The Effects of Menstrual Cycle Phase on Exercise Treadmill Test Results in Premenopausal Women

General Hypothesis:

False positive results are commonly observed in women after exercise treadmill test (ETT). The studies report severe variability in the sensitivity and specificity of ETT in detecting significant coronary artery disease (CAD). Sensitivity ranges from 23% to 100% and specificity ranges from 17% to 100%. Exercise Treadmill Test utilizes electrocardiography (ECG) along with heart rate (HR) and blood pressure (BP) monitoring during exercise, typically involving a treadmill. The major finding of Exercise Treadmill Test is the ST segment deviation from the baseline. False positive stress ECGs are common especially in premenopausal women, limiting the specificity of the test.

Menstrual periods can cause biological variability in Exercise Treadmill Test in premenopausal women. The menstrual period is divided into phases: early follicular (days 1-5), late follicular (days 6-12), ovulation (days 13-15), early luteal (days 16-19), mid-luteal (days 20-23) and late luteal phases (24-28 days). Early follicular phase is characterized by low estrogen and low progesterone, late follicular phase is characterized by low progesterone and high estrogen. Mid-luteal phase is characterized by high progesterone and high estrogen levels. Early and late follicular phases of the menstrual cycle in women demonstrate nearly 4 fold differences in estrogen levels. The menstrual period phase affects thermoregulation and physical working capacity in women. Estrogen has protective and vasodilatory effects on the cardiovascular system. With the discovery of high sensitive cardiac troponins (hs-cTn), minute changes of hs-cTn have become detectable.

The effects of menstrual periodic changes on the results of exercise treadmill test in premenopausal women have not been clearly defined. Primary purpose of the study is to investigate the biological causes of false positive test results in the treadmill exercise test in premenopausal women. Estrogen is known to have direct vasodilatory effects on coronary arteries. Early and late follicular phases of the menstrual cycle are characterized by low and high estrogen levels, respectively. The Early Follicular Phase starts on the first day of the menstrual cycle and ends when oestradiol begins to increase. It is characterised by increasing LH and FSH and constant low levels of oestradiol. The late follicular phase starts with the increase in oestradiol and ends at its preovulatory peak. False positive stress ECGs are commonly reported especially in premenopausal women, limiting the specificity of the Exercise Treadmill Test. Studies demonstrated that HR adjustment of in exercise-induced ST segment depression using the ST/HR slope or ST/HR index could improve the sensitivity and specificity of Exercise Treadmill Test.

The hypothesis of the study is that Exercise Treadmill Test results and hs-cTnT release after Exercise Treadmill Test will change at the early and late follicular phases in premenstrual women.

1. STUDY ENDPOINTS

Primary endpoint: The primary endpoint in the study was the ST/HR index.

ST segment depression on electrocardiography (ECG) (1 mm equals 0.1 mV or 100 microvolts at standard calibration) and heart rate (HR) (beats per mınute) will be combined to report ST/HR index by dividing maximum ST-segment depression by the change in heart rate (HR) from rest to peak exercise.

(Indices of ST/HR slope and the ST/HR index were reported automatically by the General Electronic GE Healthcare T2100-ST Treadmill \& CASE 6.73 Stress Test System device.)

Secondary endpoints:

1. The maximum ST segment depression (mm) on electrocardiography (ECG) during the exercise treadmill test (1 mm equals 0.1 mV or 100 microvolts at standard calibration)
2. The maximum heart rate (HR) (beats per minute) during the exercise treadmill test
3. Change in maximal exercise capacity with exercise and menstrual cycle (METs score)
4. maximum ST segment depression (mm)
5. ST/HR slope (µV/beats per minute (bpm))
6. The concentraion of high-sensitive cardiac troponin T after exercise treadmill test (ng/L)
7. Hormone levels (estrogen) at the early and late follicular phases of the menstrual cycle
8. STUDY DESIGN The study is performed in the cardiology clinics of a tertiary care training hospital. The study is approved by the local ethics committee (number 2023/0129 on 22/02/2023).

Study Population:

The study population consists of premenopausal women who applied to the outpatient clinic with chest pain between February 2023 and July 2023. Patients report that they have a regular menstrual cycle. Written informed consent is obtained from the patients.

All patients have transthoracic echocardiography. Echocardiography demonstrates normal left ventricular systolic and diastolic function. Echocardiography does not show wall motion abnormality or significant valvular heart disease.

All patients are evaluated for cardiovascular risk. Risk factors such as hypertension, diabetes mellitus, cigarette smoking, hyperlipidemia, family history of CAD, and drug use are questioned. Cardiovascular risk factors are defined as follows. Hypertension is defined as a persistent elevation in office systolic BP ≥140 and/or diastolic BP ≥90 mmHg, or a 24-hour ambulatory BP monitor average of ≥130/80, or requirement of chronic use of antihypertensives. Hyperlipidemia is defined as elevated total cholesterol (>240 mg/dl), low-density lipoprotein cholesterol (LDL-C) (>160 mg /dl), triglycerides > 150 mg/dl, high-density lipoprotein cholesterol (HDL-C) (< 50 mg /dl). Cigarette smoker is defined as having smoked at least 100 cigarettes in life time. Type 2 diabetes mellitus is defined as fasting plasma glucose (FPG) ≥ 126 mg/dL, or 2-hour plasma glucose ≥ 200 mg/dL during a 75-g oral glucose tolerance test (OGTT), or random plasma glucose ≥ 200 mg/dL in a patient with classic symptoms of hyperglycemia, or hemoglobin A1c (HbA1c) ≥ 6.5% (48 mmol/mol). Self-reported family history of premature CAD is defined as having a first-degree relative with premature CAD (men, age <55 years; women, age <65 years).

Menstrual cycle The history of the menstrual cycle of the patient is obtained in detail. The early and late follicular phases are determined from the history.

The early follicular phase Exercise Treadmill Test is planned on the 3rd day of menstrual bleeding, while the menstrual bleeding continues. The late follicular phase Exercise Treadmill Test is planned on the 3rd day after the menstrual bleeding ends. Hormone levels are measured before each Exercise Treadmill Test to ascertain the phase of the menstrual cycle. Two Exercise Treadmill Tests are performed on each participant.

The duration and regularity of the menstrual cycle, the amount of bleeding, and the number of days the bleeding lasted are noted for each patient. Patients with irregular menstrual cycles, breakthrough bleeding, spotting, gynecological examination, or treatment plan that would disrupt the menstrual cycle are excluded from the study.

The blood sample is taken 30 minutes before the Exercise Treadmill Test to measure follicular stimulating hormone (FSH), luteinizing hormone (LH), estrogen, progesterone, and hs-cTn. After the test, the patient is allowed to rest for 15 minutes and blood is taken to check the 2nd hs-cTn after Exercise Treadmill Test.

The inclusion criteria are:

1. Premenopausal female patients with regular menstrual cycles,
2. Ages between 20-55 years,
3. Patients have been seen in the cardiology clinic by a cardiologist for the symptoms of chest pain with a clinical suspicion of angina or angina equivalent.
4. A cardiologist has ordered Exercise Treadmill Test as a diagnostic test independent of the study protocol
5. Patients can understand and sign the informed consent.

The exclusion criteria are:

1. Women with menopause or menopause-related symptoms or menstrual cycle irregularity or use of hormonal therapy or oral contraceptives, current pregnancy
2. Patients with a history of CAD, secondary or uncontrolled hypertension, valvular heart disease, cardiac arrhythmia or heart failure, pericarditis or myocarditis.
3. Left ventricular hypertrophy on echocardiography, abnormal systolic or diastolic function or wall motion abnormality on echocardiography.
4. History of cancer, stage ≥ 3 kidney failure, liver failure, history of psychiatric illness, history of recent infection.
5. Pre-existing ECG abnormality that would confound the interpretation during the Exercise Treadmill Test (baseline ECG was considered abnormal ın the presence of the following findings: prolonged QT interval, chamber hypertrophy, alterations in the ST-T waves, any degree of heart block, T-wave abnormalities, premature ventricular contractions (PVCs), right bundle branch block (RBBB), left bundle branch block (LBBB), left or right axis deviation, or any abnormal rhythm other than normal sinus rhythm).
6. Patients who cannot tolerate Bruce protocol in the first Exercise Treadmill Test.
7. Lack of ability or mental capacity to understand the study.