Methods

Twenty-one patients with stable cardiovascular disease were identified and randomly selected from a known Johns Hopkins research database of patients with heart disease. These patients had been genotyped for selected polymorphisms of β1 and β2 receptors. Patients were ineligible for enrollment if they had class >II angina or heart failure, a recent revascularization procedure, symptomatic ventricular or atrial arrhythmias, or significant hypokalemia (<3.0 mEq/L) within the 6 months before enrollment. All were on stable doses of cardiovascular medications including β-blocker therapy and medications were not withheld before this outpatient study protocol. The Johns Hopkins Medicine institutional review board approved the study and all patients signed written informed consent.

After baseline clinical examination, transthoracic echocardiography, and gated blood pool scan imaging, each subject received intravenous dobutamine followed by intravenous terbutaline infusions. Because the 1/2-life for dobutamine is 2 minutes and the 1/2-life for terbutaline is several hours, all subjects received dobutamine first followed by ≥20 minutes of washout and until heart rate (HR) and blood pressure returned to baseline. After a second set of identical baseline measurements, intravenous terbutaline was initiated.

Dobutamine (Novaplus Pharmaceuticals, Irving, Texas) was infused through an intravenous cannula in the forearm starting at a dose of 5 μg/kg/min for 5 minutes to achieve a stable response and then increased to a maximum dose of 10 μg/kg/min. At this dose, previous invasive studies have demonstrated that contractility in subjects without structural heart disease and subjects with cardiomyopathy will increase >30% above baseline.

Terbutaline (Novaplus Pharmaceuticals) was infused starting at a dose of 50 ng/kg/min, which after 5 minutes was increased to 100 ng/kg/min for 5 minutes, and then to a maximal dose of 150 ng/kg/min. At this dose, HR increases 25 to 30 beats per min in healthy subjects. The blood pressure, HR, and electrocardiogram were monitored continuously during the catecholamine infusions.

Measurements of LV volumes and ejection fraction (EF) were performed using nuclear gated blood pool imaging before the initiation of dobutamine, at peak dobutamine infusion, during the second baseline period 20 to 30 minutes after termination of dobutamine, and during peak terbutaline infusion. After in vivo labeling of red blood cells with 25 to 30 mCi of technetium-99m, subjects were imaged supine in the 40° left anterior oblique position as previously described. Scans were interpreted by a single investigator (L.C.B.) blinded to subject identity and experimental conditions (i.e., baseline vs at peak catecholamine infusion). Time–activity curves using LV regions of interest were used to calculate LV volumes and EF. LV volumes were quantified using a count-based method with individual correction for chest wall attenuation. EF was calculated by the formula: (end-diastolic volume minus end-systolic volume) multiplied by 100 divided by end-diastolic volume.

Using commercially available ultrasound systems, all patients underwent a limited Doppler echocardiographic study at the 2 baseline periods and during catecholamine infusion to measure peak aortic flow.

Genomic DNA was extracted from peripheral blood leukocytes using a Gentra Autopure LS workstation (Qiagen, Valencia, California) that uses the Gentra Puregene DNA isolation kit chemistry. DNA was quantified by spectrophotometry using a DU 530 Life Science UV/VIS Spectrophotometer (Beckman Coulter, Inc., Fullerton, California). TaqMan single-nucleotide polymorphism genotyping assays (Applied Biosystems, Foster City, California) were used to genotype 2 nonsynonymous single-nucleotide polymorphisms in the β2-adrenergic receptor gene (rs1042714 C/G, corresponding to glutamine or glutamic acid at position 27, and rs1042713 A/G, corresponding to arginine or glycine at position 16). There were 2 nonsynonymous single-nucleotide polymorphisms in the β1-AR gene (rs1801252 A/G, resulting in serine or glycine at position 49, and rs1801253 C/G, corresponding to arginine or glycine at position 389). Genotype assignments were made by measuring fluorescence using an ABI Prism 7900HT Sequence Detection System and ABI Prism 7900 allelic discrimination software (Applied Biosystems, Foster City, California).

Subject data at the 2 baselines and at peak infusions were analyzed before and after stratification by genotype. Data were compared using unpaired and paired Student’s t tests, chi square test, or analysis of variance as appropriate. Nonparametric analysis using Kruskal–Wallis and Wilcoxon signed-rank sum testing were performed on those data that were not normally distributed or for small samples. Dichotomous variables are summarized using ratios and continuous outcomes are reported as mean ± SD. Given our previous data showing that patients homozygous for the Glu27 allele had adverse LV remodeling after ST-segment elevation infarction, the a priori hypothesis was that cardiac function would differ in those patients homozygous for the β2 Glu27 allele versus those with a glutamine allele. We assessed cardiac function by 2 different measurements, cardiac output (CO) and peak power index. Peak power was defined as maximal power (approximated by the product of peak aortic flow by 2-dimensional echocardiography and systolic blood pressure) divided by end-diastolic volume. A sample size of 20 patients was needed to detect a 40% difference in CO with a power of 80% and an alpha of 0.05. All statistical analyses were conducted using STATA 11.1 (STATA Corp., College Station, Texas). All p values ≤0.05 were considered statistically significant.

Because of the a priori hypothesis, parameters were compared using a recessive modeling strategy contrasting those having ≥1 major allele and those with 2 minor alleles for the β2 gene polymorphisms. This strategy was maintained for the β1 gene polymorphisms as previous data have shown that patients with Arg389Arg and Arg389Gly tend to have a greater response in EF compared to those with Gly389Gly.

Results

Fifty patients were contacted for participation in the study. Of these, 21 were enrolled. The others were excluded because of nonresponse (n = 20) and subject refusal (n = 9). In 1 patient, the dobutamine infusion was discontinued during nuclear ventriculography because of symptoms of lightheadedness. HR and blood pressure were within normal parameters at the time. The patient completed the terbutaline portion of the study and was included in the analysis. All other patients completed the protocol without deviation or adverse events.

Cause of cardiovascular disease and dose of β-blocker therapy were not associated with genotype assignment, baseline hemodynamic measurements, or response to stimulation by agonists based on genotype. All alleles in the source population were in Hardy–Weinberg equilibrium and polymorphism frequencies in that population agreed with previous studies. In the study population, the minor allelic frequencies were 0.10 for Gly49 and 0.37 for Gly389 (β1 gene) and 0.30 for Arg16 and 0.48 for Glu27 (β2 gene).

There were no differences in age, gender, co-morbid conditions, blood pressure, or LV structure or function between those with 1 Arg389 allele or 2 Arg389 alleles and those homozygous for glycine at codon 389 ( Table 1 ). HRs at rest were similar between the 2 groups, suggesting a similar dose–effect of β-blocker therapy.

In response to dobutamine and terbutaline, there were statistically significant increases in HR, stroke volume (SV), CO, EF, and peak power from baseline to peak infusion of drugs in patients who had ≥1 Arg389 allele ( Table 2 ). Figure 1 shows the differential cardiovascular response to the β1 agonist dobutamine based on polymorphism. Those patients possessing ≥1 Arg389 allele had a greater change in HR and CO compared to those homozygous for Gly389. Although the peak power index increased in response to the 2 agonists, there was no differential response by polymorphism. There was no differential response to administration of terbutaline based on β1 Arg389Gly genotype. Cardiovascular responses to dobutamine and terbutaline did not differ between subjects with polymorphisms of the β1 Ser49Gly genotype.

Response of heart rate (left) and cardiac output (right) to dobutamine based on β1-receptor genotype.

Table 1 lists subject demographics by β2 Glu27 genotype. Similar to baseline data for the β1 receptor, there were no differences in age, gender, co-morbid conditions, clinical vital signs, or LV structure or function when comparing patients with Glu27Glu alleles to those with 1 glutamine allele or 2 glutamine alleles ( Table 1 ) or Gly16 homozygotes (n = 10) to those with ≥1 Arg16 allele (n = 11, data not shown).

HR, SV, CO, and EF increased and end-systolic volume decreased in response to the dobutamine and terbutaline infusions ( Table 3 ). Cardiovascular responses to dobutamine and terbutaline did not differ by β2 Gly16Arg genotype. There were also no significant associations between cardiovascular responses to dobutamine and β2 Glu27Gln genotype. In contrast, the Gln27Glu single-nucleotide polymorphism was a determinant of chronotropic and inotropic responses to intravenous terbutaline ( Figure 2 ). Change in HR from baseline to peak terbutaline infusion was greater in those with the Glu27Glu genotype compared to those patients with 1 glutamine allele or 2 glutamine alleles. In addition, the SV change from baseline to terbutaline was greater in glutamic acid homozygotes. This increase in SV was mainly the result of a decrease in end-systolic volume (data not shown). Patients with Glu27Glu had an approximately 1.5 L/min greater increase in CO compared to all others.

Table 3

Phenotypic responses before and during agonist stimulation based on beta-2–receptor genotype

	Baseline	Peak Infusion	Change	p Value ⁎
Glu27Glu (n = 6)
Dobutamine
Heart rate (beats/min)	64	79	15 ± 11	0.02
Stroke volume (ml)	98	108	10 ± 14	NS
Cardiac output (L/min)	6.2	8.6	2.4 ± 1.6	0.01
Ejection fraction (%)	44	50	6 ± 5	0.02
Peak power index	12.3	16.3	4.0 ± 4.7	NS
End-systolic volume (ml)	136	116	−20 ± 23	NS
End-diastolic volume (ml)	234	224	−10 ± 25	NS
Systemic vascular resistance (dynes × s/cm 5 )	1,260	1,423	163 ± 246	NS
Terbutaline
Heart rate (beats/min)	62	76	14 ± 9	0.01
Stroke volume (ml)	91	119	28 ± 19	0.01
Cardiac output (L/min)	6.0	9.1	3.1 ± 2	0.01
Ejection fraction (%)	42	54	12 ± 2	0.004
Peak power index	11.4	14.1	2.7 ± 1.8	0.014
End-systolic volume (ml)	141	109	−32 ± 28	0.04
End-diastolic volume (ml)	232	228	−4 ± 75	NS
Systemic vascular resistance (dynes × s/cm 5 )	1,274	1,234	−41 ± 125	NS
1 Gln27 allele or 2 Gln27 alleles (n = 15)
Dobutamine
Heart rate (beats/min)	61	70	9 ± 13	0.02
Stroke volume (ml)	91	101	11 ± 17	0.03
Cardiac output (L/min)	5.4	7.2	1.8 ± 1.1	<0.001
Ejection fraction (%)	48	56	8 ± 7	<0.001
Peak power index	14.8	21.1	6.4 ± 8.7	0.01
End-systolic volume (ml)	110	94	−17 ± 21	0.006
End-diastolic volume (ml)	201	199	−2 ± 24	NS
Systemic vascular resistance (dynes × s/cm 5 )	1,396	1,525	129 ± 287	NS
Terbutaline
Heart rate (beats/min)	63	68	5 ± 8	0.05
Stroke volume (ml)	87	101	15 ± 10	<0.001
Cardiac output (L/min)	5.3	6.9	1.6 ± 1.1	<0.001
Ejection fraction (%)	47	56	9 ± 6	<0.001
Peak power index	13.9	15.6	1.7 ± 3.1	0.05
End-systolic volume (ml)	108	90	−18 ± 17	<0.001
End-diastolic volume (ml)	195	191	−3.6 ± 14	NS
Systemic vascular resistance (dynes × s/cm 5 )	1,357	1,331	−26 ± 112	NS

Only gold members can continue reading. Log In or Register to continue

Share this:

• Click to share on Twitter (Opens in new window)
• Click to share on Facebook (Opens in new window)

Related

Related posts:

1. Outcomes of Preoperative Angiotensin-Converting Enzyme Inhibitor Therapy in Patients Undergoing Isolated Coronary Artery Bypass Grafting
2. Effect of Nicotine Replacement Therapy on Cardiovascular Outcomes After Acute Coronary Syndromes
3. Usefulness of Bidimensional Strain Imaging for Predicting Outcome in Asymptomatic Patients Aged ≤ 16 Years With Isolated Moderate to Severe Aortic Regurgitation
4. Insulin Sensitivity, but Not Hyperglycemia, May Be Superior as a Prognostic Marker and Therapeutic Target for Diabetes in Heart Failure

Stay updated, free articles. Join our Telegram channel

Join