Gynaecology, Praed Street, London W2 1NY
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  • Carmel McEniery
    Affiliations
    University of Cambridge, Division of Cardiovascular Medicine, Addenbrooke's Hospital, Cambridge, Hills Road, Cambridge CB2 0QQ
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  • Prasanna Raj Supramaniam
    Affiliations
    Oxford University Hospitals NHS Foundation Trust, John Radcliffe Hospital, Department of Obstetrics and Gynaecology, Headley Way, Headington, Oxford OX3 9DU
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  • Linda Cardozo
    Affiliations
    King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS
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  • Mike Savvas
    Affiliations
    King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS
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  • Nick Panay
    Affiliations
    Imperial College Healthcare NHS Trust and Chelsea and Westminster NHS Foundation Trust, Queen Charlotte's and Chelsea Hospital, Department of Obstetrics and Gynaecology, Du Cane Rd, White City, London W12 0HS
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  • Haitham Hamoda
    Affiliations
    King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS
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  • Author Footnotes
    # INSTITUTION WHERE THE WORK WAS UNDERTAKEN: King's College Hospital NHS Foundation Trust, Department of Obstetrics and Gynaecology, Denmark Hill, Brixton, London SE5 9RS

      Highlights

      • Risk stratification of arterial disease can be performed with carotid femoral pulse wave velocity.
      • Menopause augments the age-dependent increase in arterial stiffness.
      • Hormone replacement therapy is the mainstay of management of women diagnosed with an early menopause and premature ovarian insufficiency.
      • Micronised progesterone/medroxyprogesterone acetate can both be used in this cohort.
      • Positive changes in traditional markers with micronised progesterone were not reflected in the carotid femoral pulse wave velocity measurements.

      ABSTRACT

      Objective

      To compare the difference between micronised progesterone (MP) and medroxyprogesterone acetate (MPA) in combination with transdermal oestradiol (t-E2) on cardiovascular disease (CVD) risk markers in women diagnosed with an early menopause and premature ovarian insufficiency (EMPOI).

      Background

      The European Society for Cardiology has identified carotid femoral pulse wave velocity (cfPWV) as the gold standard cardiogenic biomarker for risk stratification of arterial disease. Menopause has been shown to augment the age-dependent increase in arterial stiffness, with hormone replacement therapy (HRT) being the mainstay of management of women diagnosed with EMPOI.

      Study design

      A pilot randomised prospective open-label trial. Women were randomised to either cyclical MP (Utrogestan® 200mg) or MPA (Provera® 10mg) in conjunction with t-E2 (Evorel® Patches 50mcg/day) for 12 months. Seventy-one subjects were screened, and baseline data are available for 57 subjects.

      Main outcome measure

      Carotid-femoral pulse wave velocity (cfPWV).

      Results

      PWV did not significantly change from baseline in either treatment arm. MP + t-E2 demonstrated a positive effect on traditional CVD markers, with a significant improvement seen in cardiac output (CO) (0.71±1.01mL/min, 95% CI 0.20 to 1.21) and reduction in diastolic blood pressure (DBP) (-3.43±6.31mmHg, 95% Cl -6.57 to -0.29) and total peripheral resistance (TPR) (-0.15±0.19mmHg⋅min⋅mL−1, 95% CI -0.24 to -0.05) after 12 months. MPA + t-E2, in contrast, did not demonstrate significant changes from baseline in traditional haemodynamic parameters.

      Conclusion

      The positive changes in traditional markers were not reflected in the cardiogenic biomarker, cfPWV, which has demonstrated a higher positive predictive value for cardiovascular events than traditional measurements.

      Keywords

      1. Introduction

      Vascular aging can now be predicted through an assessment of carotid-femoral Pulse Wave Velocity (cfPWV), a non-invasive cardiogenic biomarker of aortic stiffness [
      • Millasseau SC
      • Stewart AD
      • Patel SJ
      • Redwood SR
      • Chowienczyk PJ.
      Evaluation of Carotid–Femoral Pulse Wave Velocity Influence of Timing Algorithm and Heart Rate.
      ]. PWV is determined by the time taken for the arterial pulse pressure, generated by the systolic contraction of the heart, to propagate along the arterial tree [
      • Millasseau SC
      • Stewart AD
      • Patel SJ
      • Redwood SR
      • Chowienczyk PJ.
      Evaluation of Carotid–Femoral Pulse Wave Velocity Influence of Timing Algorithm and Heart Rate.
      ,
      • Pereira T
      • Correia C
      • Cardoso J.
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      ]. It provides a reflection of the properties of arterial vascular health, including arterial wall elasticity and its dimensions [
      • Millasseau SC
      • Stewart AD
      • Patel SJ
      • Redwood SR
      • Chowienczyk PJ.
      Evaluation of Carotid–Femoral Pulse Wave Velocity Influence of Timing Algorithm and Heart Rate.
      ,
      • Pereira T
      • Correia C
      • Cardoso J.
      Novel Methods for Pulse Wave Velocity Measurement.
      ], with higher values correlated to a relative fall in arterial compliance [
      • Pereira T
      • Correia C
      • Cardoso J.
      Novel Methods for Pulse Wave Velocity Measurement.
      ,
      • Yildiz M
      • Masatlýoglu S
      • Seymen P
      • Aytac E
      • Sahin B
      • Seymen HO
      The carotid-femoral (aortic) pulse wave velocity as a marker of arterial stiffness in familial Mediterranean fever.
      ]. Aortic stiffness is increased in conditions such as hypertension and is a strong independent indicator for subsequent cardiovascular events [
      • Millasseau SC
      • Stewart AD
      • Patel SJ
      • Redwood SR
      • Chowienczyk PJ.
      Evaluation of Carotid–Femoral Pulse Wave Velocity Influence of Timing Algorithm and Heart Rate.
      ,
      • Cheng YB
      • Li Y
      • Sheng CS
      • Huang QF
      • Wang JG.
      Quantification of the Interrelationship between Brachial-Ankle and Carotid-Femoral Pulse Wave Velocity in a Workplace Population.
      ,
      • Mennia C
      • Manginoa M
      • Ceceljab M
      • Psathaa M
      • Brosnanc MJ
      • Trimmerd J
      • Mohneye RP
      • Chowienczykb P
      • Padmanabhanf S
      • Spectora TD
      • Valdesa AM.
      Metabolomic study of carotid-femoral pulse-wave velocity in women.
      ]. The association between increased arterial stiffness and altered coronary perfusion is postulated to be through its effects on systolic blood pressure (SBP), subsequently increasing left ventricular afterload.
      The Framingham Risk Score [
      • Wilson PWF
      • D'Agostino RB
      • Levy D
      • Belanger AM
      • Silbershatz H
      • Kannel WB
      Prediction of Coronary Heart Disease Using Risk Factor Categories.
      ] has shown brachial pulse pressure (PP) to be a strong independent determinant of recurrent cardiac events and all-cause mortality in the general population. The risk of a major clinical event is said to increase by 10-40% for every 10mmHg increase in PP [
      • Mitchell GF.
      Arterial stiffness and wave reflection: Biomarkers of cardiovascular risk.
      ]. However, PP measurements and augmentation index (AI) are indirect surrogate measures of arterial stiffness which are influenced by factors related to cardiac function, such as heart rate (HR), limiting their interpretation [
      • Mitchell GF.
      Arterial stiffness and wave reflection: Biomarkers of cardiovascular risk.
      ,
      • Laurent S
      • Cockcroft J
      • Van Bortel L
      • Boutouyrie P
      • Giannattasio C
      • Hayoz D
      • Pannier B
      • Vlachopoulos C
      • Wilkinson I
      • Struijker-Boudier H
      European Network for Non-invasive Investigation of Large Arteries. Expert consensus document on arterial stiffness: methodological issues and clinical applications.
      ]. In contrast, aortic PWV is influenced to a lesser degree by other cardiac function parameters [
      • Mitchell GF.
      Arterial stiffness and wave reflection: Biomarkers of cardiovascular risk.
      ] and may be superior in the prediction of cardiovascular events over the Framingham Risk Score [
      • Lunder M
      • Janic M
      • Kejzar N
      • Sabovic M.
      Associations among different functional and structural arterial wall properties and their relations to traditional cardiovascular risk factors in healthy subjects: a cross-sectional study.
      ], brachial artery stiffness (carotid radial PWV), AI, central PP and PP amplification [
      • Van Bortel LM
      • Laurent S
      • Boutouyrie P
      • Chowienczyk P
      • Cruickshank JK
      • De Backer T
      • Filipovsky J
      • Huybrechts S
      • Mattace-Raso FUS
      • Protogerou AD
      • Schillaci G
      • Segers P
      • Vermeersch S
      • Weber T
      on behalf of the Artery Society, the European Society of Hypertension Working Group on Vascular Structure and Function and the European Network for Noninvasive Investigation of Large Arteries. Expert consensus document on the measurement of aortic stiffness in daily practice using carotid-femoral pulse wave velocity.
      ]. PWV, however, is affected by the age and body mass index (BMI) of the study population. The structural composition of the arterial walls changes with advancing age, contributing to the increased PWV [
      • Yildiz M
      • Masatlýoglu S
      • Seymen P
      • Aytac E
      • Sahin B
      • Seymen HO
      The carotid-femoral (aortic) pulse wave velocity as a marker of arterial stiffness in familial Mediterranean fever.
      ].
      The menopause has been shown to augment the age-dependent increase in arterial stiffness. Oestrogen deficiency is associated with increased concentrations of proinflammatory cytokines, which through a sequence of pathways, have been implicated in the inhibition of endothelium dependent vasodilation and nitric oxide synthesis [
      • Suzuki H
      • Kondo K.
      Pulse wave velocity in postmenopausal women.
      ].
      Studies have shown that women diagnosed with premature ovarian insufficiency (POI) have a higher incidence of total cardiovascular disease (CVD) (hazard ratio [HR] 1.61, 95% confidence interval [CI] 1.22 to 2.12, p=0.0007) [
      • Roeters van Lennep JE
      • Heida KY
      • Bots ML
      • Hoek A
      on behalf of the collaborators of the Dutch Multidisciplinary Guideline Development Group on Cardiovascular Risk Management after Reproductive D. Cardiovascular disease risk in women with premature ovarian insufficiency: A systematic review and meta-analysis.
      ] and mortality (80%) [
      • Maclaran K
      • Horner E
      • Panay N.
      Premature ovarian failure: long-term sequelae.
      ] secondary to this [
      • Nelson LM.
      Primary ovarian insufficiency.
      ,
      • De Vos M
      • Devroey P
      • Fauser BCJM
      Primary ovarian insufficiency.
      ], irrespective of the cause. The Framingham Study documented an increased risk of coronary heart disease (CHD) in postmenopausal women, particularly in the age category 40-44 years [
      • Gordon T
      • Kannel WB
      • Hjortland MC
      • McNamara PM.
      Menopause and coronary heart disease. The Framingham Study.
      ]. However, this risk has been shown to be two-fold higher in women diagnosed with POI (HR 2.2, 95% CI 1.0 to 4.9) compared to an early menopause (EM) (HR 1.2, 95% CI 0.7 to 2.0) [
      • Løkkegaard E
      • Jovanovic Z
      • Heitmann BL
      • Keiding N
      • Ottesen B
      • Pedersen AT.
      The association between early menopause and risk of ischaemic heart disease: Influence of Hormone Therapy.
      ]. A sub-analysis of the Multi-Ethnic Study of Atherosclerosis reported a 4% lower risk of heart failure for every year of increased menopausal age (HR 0.96, 95% CI 0.94 to 0.99) [
      • Ebong IA
      • Watson KE
      • Goff Jr, DC
      • Bluemke DA
      • Srikanthan P
      • Horwich T
      • Bertoni AG.
      Age at menopause and incident heart failure: the Multi-Ethnic Study of.
      ].
      Oestrogen deficiency is also associated with atherogenic changes in the lipid profile [
      • Bruschi F
      • Meschia M
      • Soma M
      • Perotti D
      • Paoletti R
      • Crosignani PG.
      Lipoprotein(a) and other lipids after oophorectomy and oestrogen replacement therapy.
      ] contributing to cardiovascular risk in this population. Knauff et al., (2008) [
      • Knauff EA
      • Westerveld HE
      • Goverde AJ
      • Eijkemans MJ
      • Valkenburg O
      • van Santbrink EJ
      • Fauser BC
      • van der Schouw YT.
      Lipid profile of women with premature ovarian failure.
      ] compared the lipid profile of 90 POI patients with 198 control subjects and demonstrated a significantly higher triglyceride level (mean difference 0.17 log mmol/L, 95% CI 0.06 to 0.29) and borderline lower high-density lipoprotein (HDL) cholesterol level in women with POI [
      • Knauff EA
      • Westerveld HE
      • Goverde AJ
      • Eijkemans MJ
      • Valkenburg O
      • van Santbrink EJ
      • Fauser BC
      • van der Schouw YT.
      Lipid profile of women with premature ovarian failure.
      ].
      The mainstay of management of women diagnosed with POI or an early menopause (EM) (EMPOI) is hormone replacement therapy (HRT). Most international guidance documents recommend treatment until at least the natural age of the menopause [
      • Panay N
      • Kalu E.
      Management of premature ovarian failure.
      ,
      • Hamoda H
      • Panay N
      • Pedder H
      • Arya R
      • Savvas M.
      The British Menopause Society & Women's Health Concern 2020 recommendations on hormone replacement therapy in menopausal women.
      ,
      • Pitkin J
      • Rees MCP
      • Gray S
      • Lumsden MA
      • Marsden J
      • Stevenson JC
      • Williamson J.
      British Menopause Society Council Consensus Statement: Management of premature menopause.
      ,
      • Fauser BCJM
      • Laven JSE
      • Tarlatzis BC
      • Moley KH
      • Critchley HOD
      • Taylor RN
      • Berga SL
      • Mermelstein PG
      • Devroey P
      • Gianaroli L
      • D'Hooghe T
      • Vercellini P
      • Hummelshoj L
      • Rubin S
      • Goverde AJ
      • De Leo V
      • Petraglia F
      Sex Steroid Hormones and Reproductive Disorders: Impact on Women's Health.
      ,
      • Vujovic S
      • Brincat M
      • Erel T
      • Gambacciani M
      • Lambrinoudaki I
      • Moen MH
      • Schenck-Gustafsson K
      • Tremollieres Rozenberg S
      • Rees M
      ,
      • Langrish JP
      • Mills NL
      • Bath LE
      • Warner P
      • Webb DJ
      • Kelnar CJ
      • Critchley HOD
      • Newby DE
      • Wallace WHB.
      Cardiovascular Effects of Physiological and Standard Sex Steroid Replacement Regimens in Premature Ovarian Failure.
      ]. The number of trials assessing the impact of HRT on cardiovascular events in women diagnosed with EMPOI are, however, limited. Kalantaridou et al., (2004) [
      • Kalantaridou SN
      • Naka KK
      • Papanikolaou E
      • et al.
      Impaired endothelial function in young women with premature ovarian failure: normalization with hormone therapy.
      ] assessed endothelial function in women with POI before and after 6 months of HRT (oral 0.625mg conjugated equine estrogen [CEE] with cyclical medroxyprogesterone acetate [MPA] 5mg). Women receiving HRT demonstrated a more than two-fold increase in flow-mediated dilation, comparable to the control group, but the findings are limited by the small sample size (n=18 versus controls n=20) and did not reach statistical significance [
      • Kalantaridou SN
      • Naka KK
      • Papanikolaou E
      • et al.
      Impaired endothelial function in young women with premature ovarian failure: normalization with hormone therapy.
      ].
      Langrish et al., (2009) [
      • Langrish JP
      • Mills NL
      • Bath LE
      • Warner P
      • Webb DJ
      • Kelnar CJ
      • Critchley HOD
      • Newby DE
      • Wallace WHB.
      Cardiovascular Effects of Physiological and Standard Sex Steroid Replacement Regimens in Premature Ovarian Failure.
      ] compared the impact of HRT (transdermal oestradiol [t-E2] 100-150mcg/day with cyclical vaginal progesterone 400mg/day for two weeks every month) with the combined oral contraceptive pill (COCP) (ethinylestradiol 30µg with 1•5mg norethisterone) on cardiovascular risk markers in women with POI. They concluded that HRT significantly lowered the mean 24-hour systolic and diastolic blood pressure and had a greater benefit on renal function than the COCP. The number who completed the study, however, is also small (n=18) [
      • Langrish JP
      • Mills NL
      • Bath LE
      • Warner P
      • Webb DJ
      • Kelnar CJ
      • Critchley HOD
      • Newby DE
      • Wallace WHB.
      Cardiovascular Effects of Physiological and Standard Sex Steroid Replacement Regimens in Premature Ovarian Failure.
      ].
      The Postmenopausal Estrogen/Progestin Interventions (PEPI) Trial analysed the impact of oestrogen (CEE 0.625mg daily) alone or in combination with one of three progestogen regimens (MPA 2.5mg/day; MPA 10mg on days 1-12; or micronised progesterone [MP] 200mg on days 1-12) on heart disease risk factors (HDL cholesterol levels; SBP; serum insulin levels; and fibrinogen levels) in postmenopausal women aged between 45-64 years. They demonstrated that unopposed oestrogen produced the greatest beneficial effects, but the high rate of endometrial hyperplasia restricted its use to women without a uterus. For women with a uterus, the cyclic use of MP produced the most favourable cardiovascular effects [
      The Writing Group for the PEPI Trial
      Effects of Estrogen/Progestin Regimens on heart Disease Risk Factors in Postmenopausal Women: The Postmenopausal Estrogen/Progestin Interventions (PEPI) Trial.
      ]. MP in combination with oestrogen has previously been shown to have no adverse effect on blood pressure [
      • Sitruk-Ware R.
      Progestogens in hormonal replacement therapy: new molecules, risks, and benefits.
      ] and insulin levels, both surrogate markers of CVD risk, with the most favourable impact seen on HDL levels [
      • Langrish JP
      • Mills NL
      • Bath LE
      • Warner P
      • Webb DJ
      • Kelnar CJ
      • Critchley HOD
      • Newby DE
      • Wallace WHB.
      Cardiovascular Effects of Physiological and Standard Sex Steroid Replacement Regimens in Premature Ovarian Failure.
      ].
      This pilot study aimed to assess the impact of MP and MPA in combination with t-E2 on the cardiovascular risk profile of women with EMPOI, through an analysis of both traditional surrogate markers and cfPWV, identified as the gold standard cardiogenic biomarker for risk stratification of arterial disease by the European Society for Cardiology [
      • Pereira T
      • Correia C
      • Cardoso J.
      Novel Methods for Pulse Wave Velocity Measurement.
      ,
      • Cheng YB
      • Li Y
      • Sheng CS
      • Huang QF
      • Wang JG.
      Quantification of the Interrelationship between Brachial-Ankle and Carotid-Femoral Pulse Wave Velocity in a Workplace Population.
      ].

      2. Methodology

      2.1 Study objective

      To compare the CVD risk of MP versus MPA in combination with t-E2 in the management of women diagnosed with EMPOI.

      2.2 Primary endpoint

      The main outcome measure was cfPWV.

      2.3 Secondary outcome measures

      The secondary endpoints were HR, SBP, diastolic blood pressure (DBP), PP, AI, cardiac output (CO), stroke volume (SV), total peripheral resistance (TPR), total cholesterol, HDL, low density lipoprotein levels (LDL), triglyceride (TG) levels and cholesterol ratio (total cholesterol/HDL ratio).

      2.4 Study design

      Women under the age of 45-years diagnosed with EMPOI, with an intact uterus, were prospectively invited and recruited from two tertiary referral Reproductive Endocrine and Menopause Clinics between April 2013 and August 2015. Subjects fulfilling the inclusion and exclusion criteria (Table 1) were randomised into one of two treatment arms using a web-based computer randomisation software, Graph Pad. Both groups were prescribed 50mcg/day of t-E2 in the form of Evorel® Patches in conjunction with either cyclical MP (Utrogestan®) 200mg orally on days 15-26 of a 28-day cycle or, MPA (Provera®) 10mg orally on days 16-26 of a 28-day cycle. The medication was prescribed and administered in accordance with the Summary of Product Characteristics of each of the medications. The total duration of the study period was 12-months. The study was open labelled where neither the participant nor researcher were blinded to the randomisation secondary to limited funds.
      Table 1Inclusion and exclusion criteria.
      Inclusion CriteriaExclusion Criteria
      Females aged between 18 and up to 45-years of ageAge <18 or >45-years of age
      Confirmed diagnosis of POI or EMPregnant or lactating females
      Willingness to participateContraindication to the use of hormonal preparations
      No concomitant co-morbidities that would contraindicate the use of hormonal preparationsFactors present in the medical history:

      that would contribute to an increased risk of CVD

      known thrombophilia

      known porphyria

      known liver disease

      known past or suspected breast cancer

      undiagnosed vaginal bleeding

      genital tract carcinoma
      Smokers
      Body mass index >35Kg/m2
      The use of concomitant medications that could influence the results, such as anti-hypertensives
      Known hypersensitivity to any of the active substances or excipients contained within Utrogestan®, Provera® or Evorel patches®; known allergy to peanuts or soya
      Key: POI – premature ovarian insufficiency; EM – early menopause; CVD – cardiovascular disease.
      Prior to randomisation, participants previously prescribed hormonal treatment were asked to stop it for a minimum of four weeks, designated a washout period, to enable a baseline assessment to be undertaken.
      Measurements were undertaken at baseline and repeated at intervals of 3-, 6-, and 12-months.

      2.5 Carotid-Femoral Pulse Wave Velocity

      Carotid-femoral PWV was measured using an oscillometric technique to acquire the pulse waveform (Vicorder device, Skidmore Medical). Patients were placed into the supine position at approximately 450, in room temperature, and allowed to rest and re-climatise for 5-10 mins prior to assessment. A 10cm-wide cuff was placed over the upper thigh, in the region of the femoral pulse. Simultaneously, a 3cm-wide cuff was placed around the neck for the carotid pulse. The distance between the suprasternal notch and mid upper thigh cuff was measured and utilised in the assessment, as per the manufacturer's instructions. The cuffs were simultaneously inflated, and the pulse waveforms recorded. Measurements were made in duplicate/triplicate, and the mean values used for analysis.

      2.6 Sample collection

      Venous blood was collected for total cholesterol, HDL, LDL, and triglyceride levels. Blood was collected into a serum separating vacutainer with clot activator (BD Diagnostics, Plymouth, UK). All laboratory measurements were carried out at King's College Hospital, Haematology and Clinical Biochemistry Laboratories.

      2.7 Statistical analysis and sample size

      Univariate analysis was performed to compare age, BMI, and ethnicity between the two groups. Continuous variables were compared using paired t-test between visits and baseline for each group. Multiple logistic regression was undertaken to compare continuous variables adjusting for age, BMI, and ethnicity. A p-value of <0.05 was considered statistically significant. All analyses were performed using IBM SPSS Statistics version 22.0 (Statistical Package for Social Sciences, Chicago, USA).
      We aimed to recruit 90 women in total to allow for a 10% drop out and loss to follow up. The sample size was calculated using the Altman nomogram [
      • Gore SM
      • Altman DG.
      Statistics in Practice.
      ] and based on changes in the AI in response to exposure to the two different progestogens assessed in the study. Baseline data were obtained from a reference population, Pulse Wave Analysis (PWA) and AI data as reported by McEniery et al., (2005) [
      • McEniery CM
      • Yasmin Hall IR
      • Ahmad QA
      • et al.
      Normal Vascular Aging: Differential Effects on Wave Reflection and Aortic Pulse Wave Velocity.
      ]. Based on the reported findings, we considered a change of 8% with Standard Deviation of 12 to be a clinically significant difference to detect. Using the Altman nomogram, this would give a standardised difference of 0.80. A sample of 80 women in two groups would detect a standardised difference of 0.80 with 80% power at the 5% level of significance.

      3. Results

      3.1 Patient characteristics

      Seventy-one participants consented to the study. Five were excluded as they did not meet the study inclusion criteria (Table 1) (diabetes [n=1]; hereditary thrombophilia [n=3]; spontaneous pregnancy [n=1]). Fig. 1 outlines the study algorithm.
      Fig. 1
      Fig. 1Study algorithm. *Side effects: no adverse effects were reported to the MHRA; **Loss to follow up: unable to contact the patient for a visit. The differences in numbers between the PWV and lipoprotein arm is secondary to either missing data or patients withdrawing consent at different points.
      The baseline demographic characteristics and haemodynamic parameters for the two study populations did not differ (Table 2). The age of the participants ranged between 19 years to 44 years of age. Of these, 43 of the participants were ≤40 years of age (POI), and 23 were >40 years of age (EM).
      Table 2Baseline demographic characteristics and haemodynamic parameters for the two study populations.
      CategoryPWV cohortLipoprotein profile cohort
      MP + t-E2MPA + t-E2P-valueMP + t-E2MPA + t-E2P-value
      Age (years)35.75±6.5237.97±6.020.5335.75±6.5237.97±6.020.17
      BMI (Kg/m2)25.23±5.1724.90±4.210.7925.23±5.1724.90±4.210.79
      Ethnicity0.300.83

      Asian2727
      Black117127
      White13131413
      Other2242
      HR (bpm)68.38±10.0469.89±10.340.58
      SBP (mmHg)122.67±12.32124.05±12.610.68
      DBP (mmHg)69.02±8.3269.58±7.180.79
      PP (mmHg)53.69±7.3354.96±8.570.55
      AI (%)20.73±7.3519.87±5.620.63
      CO (mL/min)5.87±1.036.02±1.130.61
      SV (ml)89.03±12.4889.62±16.980.88
      TPR (mmHg⋅min⋅mL−1)0.98±0.180.96±0.150.63
      PWV (m/s)6.39±1.726.39±1.171.00
      Total cholesterol (mmol/l)4.65±0.674.77±0.790.09
      HDL (mmol/l)1.74±0.421.54±0.400.40
      LDL (mmol/l)2.47±0.582.66±0.630.20
      TG (mmol/l)0.98±0.451.26±0.780.35
      Cholesterol ratio (mmol/l)2.79±0.673.31±1.050.55
      Key: Data expressed as mean±standard deviation [SD]; t-E2 – transdermal oestradiol; MP – micronised progesterone; MPA – medroxyprogesterone acetate; BMI – body mass index; HR – heart rate; SBP – systolic blood pressure; DBP – diastolic blood pressure; PP – Pulse pressure (difference between the systolic and diastolic blood pressure); AI – Augmentation index (quantifies the extent of augmented pressure relative to the central pulse pressure, and is the difference between the second and first systolic peaks expressed as a percentage of the pulse pressure); CO – cardiac output; SV – stroke volume; TPR – total peripheral resistance; PWV – pulse wave velocity (speed of travel of the pulse along a specified arterial segment); HDL – high density lipoprotein; LDL – low density lipoprotein; TG – triglyceride; cholesterol ratio – total cholesterol/HDL.
      PWV did not demonstrate significant changes from baseline for the duration of the study, in either treatment arm (Table 3).
      Table 3Impact of MP and MPA in combination with t-E2 on the haemodynamic variables after 3-, 6- and 12-months, presented as mean difference (±SD), with 95% confidence intervals.
      VariablesMP + t-E2MPA + t-E2
      Mean difference after 3-months (95% CI)Mean difference after 6-months (95% CI)Mean difference after 12-months (95% CI)Mean difference after 3-months (95% CI)Mean difference after 6-months (95% CI)Mean difference after 12-months (95% CI)
      HR1.16±12.88 (-4.56 to 6.87)7.69±12.80 (1.10 to 14.27)*4.71±13.97 (-2.24 to 11.65)2.89±12.67 (-2.72 to 8.51)1.99±11.49 (-3.55 to 7.53)4.03±18.07 (-5.98 to 14.03)
      SBP-0.65±10.20 (-5.18 to 3.87)-3.61±7.39 (-7.41 to 0.19)-1.69±7.78 (-5.56 to 2.18)-2.14±11.64 (-7.30 to 3.02)-2.42±9.78 (-7.13 to 2.30)1.18±8.95 (-3.77 to 6.14)
      DBP-1.75±6.57 (-4.66 to 1.17)-2.94±5.60 (-5.81 to -0.06)-3.43±6.31 (-6.57 to -0.29)*-2.08±7.18 (-5.26 to 1.10)-2.12±6.86 (-5.42 to 1.19)0.97±8.40 (-3.69 to 5.62)
      PP1.32±9.85 (-3.05 to 5.68)-1.29±6.50 (-4.63 to 2.05)1.83±4.69 (-0.50 to 4.17)-0.32±8.63 (-4.15 to 3.51)-0.42±7.98 (-4.27 to 3.43)-0.80±6.43 (-4.36 to 2.76)
      AI-1.16±10.48 (-5.81 to 3.48)-1.16±10.48 (-5.81 to 3.48)2.14±6.52 (-1.21 to 5.49)1.19±5.83 (-1.39 to 3.78)0.73±6.12 (-2.21 to 3.68)2.49±5.60 (-0.61 to 5.59)
      CO0.46±1.37 (-0.15 to 1.06)0.34±0.95 (-0.14 to 0.83)0.71± 1.01 (0.20 to 1.21)*-0.07±1.37 (-0.68 to 0.54)0.03±1.18 (-0.54 to 0.60)-0.06±1.29 (-0.77 to 0.66)
      SV3.31±18.76 (-5.01 to 11.62)-3.51±14.63 (-11.03 to 4.02)3.54±12.07 (-2.47 to 9.54)-3.10±16.07 (-10.22 to 4.03)-0.68±16.69 (-8.73 to 7.36)0.20±11.38 (-6.10 to 6.50)
      TPR-0.08±0.21 (-0.18 to 0.02)-0.07±0.16 (-0.15 to 0.01)-0.15±0.19 (-0.24 to -0.05)*-0.02±0.18 (-0.10 to 0.06)0.00±0.22 (-0.11 to 0.11)0.04±0.24 (-0.09 to 0.18)
      PWV0.40±3.32 (-1.07 to 1.87)0.22±1.38 (-0.49 to 0.93)0.28±0.87 (-0.15 to 0.71)-0.02±1.04 (-0.48 to 0.44)-0.07±1.05 (-0.58 to 0.43)0.64±1.62 (-0.26 to 1.54)
      Total cholesterol-0.16±0.45 (-0.37 to 0.06)-0.13±0.50 (-0.37 to 0.11)0.02±0.47 (-0.23 to 0.26)-0.28±0.55 (-0.52 to -0.04)*-0.21±0.51 (-0.46 to 0.04)-0.23±0.51 (-0.51 to 0.06)
      HDL-0.07±0.17 (-0.15 to 0.01)-0.03±0.21 (-0.13 to 0.07)-0.09±0.20 (-0.20 to 0.01)-0.12±0.46 (-0.32 to 0.08)*-0.16±0.27 (-0.29 to -0.02)*-0.11±0.21 (-0.22 to 0.01)
      LDL-0.04±0.35 (-0.20 to 0.13)-0.01±0.35 (-0.17 to 0.16)0.09±0.31 (-0.07 to 0.26)-0.12±0.46 (-0.32 to 0.08)-0.11±0.45 (-0.34 to 0.13)-0.02±0.45 (-0.27 to 0.23)
      TG-0.12±0.49 (-0.34 to 0.11)-0.16±0.59 (-0.45 to 0.12)0.06±0.48 (-0.18 to 0.31)-0.13±0.54 (-0.37 to 0.10)0.17±0.73 (-0.19 to 0.53)-0.21±0.37 (-0.42 to -0.01)*
      Cholesterol ratio0.03±0.30 (-0.11 to 0.18)0.03±0.37 (-0.16 to 0.21)0.18±0.30 (0.02 to 0.33)*0.07±0.46 (-0.14 to 0.27)0.25±0.67 (-0.10 to 0.59)0.05±0.38 (-0.17 to 0.27)
      Key: Data expressed as mean±standard deviation [SD]; t-E2 – transdermal oestradiol; MP – micronised progesterone; MPA – medroxyprogesterone acetate; HR – heart rate (bpm); SBP – systolic blood pressure (mmHg); DBP – diastolic blood pressure (mmHg); PP – Pulse pressure (mmHg); AI – Augmentation index (%); CO – cardiac output (mL/min); SV – stroke volume (ml); TPR – total peripheral resistance (mmHg⋅min⋅mL−1); PWV – pulse wave velocity (m/s); HDL – high density lipoprotein (mmol/l); LDL – low density lipoprotein (mmol/l); TG – triglyceride (mmol/l); cholesterol ratio – total cholesterol/HDL. *p<0.05.
      MP + t-E2 demonstrated a positive effect on traditional CVD markers, with a significant improvement seen in CO (0.71±1.01mL/min, 95% CI 0.20 to 1.21, p=0.01) and reduction in DBP (-3.43±6.31mmHg, 95% Cl -6.57 to -0.29, p=0.03) and TPR (-0.15±0.19mmHg⋅min⋅mL−1, 95% CI -0.24 to -0.05, p=0.01) after 12-months duration compared to baseline. A significant increase in cholesterol ratio was seen after 12-months duration (0.18±0.30mmol/l, 95% CI 0.02 to 0.33, p=0.03) during the same study period.
      MPA + t-E2 in contrast, did not demonstrate significant changes from baseline in HR, SBP, PP, AI, CO or TPR. Total cholesterol levels, however, were initially lowered after 3-months duration (-0.28±0.55mmol/l, 95% Cl -0.52 to -0.04, p=0.02). HDL levels demonstrated a significant decline after 3-months (-0.13±0.24mmol/l, 95% CI -0.23 to -0.02, p=0.02) and 6-months (-0.16±0.27mmol/l, 95% CI -0.29 to -0.02, p=0.03) duration from baseline. This difference was not maintained by 12-months duration (-0.11±0.21mmol/l, 95% CI -0.22 to 0.01, p=0.07). After 12-months duration, however, significant reductions were seen in the TG levels (-0.21±0.37mmol/l, 95% CI -0.42 to -0.01, p=0.04).

      4. Discussion

      MP + t-E2 resulted in a significant improvement in CO and reduction in DBP and TPR after 12-months of treatment. PWV, however, did not significantly change from baseline in either treatment arm over the course of the study, consistent with the neutral changes observed in the AI and PP readings over the same period.
      No significant changes were observed in the lipoprotein profile of the MP + t-E2 treatment arm, in keeping with MP having a more selective effect on progesterone receptors [
      • Jensen J
      • Riis BJ
      • Strom V
      • Nilas L
      • Christiansen C.
      Long term effects of percutaneous estrogens and oral progesterone on serum lipoproteins in postmenopausal women.
      ,
      • Ottosson UB
      • Johansson BG
      • von Schoultz B
      Subfractions of high-density lipoprotein cholesterol during estrogen replacement therapy: A comparison between progestogens and natural progesterone.
      ]. Furthermore, no significant changes were demonstrated in the LDL levels, the serum levels of which have been closely correlated to the development of coronary artery disease, supporting the notion of a neutral impact of MP on CVD risk [
      • Krauss RM.
      Effects of Progestational Agents on Serum Lipids and Lipoproteins.
      ,
      • Nath A
      • Sitruk-Ware R.
      Different cardiovascular effects of progestins according to structure and activity.
      ].
      MPA + t-E2, in contrast, resulted in changes in the lipoprotein profile that included a reduction in total cholesterol levels at 3-months of treatment, HDL levels at 6-months and TG levels after 12-months of treatment. The TG lowering effect seen with MPA + t-E2 may have occurred secondary to its antagonistic effect on oestrogen stimulated hepatic TG synthesis or due to increased activity of the lipoprotein lipase enzyme [
      • Krauss RM.
      Effects of Progestational Agents on Serum Lipids and Lipoproteins.
      ]. MPA + t-E2, however, did not demonstrate significant changes in any of the other measured traditional haemodynamic parameters, including CO and TPR.
      The findings overall, showed that MP and MPA given in combination with t-E2 did not adversely impact CVD risk markers as assessed by the gold standard cardiogenic biomarker, cfPWV when used in the management of women with EMPOI. Furthermore, MP + t-E2 resulted in a significant improvement in CO and reduction in DBP and TPR.
      Central arterial stiffness has been shown to be an independent predictor of cardiovascular risk and all-cause mortality [
      • Hickson SS
      • Butlin M
      • Broad J
      • Avolio AP
      • Wilkinson IB
      • McEniery CM.
      Validity and repeatability of the Vicorder apparatus: a comparison with the SphygmoCor device.
      ,
      • Blacher J
      • Asmar R
      • Djane S
      • London GM
      • Safar ME.
      Aortic Pulse Wave Velocity as a Marker of Cardiovascular Risk in Hypertensive Patients.
      ,
      • Boutouyrie P
      • Tropeano AI
      • Asmar R
      • Gautier I
      • Benetos A
      • Lacolley P
      • Laurent S.
      Aortic Stiffness Is an Independent Predictor of Primary Coronary Events in Hypertensive Patients. A Longitudinal Study.
      ]. Invasive catheterisation of the ascending aorta remains the gold standard technique for central haemodynamic assessment, but the invasive nature of the technique has limited its clinical applicability. The analysis of peripheral waveforms with dedicated devices has provided the opportunity to assess central haemodynamic parameters non-invasively and has shown good correlation with direct central blood pressure measurements obtained during invasive catheterisation of the ascending aorta [
      • Hickson SS
      • Butlin M
      • Broad J
      • Avolio AP
      • Wilkinson IB
      • McEniery CM.
      Validity and repeatability of the Vicorder apparatus: a comparison with the SphygmoCor device.
      ]. Salvi et al., (2019) [
      • Salvi P
      • Scalise F
      • Rovina M
      • Moretti F
      • Salvi L
      • Grillo A
      • Gao L
      • Baldi C
      • Faini A
      • Furlanis G
      • Sorropago A
      • Millasseau SC.
      Noninvasive Estimation of Aortic Stiffness Through Different Approaches: Comparison With Intra-Aortic Recordings.
      ] demonstrated a strong correlation between non-invasive devices measuring cfPWV (Complior Analyse, PulsePen ET, PulsePen ETT, and SphygmoCor) and invasive aortic PWV measurements (r>0.83) [
      • Salvi P
      • Scalise F
      • Rovina M
      • Moretti F
      • Salvi L
      • Grillo A
      • Gao L
      • Baldi C
      • Faini A
      • Furlanis G
      • Sorropago A
      • Millasseau SC.
      Noninvasive Estimation of Aortic Stiffness Through Different Approaches: Comparison With Intra-Aortic Recordings.
      ]. The Vicorder system (Skidmore Medical Limited) used in this study has been calibrated to non-invasively record mean aortic and diastolic pressures and has also been validated against other devices used in this context [
      • Hickson SS
      • Butlin M
      • Broad J
      • Avolio AP
      • Wilkinson IB
      • McEniery CM.
      Validity and repeatability of the Vicorder apparatus: a comparison with the SphygmoCor device.
      ].
      The difference in obtained measurements between invasive aortic PWV and non-invasive cfPWV readings can be attributed to three main causes [
      • Salvi P
      • Scalise F
      • Rovina M
      • Moretti F
      • Salvi L
      • Grillo A
      • Gao L
      • Baldi C
      • Faini A
      • Furlanis G
      • Sorropago A
      • Millasseau SC.
      Noninvasive Estimation of Aortic Stiffness Through Different Approaches: Comparison With Intra-Aortic Recordings.
      ]. Firstly, cfPWV assessments do not incorporate the ascending aorta in the path of travel. Secondly, cfPWV measurements include arterial segments in which the path of travel of the pulse can be in an opposite direction (brachiocephalic trunk and common carotid artery) to the thoracic aorta. Thirdly, cfPWV measurements include segments of the femoral artery in their evaluation. The muscular component of the femoral artery is greater than that of the aorta. This difference between muscular and elastic arteries can increase the PWV assessment in younger individuals. This difference, however, is reversed with advancing age, thus, cfPWV measurements may be overestimated in younger adults and underestimated in older individuals. Furthermore, PWV exponentially increases in aortic arteries with increasing age, but only weakly and linearly increases with age within the muscular arteries of the lower limbs [
      • Wohlfahrt P
      • Krajčoviechová A
      • Seidlerová J
      • Galovcová M
      • Bruthans J
      • Filipovský J
      • Laurent S
      • Cífková R
      Lower-extremity arterial stiffness vs. aortic stiffness in the general population.
      ].
      Overall, cfPWV is deemed to be a highly reproducible, non-invasive emerging cardiogenic biomarker of arterial stiffness and thus, cardiovascular risk stratification [
      • Pereira T
      • Correia C
      • Cardoso J.
      Novel Methods for Pulse Wave Velocity Measurement.
      ,
      • Collaboration The Reference Values for Arterial Stiffness’
      Determinants of pulse wave velocity in healthy people and in the presence of cardiovascular risk factors: ‘establishing normal and reference values.
      ]. It is considered to be the gold standard measure for arterial stiffness [
      • Laurent S
      • Cockcroft J
      • Van Bortel L
      • Boutouyrie P
      • Giannattasio C
      • Hayoz D
      • Pannier B
      • Vlachopoulos C
      • Wilkinson I
      • Struijker-Boudier H
      European Network for Non-invasive Investigation of Large Arteries. Expert consensus document on arterial stiffness: methodological issues and clinical applications.
      ,
      • Collaboration The Reference Values for Arterial Stiffness’
      Determinants of pulse wave velocity in healthy people and in the presence of cardiovascular risk factors: ‘establishing normal and reference values.
      ], however, its clinical application is limited by population-based differences seen within the vascular behaviour in varying physiological and pathological conditions [
      • Farro I
      • Bia D
      • Zocalo Y
      • Torrado J
      • Farro F
      • Florio L
      • Olascoaga A
      • Alallon W
      • Lluberas R
      • Armentano RL.
      Pulse Wave Velocity as Marker of Preclinical Arterial Disease: Reference Levels in a Uruguayan Population Considering Wave Detection Algorithms, Path Lengths, Aging, and Blood Pressure.
      ], including the different methodologies available to measure the path length and algorithms used to calculate PWV [
      • Collaboration The Reference Values for Arterial Stiffness’
      Determinants of pulse wave velocity in healthy people and in the presence of cardiovascular risk factors: ‘establishing normal and reference values.
      ].
      The European Society of Hypertension have stated that a PWV of 12m/s can signify vascular damage and/or cardiovascular risk. The baseline PWV for both treatment arms and the change from baseline throughout the study duration was approximately 50% lower than the single cut-off value used by the European Society of Hypertension. This single value, however, cannot be extrapolated to all populations, as it can underestimate the level of risk in young subjects and overestimate the level of risk in older individuals [
      • Farro I
      • Bia D
      • Zocalo Y
      • Torrado J
      • Farro F
      • Florio L
      • Olascoaga A
      • Alallon W
      • Lluberas R
      • Armentano RL.
      Pulse Wave Velocity as Marker of Preclinical Arterial Disease: Reference Levels in a Uruguayan Population Considering Wave Detection Algorithms, Path Lengths, Aging, and Blood Pressure.
      ]. More directed ranges accounting for age and ethnicity are therefore needed for clinical application of this cardiogenic biomarker in everyday practice [
      • Kasliwal RR
      • Mahansaria K
      • Bansal M.
      Central Aortic Blood Pressure and Pulse Wave Velocity as Additional Markers in Patients with Hypertension.
      ].
      A meta-analysis by Laugesen E et al., (2013) [
      • Laugesen E
      • Høyem P
      • Stausbøl-Grøn B
      • Mikkelsen A
      • Thrysøe S
      • Erlandsen M
      • Christiansen JS
      • Knudsen ST
      • Hansen KW
      • Kim WY
      • Hansen TK
      • Poulsen PI.
      Carotid-femoral pulse wave velocity is Associated with cerebral white matter Lesions in type 2 diabetes.
      ] concluded that a 1m/s increase in PWV is associated with a 14% (95% CI 9 to 20%), 15% (95% CI 9 to 21%) and 15% (95% CI 9 to 21%) increased risk of total cardiovascular events, cardiovascular mortality, and all-cause mortality, respectively when controlled for risk factors [
      • Laugesen E
      • Høyem P
      • Stausbøl-Grøn B
      • Mikkelsen A
      • Thrysøe S
      • Erlandsen M
      • Christiansen JS
      • Knudsen ST
      • Hansen KW
      • Kim WY
      • Hansen TK
      • Poulsen PI.
      Carotid-femoral pulse wave velocity is Associated with cerebral white matter Lesions in type 2 diabetes.
      ]. In this study, the PWV for both treatment arms changed by <1m/s from baseline, consistent with no significant changes observed in this parameter throughout the study.
      Vlachopoulos C et al., (2010) [
      • Vlachopoulos C
      • Aznaouridis K
      • O'Rourke MF
      • Safar ME
      • Baou K
      • Stefanadis C
      Prediction of cardiovascular events and all-cause mortality with central haemodynamics: a systematic review and meta-analysis.
      ] conducted a meta-analysis of 11 longitudinal studies including 5,648 subjects with a mean follow up duration of 45-months. The studies included both men and women of broad age categories, the youngest of which were 42.6±11.2 years. They found that the relative risk of cardiovascular events increased by 8.8% (n=3285; 95% CI 1.04 to 1.14) with a 10mmHg increase in central systolic pressure, 13.7% (n=4778; 95% CI 1.06 to 1.22) with a 10mmHg increase in central PP and 31.8% (n=1326; 95% CI 1.09 to 1.59) with a 10% absolute increase in central AI. A 10% increase in central AI was also correlated to a 38.4% (n=569; 95% CI 1.19 to 1.61) increased risk of all-cause mortality [
      • Vlachopoulos C
      • Aznaouridis K
      • O'Rourke MF
      • Safar ME
      • Baou K
      • Stefanadis C
      Prediction of cardiovascular events and all-cause mortality with central haemodynamics: a systematic review and meta-analysis.
      ]. Adkisson EJ et al., (2010) [
      • Adkisson EJ
      • Casey DP
      • Beck DT
      • Gurovich AN
      • Martin JS
      • Braith RW.
      Central, peripheral and resistance arterial reactivity: fluctuates during the phases of the menstrual cycle.
      ] reported a 5-9% reduction in cardiac morbidity and 4% reduction in all-cause mortality when the SBP is lowered by 3mmHg [
      • Adkisson EJ
      • Casey DP
      • Beck DT
      • Gurovich AN
      • Martin JS
      • Braith RW.
      Central, peripheral and resistance arterial reactivity: fluctuates during the phases of the menstrual cycle.
      ]. This study did not demonstrate significant changes in PP, AI or SBP, in either treatment arm from baseline, with absolute levels differing by <10mmHg and <10% change in AI at 12-months, consistent with the neutral changes observed in the PWV readings over the same study period.
      Hypertension, overall, is an important risk predictor for CVD. Current practice estimates a greater prediction of all cardiovascular events with raised SBP, but both systolic and diastolic hypertension being independent predictors of adverse cardiovascular outcomes [
      • Flint AC
      • Conell C
      • Ren X
      • Banki NM
      • Chan SL
      • Rao VA
      • Melles RB
      • Bhatt DL.
      Effect of Systolic and Diastolic Blood Pressure on Cardiovascular Outcomes.
      ]. Systolic hypertension occurs because of increased SV and/or arterial stiffness [
      • Franklin SS.
      The importance of diastolic blood pressure in predicting cardiovascular risk.
      ]. DBP, has been proposed by researchers, including the Framingham investigators, to drive coronary risk in younger subjects [
      • Kannel WB
      • Gordon T
      • Schwartz MJ
      Systolic versus diastolic blood pressure and risk of coronary heart disease.
      ,
      • Franklin SS
      • Larson MG
      • Khan SA
      • Wong ND
      • Leip EP
      • Kannel WB
      • et al.
      Does the relation of blood pressure to coronary heart disease risk change with aging?.
      ,
      • Stamler J
      • Neaton JD
      • Wentworth D.
      Blood pressure (systolic and diastolic) and risk of fatal coronary heart disease.
      ,
      • Li Y
      • Wei FF
      • Wang S
      • Cheng YB
      • Wang JG.
      Cardiovascular Risks Associated With Diastolic Blood Pressure and Isolated Diastolic Hypertension.
      ], peaking in the fifth decade of life [
      • Vlachopoulos C
      • Aznaouridis K
      • O'Rourke MF
      • Safar ME
      • Baou K
      • Stefanadis C
      Prediction of cardiovascular events and all-cause mortality with central haemodynamics: a systematic review and meta-analysis.
      ], with SBP becoming of greater importance in older people [
      • Kannel WB
      • Gordon T
      • Schwartz MJ
      Systolic versus diastolic blood pressure and risk of coronary heart disease.
      ,
      • Franklin SS
      • Larson MG
      • Khan SA
      • Wong ND
      • Leip EP
      • Kannel WB
      • et al.
      Does the relation of blood pressure to coronary heart disease risk change with aging?.
      ,
      • Stamler J
      • Neaton JD
      • Wentworth D.
      Blood pressure (systolic and diastolic) and risk of fatal coronary heart disease.
      ,
      • Li Y
      • Wei FF
      • Wang S
      • Cheng YB
      • Wang JG.
      Cardiovascular Risks Associated With Diastolic Blood Pressure and Isolated Diastolic Hypertension.
      ]. This study did not identify changes in the SBP over the course of the research, consistent with no changes observed in SV and AI. The DBP, however, did demonstrate a significant reduction after 12-months duration (-3.43±6.31mmHg, 95% Cl -6.57 to -0.29, p=0.03) in the MP + t-E2 treatment arm. This could reflect the younger cohort included in this study and support the more favourable impact of MP in combination with t-E2 when traditional surrogate markers of CVD risk are assessed.
      The HR increased by 4-5 beats per minute after 12-months duration from baseline in both treatment arms, with a significant increase recorded in the MP + t-E2 (7.69±12.80bpm, 95% CI 1.10 to 14.27, p=0.03) treatment arm at 6-months duration. Cardiac function, such as HR, has been shown to influence the parameters PP and AI [
      • Mitchell GF.
      Arterial stiffness and wave reflection: Biomarkers of cardiovascular risk.
      ,
      • Laurent S
      • Cockcroft J
      • Van Bortel L
      • Boutouyrie P
      • Giannattasio C
      • Hayoz D
      • Pannier B
      • Vlachopoulos C
      • Wilkinson I
      • Struijker-Boudier H
      European Network for Non-invasive Investigation of Large Arteries. Expert consensus document on arterial stiffness: methodological issues and clinical applications.
      ], with this difference potentially providing an explanation for the fluctuations seen in the PP and AI readings over the course of the study.
      Epidemiological studies, such as the Framingham cohort studies in the USA, have identified several CVD risk factors including the lipid profile. Risk stratification models to estimate a patient's 10-year risk of developing CVD incorporate lipid parameters in their equation. Both the Framingham-based equations [
      • Anderson KM
      • Wilson PW
      • Odell PM
      • Kannel WB.
      An updated coronary risk profile. A statement for health professionals.
      ] and the European Systematic COronary Risk Evaluation (SCORE) algorithm [
      • Conroy RM
      • PyoÈraÈ la K
      • Fitzgerald AP
      • Sans S
      • Menotti A
      • De Backer G
      • et al.
      Estimation of ten-year risk of fatal cardiovascular disease in Europe: the SCORE project.
      ] consider total cholesterol, HDL cholesterol and the ratio of total cholesterol to HDL to be the strongest predictors. The Framingham Offspring study followed a cohort of individuals for 20-years and identified that any combination of low levels of HDL cholesterol, high levels of LDL cholesterol and high levels of TG were associated with an increased risk of CVD [
      • Andersson C
      • Lyass A
      • Vasan RS
      • Massaro JM
      • Sr D'Agostino RB
      • Robins SJ
      Long-term risk of cardiovascular events across a spectrum of adverse major plasma lipid combinations in the Framingham Heart Study.
      ]. The National Clinical Guideline Centre for Cardiovascular Risk Assessment for primary prevention of CVD, however, recommend using the QRISK2 risk assessment tool to assess cardiovascular risk for the primary prevention of CVD in people aged ≤84 years which incorporates the cholesterol ratio, total cholesterol/HDL in its calculation [
      • Hippisley-Cox J
      • Coupland C
      • Vinogradova Y
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      • Brindle P.
      Derivation and validation of QRISK, a new cardiovascular disease risk score for the United Kingdom: prospective open cohort study.
      ].
      LDL is the predominant cholesterol-carrying lipoprotein and main atherogenic lipoprotein. Epidemiological data has suggested that isolated low HDL levels is a strong and independent risk factor for CVD [
      • Goldbourt U
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      • Medalie JH.
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      ,
      • Lamarche B
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      ,
      • Kontush A.
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      ]. HDL particles may act as a protective factor against atherosclerosis through several biological mechanisms including the prevention of endothelial activation, inflammation, and oxidative stress, as well as enhancing nitric oxide production and promoting cholesterol efflux to reduce lesion formation and maintain barrier integrity [

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      ], regulating gene expression by virtue of micro-RNAs, and improving glucose metabolism [
      • Kontush A.
      HDL-mediated mechanisms of protection in cardiovascular disease.
      ].
      A subsidiary analysis of a clinical trial in Australia, New Zealand, and Finland, looking at the impact of Long-Term Intervention with Pravastatin in Ischaemic Disease (LIPID study), (n=9014 aged between 31-75 years with recorded acute coronary syndrome), acknowledged that all lipid parameters were linked with future coronary events. They concluded that the total cholesterol/HDL ratio, LDL/HDL ratio or apolipoprotein B/apolipoprotein A1 ratio were more superior in predicting the time to coronary event than a single lipid measure [
      • Glasziou PP
      • Irwig L
      • Kirby AC
      • et al.
      Which lipid measurement should we monitor? An analysis of the LIPID study.
      ].
      The ESCARVAL Study Group found that HDL levels, total cholesterol/HDL ratios and TG/HDL ratios were better predictors for mortality and CVD than other lipid parameters commonly used in clinical practice [
      • Orozco-Beltran D
      • Gil-Guillen VF
      • Redon J
      • Martin-Moreno JM
      • Pallares-Carratala V
      • Navarro-Perez J
      • et al.
      Lipid profile, cardiovascular disease and mortality in a Mediterranean high-risk population: The ESCARVAL-RISK study.
      ]. They demonstrated a positive association with the TG/HDL ratio and an inverse association with total cholesterol, HDL, and LDL on age-adjusted mortality rates (deaths/10,000 person-years) [
      • Superko HR
      • Pendyala L
      • Williams PT
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      • Garrett BC.
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      ]. The subfractions of HDL were not measured as they have not shown an added benefit in the identification of persons at risk above the measurement of standard HDL.
      The main strength of the study is in its’ aim to bridge the gap in knowledge regarding the impact of HRT on surrogate markers of CVD through an analysis of both traditional markers and cfPWV in this cohort of women. The study, despite highlighting several findings is not without its limitations.
      The main caution that needs to be considered when interpreting non-invasively measured cfPWV readings is the calculation of path length between the carotid and femoral sites which can significantly influence PWV, with differences of up to 30% being cited. The distance is commonly measured by one of four different methodologies: (i) the direct distance between the carotid and femoral sites; (ii) the distance between the sternal notch and femoral sites; (iii) the subtracted distance between the carotid and sternal notch from the total distance; or (iv) the subtracted distance between the carotid and sternal notch from the sternal notch and femoral site [
      • Sugawara J
      • Hayashi K
      • Yokoi T
      • Tanaka H.
      Carotid-Femoral Pulse Wave Velocity: Impact of Different Arterial Path Length Measurements.
      ]. The expert consensus document on the measurement of aortic stiffness recommended using 80% of the direct distance between the common carotid artery and common femoral artery [
      • Van Bortel LM
      • Laurent S
      • Boutouyrie P
      • Chowienczyk P
      • Cruickshank JK
      • De Backer T
      • Filipovsky J
      • Huybrechts S
      • Mattace-Raso FUS
      • Protogerou AD
      • Schillaci G
      • Segers P
      • Vermeersch S
      • Weber T
      on behalf of the Artery Society, the European Society of Hypertension Working Group on Vascular Structure and Function and the European Network for Noninvasive Investigation of Large Arteries. Expert consensus document on the measurement of aortic stiffness in daily practice using carotid-femoral pulse wave velocity.
      ]. This study, however, utilised the distance between the sternal notch and femoral site to calculate the distance between the two points as per the manufacturer's instructions for the device, which is largely dependent on body habitus, potentially introducing an error in the PWV estimation.
      Furthermore, the PWV measurements were undertaken at different points within the cyclical hormonal medication phases of oestrogen only or oestrogen and progesterone combined. Each visit occurred after a set period plus or minus two weeks to allow flexibility for the trial subjects. Hormonal influences can occur directly on the arterial wall physiology, affecting blood pressure and vascular reactivity through oestrogen driven activation of endothelial nitric oxide synthase activity. Adkisson et al., (2010) [
      • Adkisson EJ
      • Casey DP
      • Beck DT
      • Gurovich AN
      • Martin JS
      • Braith RW.
      Central, peripheral and resistance arterial reactivity: fluctuates during the phases of the menstrual cycle.
      ] postulated cyclic variations within the haemodynamic parameters, with lower central and systemic blood pressure (approximately 4mmHg) readings in the late follicular and early luteal phases of the menstrual cycle mirrored by a proportionate increase in oestrogen levels at this time, and a subsequent increase in the bioavailability of nitric oxide [
      • Adkisson EJ
      • Casey DP
      • Beck DT
      • Gurovich AN
      • Martin JS
      • Braith RW.
      Central, peripheral and resistance arterial reactivity: fluctuates during the phases of the menstrual cycle.
      ]. The progestogen component may antagonise the oestrogen mediated responses.
      The main limitation is the small sample size which did not achieve the intended power calculation. The number of recruits who voluntarily decided to leave the study or were lost to follow up, however, appears consistent across the two treatment arms; this, however, does not detract from needing larger studies to demonstrate significance. Furthermore, the loss of subjects is unlikely to be solely attributable to the widely reported progestogen related adverse effects as loss is seen across both treatment arms. The largest fall in numbers occurred after the initial 3-month period. This needs to be considered when interpreting the findings.
      The open-label design means that neither the participants nor the investigators were blinded to the study arms. This was due to the limitation in funding that restricted the option of packaging the medication identically.

      5. Conclusion

      MP combined with t-E2 was shown to have a more favourable effect on traditional surrogate markers of CVD risk in women with EMPOI, with changes observed in CO, DBP and TPR, when compared to MPA + t-E2. No significant differences, however, were noted when the cardiogenic biomarker, cfPWV, was assessed. Further studies are needed to establish reference values for cfPWV to help guide assessment of CVD risk and evaluate the response to hormone replacement in women with EMPOI.

      Contributors

      Monica Mittal conducted the study and analysed the data.
      Carmel McEniery checked the manuscript for intellectual content.
      Prasanna Raj Supramaniam analysed the data and checked the manuscript for intellectual content.
      Linda Cardozo checked the manuscript for intellectual content.
      Mike Savvas checked the manuscript for intellectual content.
      Nick Panay checked the manuscript for intellectual content.
      Haitham Hamoda formulated the hypothesis, oversaw the manuscript, and checked the manuscript for intellectual content.
      All authors approved the final submission.

      Funding

      This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

      Ethical approval

      Study approval was obtained by the Research and Development Department at King's College Hospital and ethical approval was granted by the London and GTAC Ethics Committee (REC Number: 12/LO/1957; EudraCT Number: 2012-004511-30 https://www.clinicaltrialsregister.eu/ctr-search/search?query=2012-004511-30) on the 16th January 2013. All patients gave informed written consent prior to data collection.

      Provenance and peer review

      This article was not commissioned and was externally peer reviewed.

      Research data (data sharing and collaboration)

      There are no linked research data sets for this paper. Data will be made available on request.

      Declaration of competing interest

      The authors declare that they have no competing interests.

      Acknowledgements

      I would like to extend my gratitude to all respondents, without whose cooperation I would not have been able to conduct this study.

      Appendix. Supplementary materials

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