One of the authors (Michael Schär) is employed by a commercial company (Philips Healthcare). This affiliation does not alter the PLOS ONE policies on sharing data and materials, as detailed in the online guide for authors.
Software development: M. Schär M. Stuber JY. Conceived and designed the experiments: SK M. Stuber GG AH RW GH. Performed the experiments: AH M. Schär GH M. Stuber RW JY SK. Analyzed the data: AH SK M. Stuber GH RW GG. Contributed reagents/materials/analysis tools: M. Schär M. Stuber JY RW GG. Wrote the paper: AH SK M. Stuber RW GG.
Our objective is to test the hypothesis that coronary endothelial function (CorEndoFx) does not change with repeated isometric handgrip (IHG) stress in CAD patients or healthy subjects.
Coronary responses to endothelial-dependent stressors are important measures of vascular risk that can change in response to environmental stimuli or pharmacologic interventions. The evaluation of the effect of an acute intervention on endothelial response is only valid if the measurement does not change significantly in the short term under normal conditions. Using 3.0 Tesla (T) MRI, we non-invasively compared two coronary artery endothelial function measurements separated by a ten minute interval in healthy subjects and patients with coronary artery disease (CAD).
Twenty healthy adult subjects and 12 CAD patients were studied on a commercial 3.0 T whole-body MR imaging system. Coronary cross-sectional area (CSA), peak diastolic coronary flow velocity (PDFV) and blood-flow were quantified before and during continuous IHG stress, an endothelial-dependent stressor. The IHG exercise with imaging was repeated after a 10 minute recovery period.
In healthy adults, coronary artery CSA changes and blood-flow increases did not differ between the first and second stresses (mean % change ±SEM, first vs. second stress CSA: 14.8%±3.3% vs. 17.8%±3.6%, p = 0.24; PDFV: 27.5%±4.9% vs. 24.2%±4.5%, p = 0.54; blood-flow: 44.3%±8.3 vs. 44.8%±8.1, p = 0.84). The coronary vasoreactive responses in the CAD patients also did not differ between the first and second stresses (mean % change ±SEM, first stress vs. second stress: CSA: −6.4%±2.0% vs. −5.0%±2.4%, p = 0.22; PDFV: −4.0%±4.6% vs. −4.2%±5.3%, p = 0.83; blood-flow: −9.7%±5.1% vs. −8.7%±6.3%, p = 0.38).
MRI measures of CorEndoFx are unchanged during repeated isometric handgrip exercise tests in CAD patients and healthy adults. These findings demonstrate the repeatability of noninvasive 3T MRI assessment of CorEndoFx and support its use in future studies designed to determine the effects of acute interventions on coronary vasoreactivity.
Coronary responses to endothelial-dependent interventions are important measures of vascular risk, predicting early and late cardiovascular events
Sequential studies allowing paired comparisons of coronary artery area and blood flow responses to endothelial-dependent stresses before and following an acute intervention are used to assess the endothelial response to that intervention. Because coronary endothelial function may change over a short time period (minutes to hours) in response to environmental stimuli or intervention
The protocol was approved by the Institutional Review Board at The Johns Hopkins School of Medicine and all participants provided written informed consent. No subject had a contraindication to MRI. Healthy subjects were those under age 50 years without a known history of CAD and traditional CAD risk factors, and for those over age 50 years, an Agatston coronary artery calcium score <10
MRI was performed in the morning in the fasting state before administration of any prescribed vasoactive medications. A diagram illustrating MRI study flow with measured parameters is shown in
Hemodynamic parameters have been measured at all time-points (blood pressure and heart rate).
In image (
Baseline imaging at rest for cross-sectional coronary artery area measurements
A commercial human 3.0 Tesla (T) whole-body MR scanner (Achieva, Philips, Best, NL) with a 6-element cardiac coil for signal reception was used. Cross-sectional anatomical
Images were analyzed for cross-sectional area changes using a semi-automated software tool (Cine version 3.15.17, General Electric, Milwaukee, WI, USA). A circular region-of-interest was manually traced around the coronary artery in diastole during a period of least coronary motion. The computer algorithm employed an automated full width half maximum algorithm for the cross-sectional coronary area measurements.
For flow measurements, images were analyzed using commercially available software (FLOW Version 3.0, Medis, NL). Peak diastolic coronary flow velocity was used for the velocity measurements and coronary artery blood-flow was calculated (and converted to the units mL/minute) using the adapted equation: coronary artery cross-sectional area x coronary artery peak diastolic velocity x 0.3
Statistical analysis was performed using SPSS 18.0 for Windows (SPSS Inc). Data are expressed as mean ± standard error. Proportions were compared using chi-square tests. Paired Student’s t-tests were used to compare stress coronary artery cross-sectional area, diastolic coronary flow velocity and blood-flow measurements to the initial baseline measurements obtained prior to stress, and to compare changes in all three parameters between the first and second stress. Student’s unpaired t-tests were used to compare the changes from rest to stress in coronary cross-sectional area, peak diastolic coronary flow velocity, and blood-flow measurements between the healthy and CAD subjects. The data were tested for normality using the Shapiro-Wilk test and the results indicated that parametric testing was appropriate. The Bland-Altman method was used to assess interobserver and intraobserver agreement for area, peak diastolic velocity and coronary blood-flow measurements with p-values derived from Pitman’s test of differences. Statistical significance was defined as a two-tailed p-value <0.05.
Seventeen of twenty healthy subjects (85%) and eleven of twelve CAD patients (92%) completed the study with adequate image quality. Three healthy subjects were excluded due to broken coil (N = 1), non-diagnostic image quality because of bulk movement (N = 1) and incomplete study due to shoulder pain (N = 1). One CAD patient was excluded because of non-diagnostic image quality. Thirty coronary artery segments in 17 healthy subjects and 15 coronary artery segments in 11 CAD patients were evaluable for analysis (
A scout scan obtained parallel to the left anterior descending (LAD) artery (
Characteristics | Healthy Subjects N = 17 | CAD Patients N = 11 | P value |
Age (yr), Mean ±SD | 31±10 | 57±6 | <0.001 |
Gender (male) | 8 (47) | 8 (73) | NS |
Previous MI | 0 | 5 (45) | 0.016 |
PCI/stent | 0 | 7 (64) | 0.002 |
CABG | 0 | 1 (9) | 0.34 |
CAD risk factors |
|||
History of smoking | 0 | 3 (27) | 0.08 |
Dyslipidemia | 0 | 9 (82) | <0.001 |
Diabetes mellitus | 0 | 0 | NS |
Hypertension | 0 | 8 (73) | <0.001 |
Family history of early CAD | 0 | 3 (27) | 0.08 |
Vessel/s studied | |||
RCA | 15 | 7 | 0.84 |
LAD | 15 | 5 | 0.30 |
LCX | 0 | 3 | 0.08 |
Total vessels studied | 30 | 15 |
Abbreviations; SD = standard deviation, CAD = coronary artery disease, PCI = percutaneous coronary intervention, CABG = coronary artery bypass graft surgery, MI = myocardial infarction, RCA = right coronary artery, LAD = left anterior descending coronary artery, LCX = left circumflex coronary artery.
CAD risk factors excluding age and gender.
IHG exercise caused a significant hemodynamic effect in both groups. In healthy subjects, the baseline rate pressure product (RPP, heart rate x systolic blood pressure) of 8437±346 mmHg*beats/minute increased to 10,471±515 mmHg*beats/minute with the first stress (p<0.0001 vs. baseline). RPP increased similarly during the second stress in healthy subjects (
* signifies p<0.05 compared to baseline RPP. Error bars indicate standard error of the mean.
In the healthy group, coronary arteries dilated significantly during the first IHG stress (baseline cross-sectional area: 10.1±0.5 vs. first stress: 11.6±0.7 mm2, p<0.0001) and second stress: 11.9±0.7 mm2, p<0.0001). There was no significant difference in % cross-sectional area (CSA) change with IHG between the first and second stress (% increase in mean CSA with stress 1∶14.8% ±3.3% vs. stress 2∶17.8% ±3.6%, p = 0.24). In contrast to the increase in CSA in the healthy group, CSA decreased with the first and second stresses in the CAD group (baseline area: 14.0±1.1 vs. stress area 1∶13.1±1.0 mm2, p = 0.005, n = 15), and second stress: 13.3±1.0 mm,2 p = 0.06), although again the percent change in mean CSA during the two stresses did not differ from one another (−6.4% ±2.0% vs. −5.0% ±2.4% for the first and second studies respectively, p = 0.22). In the healthy group, the coronary artery area measured just before the second exercise period was similar to that measured at baseline (baseline cross-sectional area: 10.1±0.5 mm2 vs. pre-exercise 2∶10.3±0.6 mm2, p = 0.51), while in the CAD group, the coronary artery area was lower before the second exercise period as compared to baseline (baseline cross-sectional area: 14.0±1.1 mm2 vs. pre-exercise 2∶13.1±1.0 mm2, p = 0.01). Importantly, there was a significantly different response between healthy subjects and CAD patients in terms of direction and magnitude of coronary vasoreactivity to IHG stress (healthy CSA change (stress 1): 14.8% ±3.3% vs. CAD area change (stress1): −6.4% ±2.0%, p<0.0001). The relative stress-induced area changes in both groups are shown in
Error bars indicate standard error of the mean. In the healthy group, a normal coronary endothelial response is seen with an increase in coronary artery area, velocity and flow with stress, and no significant difference between stress 1 and stress 2 response. In the CAD group, there is an abnormal coronary endothelial response with no increase or decrease in the same three parameters with stress, and no significant difference in response between stress 1 and 2.
Peak diastolic coronary flow velocity increased in healthy subjects during the first and second stresses (20.7±0.9 cm/s baseline vs. 26.4±1.3 cm/s and 25.7±1.1 cm/s, for the first and second studies respectively, p<0.0001 vs. baseline) with no significant difference in the percent velocity change between the two tests (p = 0.54). There was no significant change in peak diastolic flow velocity with stress for CAD subjects (baseline vs. stress1∶20.0±1.4 cm/s vs. 19.2±1.5 cm/s, p = 0.42; and vs. stress2∶19.2±1.4 cm/s, p = 0.53). In the healthy and CAD groups, there was no significant difference in velocity values between the baseline and pre-exercise 2 measurements (healthy: baseline velocity: 20.7±0.9 cm/s vs. pre-exercise 2∶20.1±0.7 cm/s, p = 0.28; CAD: baseline velocity: 20.0±1.4 cm/s vs. pre-exercise 2∶18.4±1.0 cm/s, p = 0.09).
Coronary blood flow increased significantly with IHG stress in healthy subjects and decreased in CAD patients (healthy flow change (stress 1): 44.3% ±8.3% vs. CAD flow change (stress1): −9.7% ±5.1%, p<0.0001). In healthy subjects, coronary blood-flow increased significantly with isometric handgrip during both stress periods (baseline: 63.2±4.6 ml/minute vs. stress 1∶91.2±6.2 ml/minute, p<0.0001, and vs. stress 2∶91.5±5.9 ml/minute, p<0.0001). In CAD patients, blood-flow did not increase, but decreased slightly with the first and second stresses, although not significantly (baseline: 83.9±9.7 ml/minute vs. stress 1∶75.8±8.0 ml/minute, p = 0.13, and vs. stress 2∶76.6±7.0 ml/minute, p = 0.40). In the healthy group, the coronary flow measured pre-exercise 2 was similar to the baseline value (baseline flow: 63.2±4.6 ml/minute vs. pre-exercise 2 flow: 63.1±5.5 ml/minute, p = 0.93). In the CAD group, the pre-exercise 2 coronary flow did not return to the original baseline value (baseline flow: 83.9±9.7 ml/minute vs. pre-exercise 2 flow: 69.6±5.3 ml/minute, p = 0.03). Relative to baseline coronary blood-flow, changes with stress were not significantly different between stress 1 and stress 2 in either the healthy subjects (stress 1 flow change: 44.3% ±8.3% vs. stress 2∶44.8% ±8.1%, p = 0.84) or the CAD patients (−9.7% ±5.1% vs. stress 2: −8.7% ±6.3%, p = 0.38). Relative changes in velocity and flow for both groups are shown in
The intra-observer results for area and velocity measurements showed no significant differences (p = 0.10 and p = 0.70 respectively). Similarly, the inter-observer variability for the area and velocity measurements did not show significant differences (p = 0.68 and p = 0.63 respectively) similar to that previously reported
Bland-Altman plots for intra-observer variability (A and C) and inter-observer variability (B and D) of coronary artery cross-sectional area (A and B) and peak diastolic flow velocity (C and D) measurements in CAD patients and healthy subjects. Solid lines above and below the mean represent ±2 standard deviations and the mean differences are shown. P-values are derived from Pitman’s test of differences.
3T MRI was performed at rest and during sequential isometric handgrip exercise, an established endothelial-dependent stressor. IHG exercise caused significant hemodynamic effects in healthy and CAD subjects during both stress periods. The coronary endothelial response to stress in the healthy group, as expected, was marked by vasodilation and increased flow. The responses during the first and second stress periods did not differ. In the CAD group, the coronary endothelial responses during the first and second stresses were abnormal with a lack of vasodilation and decreased flow, and these did not differ from the first stress to the second. Therefore, when compared to the unperturbed state (baseline 1), the coronary vasoactive responses to IHG exercise are similar between two successive exercise sessions for both healthy subjects and patients with CAD. However with this protocol where the second IHG exercise commenced 10 minutes after the first, the second pre-exercise coronary indices had not returned to baseline values (those prior to first IHG) in CAD patients, although they did in healthy volunteers. In future studies which may investigate the role of an intervention, it is critical to compare the two IHG responses to the true baseline, unperturbed state. A longer recovery period between the two successive stress intervals could also be investigated in future studies.
The values for coronary endothelial function reported here are similar to those previously reported using MRI
In animal studies of coronary arteries, there is evidence that short term exercise training enhances nitric oxide (NO)-mediated coronary dilation
Thus, the non-invasive MRI technique described here is particularly suitable for evaluating asymptomatic populations and for performing repeated studies in low risk individuals. Although PET can be used to assess coronary blood flow in response to endothelial stressors
One limitation to this study is that we did not compare MRI-derived measures of coronary vasoreactivity with those obtained using invasive methods such as coronary angiography or Doppler guidewire. As many of the subjects were healthy, an invasive coronary test was not clinically indicated and could not be justified. Moreover, MRI measures of coronary area
In summary, we report that coronary endothelial function measured non-invasively using MRI does not change with repeated isometric handgrip exercise over the short term in both healthy subjects and those with CAD when compared to the baseline unperturbed state. This ability to non-invasively characterize the coronary endothelial responses to repeated IHG exercise coupled with the reproducibility of the results and the short time required for the MRI protocol may facilitate the design of future studies targeting coronary endothelial responses to acute interventions and contribute to the non-invasive characterization of factors that affect vascular function.
The authors thank Angela Steinberg, RN and Rob van der Geest, PhD for their assistance, and the patients and healthy volunteers for their participation in this study.