D-dimer in Marfan syndrome: effect of obstructive sleep apnea induced blood pressure surges

Abstract

Aortic dissection and rupture are the major causes of premature death in persons with Marfan syndrome (MFS), a rare genetic disorder featuring cardiovascular, skeletal, and ocular impairments. We and others have found that obstructive sleep apnea (OSA) confers significant vascular stress in this population and may accelerate aortic disease progression. We hypothesized that D-dimer, a diagnostic biomarker for several types of vascular injury that is also elevated in persons with MFS with aortic enlargement, may be sensitive to cardiovascular stresses caused by OSA. To test this concept, we recruited 16 persons with MFS without aortic dissection and randomized them to two nights of polysomnography, without (baseline) and with OSA treatment: continuous positive airway pressure (CPAP). In addition to scoring OSA by the apnea-hypopnea index (AHI), beat-by-beat systolic BP (SBP) and pulse-pressure (PP) fluctuations were quantified. Morning blood samples were also assayed for D-dimer levels. In this cohort (male:female, 10:6; age, 36 ± 13 yr; aortic diameter, 4 ± 1 cm), CPAP eliminated OSA (AHI: 20 ± 17 vs. 3 ± 2 events/h, P = 0.001) and decreased fluctuations in SBP (13 ± 4 vs. 9 ± 3 mmHg, P = 0.011) and PP (7 ± 2 vs. 5 ± 2 mmHg, P = 0.013). CPAP also reduced D-dimer levels from 1,108 ± 656 to 882 ± 532 ng/mL (P = 0.023). Linear regression revealed a positive association between the maximum PP during OSA and D-dimer in both the unadjusted (r = 0.523, P = 0.038) and a model adjusted for contemporaneous aortic root diameter (r = 0.733, P = 0.028). Our study revealed that overnight CPAP reduces D-dimer levels commensurate with the elimination of OSA and concomitant hemodynamic fluctuations. Morning D-dimer measurements together with OSA screening might serve as predictors of vascular injury in MFS.

NEW & NOTEWORTHY What is New? Surges in blood pressure caused by obstructive sleep apnea during sleep increase vascular stress and D-dimer levels in Marfan syndrome. Elevations in D-dimer can be lowered with CPAP. What is Noteworthy? D-dimer levels might serve as a marker for determining the significance of obstructive sleep apnea in persons with Marfan syndrome. D-dimer or obstructive sleep apnea screening is a potential method to identify persons with Marfan syndrome at risk for adverse cardiovascular events.

INTRODUCTION

Marfan syndrome (MFS) is an autosomal dominant connective tissue disease that causes vascular wall weakness leading to thoracic aortic aneurysms and eventual dissection or rupture (1, 2). Efforts to mitigate aortic morbidity have focused on early detection, inhibiting transforming growth factor-β with angiotensin II-receptor blockers and reducing factors that contribute to aortic stress such as blood pressure and pulse rate surges (3, 4).

D-dimer is a fibrin degradation product released when cross-linked fibrin is cleaved by plasmin (5, 6). Thus, D-dimer elevation reveals activation of the clotting and/or fibrinolytic system and has a diagnostic role in conditions such as deep vein thrombosis, pulmonary embolism, and disseminated intravascular coagulation (7, 8). D-dimer has also been used as a marker for acute aortic dissection (9–11) and was recently shown to be elevated in patients with MFS with aortic enlargement compared with non-MFS controls without aortic enlargement (12). Another study showed that non-MFS patients with aortic aneurysm had higher D-dimer levels than age-matched controls (13). Both studies suggest that aortic structural compromise contributes to D-dimer elevation. It is not clear, however, if blood pressure swings acting on the aortic wall (14) disrupt aortic vascular integrity and thereby modulate D-dimer levels.

Because obstructive sleep apnea (OSA) frequently leads to surges in blood pressure and heart rate with each apneic or hypopneic event (15–18), the disorder might be a modifiable risk factor for aortic morbidity in MFS. Moreover, OSA has been associated with adverse aortic events in MFS (19, 20), which can be mitigated by treatments such as continuous positive airway pressure (CPAP) (21). It is plausible that acute nocturnal cardiovascular stress from OSA contributes to aortic shear stress, as reflected by elevated markers of arterial injury such as D-dimer.

Our goal was to examine the effect of OSA-induced hemodynamic fluctuations on D-dimer levels in patients with MFS. We monitored the breathing pattern and associated blood pressure changes during sleep in patients with MFS at baseline and during a single night of continuous positive airway pressure (CPAP) therapy and assayed morning D-dimer levels. We also assessed the augmentation index (AIx), an indirect indicator of arterial stiffness (22) as a measure of vascular stress, in the morning after sleep studies. We hypothesized that 1) D-dimer concentrations would be associated with hemodynamic fluctuations during OSA events at baseline; 2) elimination of OSA events with CPAP would decrease hemodynamic fluctuations and D-dimer levels; and 3) after alleviation of OSA, reduction in D-dimer would be associated with a reduction in AIx.

METHODS

Participants

We recruited consecutive self-reported snorers with MFS from the Johns Hopkins Vascular Connective Tissue Disorders Clinic and through the Marfan Foundation (Fig. 1). To be eligible for the study, the participants had to be 18 yr or older with an established diagnosis of MFS and answer “yes” to habitual snoring on the screening questionnaire. We excluded persons with aortic dissection, uncontrolled hypertension, lung disease, and other sleep disorders such as insomnia. We also excluded participants with bleeding disorders or those on long-term anticoagulation therapy. Participants also provided reports of echocardiography performed within the past year, from which we obtained their aortic root diameter. The study was approved by the Johns Hopkins Institutional Review Board, and all the participants provided written informed consent.

Figure 1.

Participants’ consort diagram.

Polysomnography

Participants underwent overnight polysomnography while sleeping with or without CPAP in a randomized crossover fashion with washout periods of 1–7 days. Polysomnography was performed according to standard American Academy of Sleep Medicine (AASM) guidelines (23). Parameters monitored included electroencephalogram, electrooculogram, submental and pretibial electromyogram, modified electrocardiogram, body position, airflow, pulse oximetry, esophageal pressure, and blood pressure. On CPAP nights, CPAP pressure was titrated until apneas and hypopneas (OSA events) were eliminated.

Polysomnography analyses.

Sleep architecture was summarized by calculating the total sleep time (TST), sleep efficiency, and percentages of TST spent in each sleep stage on baseline and CPAP nights. An obstructive apnea was defined as cessation of airflow for ≥10 s in the presence of continued respiratory effort. Hypopnea was defined as ≥30% drop in airflow for ≥10 s in association with ≥3% oxygen desaturation or arousal from sleep (23). The apnea/hypopnea index (AHI) was calculated by the sum of apneas and hypopneas divided by TST in hours. The respiratory rate was calculated as an overnight average from the airflow channel. Based on the AASM definition of arousals (23), we also calculated the total number of arousals and identified those related to changes in airflow, which we termed respiratory arousals.

Measurement of esophageal pressure.

To quantify respiratory effort, we monitored esophageal pressure (Pes) continuously with a balloon-tipped catheter placed in the middle third of the esophagus (24–26). On a breath-by-breath basis, we identified the peak inspiratory Pes (Pes_peak-insp), which is the point of maximum inspiratory effort (24, 27), and calculated the average during sleep for each participant.

Blood pressure monitoring.

We monitored beat-by-beat blood pressure during sleep with the Caretaker Medical unit (Caretaker Medical, ISO 13485, Charlottesville, VA). The device uses a low-coupling pressure finger cuff to obtain real-time blood pressure (28, 29). As OSA is known to cause surges in blood pressure and heart rate (17, 18), we quantified the degree of hemodynamic fluctuations during periods with OSA events. We identified the minimum and maximum systolic blood pressures (SBPs), pulse pressures (PPs), and heart rates (HRs) during OSA events and stable nonflow limited breathing periods with CPAP (Fig. 2). For each participant, we calculated the average for six variables of hemodynamic fluctuation, which we define below:

Figure 2.

Representative 3-min periods of obstructive sleep apnea (OSA) and continuous positive airway pressure (CPAP) treatment. The upper tracing depicts heart rate (HR), the upper middle tracing depicts airflow, the middle tracing depicts systolic blood pressure (SBP), the lower middle tracing depicts diastolic blood pressure (DBP), and the bottom tracing depicts pulse pressure (PP). BP, blood pressure; bpm, beats/min.

• SBP_max: maximum systolic blood pressure

• PP_max: maximum pulse pressure

• HR_max: maximum heart rate

• ΔSBP: difference between maximum and minimum SBP

• ΔPP: difference between the maximum and minimum PP

• ΔHR: difference between the maximum and minimum HR

D-Dimer Assay and Assessment of AIx

In the morning after sleep studies, we collected venous blood to assay D-dimer levels. We also measured the AIx, an indirect indicator of arterial stiffness with the Endo-PAT2000 (Itamar Medical) (30). The EndoPAT 2000 assesses AIx by quantifying the degree of augmentation in central pressure due to wave reflection from the peripheral circulation (22). AIx measurements were calculated from 5-min averages at rest and during blood flow restriction with an arm cuff, and measurements were normalized to a heart rate of 75 beats/min (22). A detailed description of the procedure for obtaining AIx is described in previous reports (31, 32).

Statistical Analyses

All analyses were performed using R (www.r-project.org, with the regression models using the “stats” package) and a two-sided P < 0.05 was considered statistically significant. All values were reported as means  ±  SD or N (%).

To address our hypotheses, we examined the association of D-dimer at baseline with hemodynamic fluctuations (see Blood pressure monitoring) during OSA events with linear regression. As pulse pressure was highly collinear with aortic size in our cohort (Fig. 3), we also adjusted for aortic root diameter in our regression analyses. We then compared the mean AHI, number of arousals, Pes_peak-insp, SBP_max, PP_max, HR_max, ΔSBP, ΔPP, ΔHR, O₂ saturation, and other summarized sleep study characteristics between the baseline and CPAP nights using paired t tests. We also compared the morning D-dimer levels and AIx between baseline and CPAP studies with paired t tests.

Figure 3.

Correlation of brachial pulse pressure and aortic root diameter. N = 11, participants with native aortas, since 5 participants had aortic grafts. N = 11 total sample size of n = 7 men and 4 women. r, correlation coefficient.

We also examined the correlation between D-dimer and AIx at baseline with scatterplots, and then performed the Fisher exact method to test if the reductions in D-dimer after CPAP treatment were associated with decreases in AIx.

RESULTS

Participant Characteristics

Participants’ demographic and anthropometric characteristics, as well as baseline hemodynamics, are presented in Table 1. The study population included 10 males and 6 females with an average age of 36 ± 13 yr, body mass index of 26 ± 5 kg/m², and 81% were Caucasian. The average systolic and diastolic blood pressures were 118 ± 10 mmHg and 71 ± 6 mmHg, respectively. The average aortic root diameter was 4 ± 1 cm among those with native aortas, who made up 11 of the total sample of 16 participants. Four subjects were on an angiotensin receptor blocker, three were on a β-blocker, six were on a combination of an angiotensin receptor blocker and a β-blocker, and three were not on any blood pressure medication.

Table 1.

Participant characteristics

Characteristics		N
Sex, men:women	10:6	16
Age, yr	36 ± 13	16
Race, n (%)
Caucasian	13 (81)	13
Asian	1 (6)	1
Other*	2 (13)	2
Body mass index, kg/m2	26 ± 5	16
Systolic blood pressure, mmHg	118 ± 10	16
Diastolic blood pressure, mmHg	71 ± 6	16
Pulse pressure, mmHg	43 ± 10	16
Heart rate, beats/min	76 ± 13	16
Hypertension, n (%)	1 (6)	16
Aortic root diameter, cm	4 ± 1	11
Aortic root replacement, n (%)	5 (31)	5
Blood pressure medications, n (%)
β-Blocker only	3 (19)	3
ARB only	4 (25)	4
β-Blocker and ARB	6 (38)	6
None	3 (19)	3

Values are means ± SD or absolute values with frequency, n (%); N = 16 participants, total sample size. *Other: participants who did not identify as African American, Hispanic, Caucasian, or Asian. ARB, angiotensin receptor blocker.

Association of OSA-Induced Hemodynamic Fluctuations and D-Dimer

Linear regression analyses revealed a positive association between D-dimer and PP_max during OSA events in both the unadjusted model (β = 26.3, r = 0.523, P = 0.038) (Fig. 4) and a model adjusted for aortic root diameter obtained from participants with native aortas (β = 62.9, r = 0.733, P = 0.028). The association between D-dimer and other measures of hemodynamic fluctuations are shown in Table 2.

Figure 4.

Correlation of the average maximum pulse pressure (PP) during obstructive sleep apnea events and D-dimer at baseline (A) and correlation of augmentation index (AIx) and D-dimer at baseline (B). N = 16 participants, total sample size of n = 10 men and 6 women. r, correlation coefficient.

Table 2.

Relationship between D-dimer and hemodynamic fluctuations

	D-Dimer
	β	r	P	N
SBPmax
Univariable	16.9	0.464	0.070	16
Adjusted	20.1	0.566	0.167	16
PPmax
Univariable	26.3	0.523	0.037	16
Adjusted	62.9	0.735	0.028	16
HRmax
Univariable	−4.0	0.063	0.827	16
Adjusted	7.7	0.366	0.769	16
ΔSBP
Univariable	72.9	0.482	0.059	16
Adjusted	105.4	0.669	0.063	16
ΔPP
Univariable	141.7	0.472	0.065	16
Adjusted	212.7	0.672	0.060	16
ΔHR
Univariable	−28.1	0.118	0.658	16
Adjusted	−15.9	0.358	0.844	16

N = 16 participants, total sample size of n = 10 men and 6 women. SBP_max, maximum systolic blood pressure; PP_max, maximum pulse pressure; HR_max, maximum heart rate; Δ, change; β, β-coefficient; r, correlation coefficient. Statistical method, linear regression. Boldface indicates significant P value.

Effect of CPAP on OSA, Hemodynamics, and D-Dimer

CPAP decreased the AHI, number of arousals, respiratory rate, maximum inspiratory effort (Pes_peak-insp), HR_max, ΔSBP, ΔPP, and ΔHR, but did not change sleep architecture, SBP_max, PP_max, and O₂ saturation (see Table 3). CPAP also decreased D-dimer levels from a mean (SD) of 1,108 ± 656 ng/mL to 882 ± 532 ng/mL (P = 0.023) and AIx from a mean (SD) of 7 ± 4% to 0 ± 5% (P = 0.117) (Fig. 5).

Table 3.

Effect of CPAP on nocturnal physiology

	Baseline	CPAP	P	N
Sleep architecture
Total sleep time, min	371 ± 77	371 ± 49	0.970	16
Sleep efficiency, %	82 ± 15	81 ± 10	0.635	16
REM, %	15 ± 8	14 ± 8	0.618	16
Sleep latency, min	19 ± 20	19 ± 16	0.958	16
Wake after sleep onset, min	61 ± 63	70 ± 55	0.454	16
Respiratory parameters
AHI, events/h	20 ± 17	3 ± 2	0.001	16
NREM AHI, events/h	19 ± 17	3 ± 2	0.001	16
REM AHI, events/h	18 ± 15	6 ± 5	0.009	16
O2, %	96 ± 1	96 ± 2	0.937	16
Respiratory rate, breaths/min	17 ± 3	16 ± 2	0.015	16
Pespeak-insp cmH2O	−10 ± 4	0 ± 2	<0.001	11#
Hemodynamic parameters*
SBPmax, mmHg	126 ± 18	125 ± 16	0.857	16
PPmax, mmHg	48 ± 13	48 ± 16	0.843	16
HRmax, beats/min	72 ± 10	67 ± 10	0.001	16
ΔSBP, mmHg	13 ± 4	9 ± 3	0.011	16
ΔPP, mmHg	7 ± 2	5 ± 2	0.013	16
ΔHR, beats/min	9 ± 3	5 ± 1	<0.001	16
Arousal and positional analyses
Total arousals	124 ± 51	80 ± 42	0.004	16
Respiratory arousals	64 ± 38	11 ± 8	<0.001	16
Supine sleep, %	47 ± 22	50 ± 28	0.646	16

Values are means ± SD; N = 16 participants, total sample size of n = 10 men and 6 women. #The other 5 participants either refused esophageal catheterization or did not have usable esophageal pressure (Pes) data. *Hemodynamic fluctuations during obstructive sleep apnea events and normal breathing with continuous positive airway pressure (CPAP). AHI, apnea hypopnea index; REM, random eye movement; NREM, non-random eye movement; Pes_peak-insp, peak inspiratory esophageal pressure (larger negative values indicate greater respiratory effort); SBP_max, maximum systolic blood pressure; PP_max, maximum pulse pressure; HR_max, maximum heart rate; Δ, change. Statistical method, paired t test. Boldface indicates significant P value.

Figure 5.

D-dimer levels (A) and augmentation index (AIx; B) after overnight baseline and continuous positive airway pressure (CPAP) studies are shown. N = 16 participants, total sample size of n = 10 men and 6 women. Statistical method, paired t test.

Association of D-Dimer and AIx

We found a significant linear relationship between D-dimer and AIx at baseline (r = 0.713, P = 0.002; see Fig. 4). In addition, the Fisher exact test revealed a significant association between the proportion of subjects with reduced D-dimer and reduced AIx after CPAP treatment (P = 0.007), suggesting that the decrease in D-dimer was dependent on a decrease in AIx.

DISCUSSION

In this randomized CPAP crossover study, we generated several novel findings that characterize the effect of OSA on D-dimer levels in our cohort of patients with MFS without overt aortic dissection. First, OSA-induced hemodynamic fluctuation manifested as pulse pressure was associated with D-dimer at baseline. Second, we found that overnight OSA treatment decreased morning D-dimer levels. Third, reduction in D-dimer after CPAP treatment was associated with a concomitant reduction in AIx. In conclusion, our data support the hypothesis that OSA is a modifiable cause of vascular stress and injury in MFS and that D-dimer may serve as a biomarker of cardiovascular stress in MFS.

Our study extends previous work on humoral changes in patients with MFS. Kornhuber et al. (12) found that patients with MFS with enlarged aortic root had elevated levels of D-dimer compared with matched non-MFS controls. Ihara et al. (13) studied non-MFS patients with aortic aneurysms and found that D-dimer and markers of collagen turnover were greater in the patients with aortic aneurysms than in controls. In our study, D-dimer was also elevated in asymptomatic, clinically stable patients with MFS. Specifically, only one-third of our participants had D-dimer levels lower than 500 ng/mL, the threshold below which acute aortic dissection is ruled out (10, 33). We also show that treatment of OSA reduces D-dimer levels, indicating a contribution of OSA to the elevated baseline values. Since D-dimer levels were also positively associated with maximum pulse pressures during OSA events (Fig. 4), and even after accounting for aortic size (Table 2), the hemodynamic fluctuations may trigger low-grade aortic injury with increased D-dimer as a precursor for overt dissection. Our findings together with those of others (12, 13) suggest that both a structurally compromised aortic wall and factors such as OSA that increase cardiovascular stress may contribute to vascular injury and increased D-dimer levels in MFS.

In patients with MFS, who already have structural aortic compromise, OSA presents an additional risk factor that may accelerate aortic morbidity. It is well recognized that OSA leads to intermittent sympathetic nervous system (SNS) activation leading to fluctuations in blood pressure and heart rate (34, 35), which increase aortic stress (14). As noted above, we found a strong association between D-dimer and OSA-induced fluctuations in pulse pressure. In addition, OSA treatment decreased blood pressure and heart rate fluxes (Table 3), with a subsequent reduction in morning D-dimer levels. In our study, therefore, higher D-dimer levels at baseline may have resulted from exaggerated hemodynamic surges increasing vascular wall stress leading to intramural microthrombosis and fibrinolysis. Our measure of vascular stress was AIx, which we found to be highly correlated with D-dimer at baseline (Fig. 4). Likewise, CPAP reduction of D-dimer was associated with CPAP reduction of AIx, suggesting AIx as a mediator for OSA-induced vascular injury. In fact, two previous studies showed that AIx was a predictor of aortic adverse events in patients with MFS (36, 37).

Other mechanisms that may contribute to OSA-induced vascular injury or explain our findings include increased inspiratory effort and hypoxia (24, 38, 39). In this study, elimination of OSA with CPAP significantly reduced the maximum inspiratory effort (Pes_peak-insp) (Table 3), which would decrease the aortic transmural pressure (24, 40), thereby potentially mitigating aortic injury (14, 24). On the other hand, the mean oxygen saturation did not change with CPAP, probably because our participants were not hypoxic at baseline (Table 3), which also implies that hypoxia did not contribute to our findings.

The main strength of this study is the randomized crossover design. Our study is also the first to conduct comprehensive monitoring of nocturnal hemodynamic physiology in patients with MFS. From a practical standpoint, overnight studies that quantify cardiovascular stress and incorporate biochemical readouts may offer a useful platform to test interventions for vascular injury in MFS. Whether this structure works better for biomechanical interventions such as CPAP rather than pharmacological interventions is unknown but should be explored. Our limitation was the small sample size, which prevented multivariable analyses to determine whether blood pressure medication and MFS subgenotype modifies the effect of OSA on D-dimer levels. The sample size along with the short-term protocol may also explain the lack of a statistical change in mean AIx after CPAP treatment but highlights the sensitivity of D-dimer to acute nocturnal cardiovascular stress in MFS.

In conclusion, our study invokes OSA as a modifiable contributor to vascular injury and increased D-dimer in MFS. OSA screening along with D-dimer assay in patients with MFS may help with monitoring cardiovascular stress and identification of those with the greatest risk for aortic adverse events. Future studies are still needed to validate our findings and determine if OSA-induced D-dimer increase is a precursor for aortic disease progression in patients with MFS.

GRANTS

The study was funded by the Victor McKusick Marfan Foundation Fellowship Award, the NIH T32 Multi-Institutional Training Program in Sleep and Genetics (HL 110952‐6), and the Johns Hopkins Pulmonary and Critical Care Medicine Inspire Award.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the authors.

AUTHOR CONTRIBUTIONS

M.S., H.S., and E.N. conceived and designed research; M.S. performed experiments; M.S. and F.S. analyzed data; M.S., H.S., J.J., A.S., L.P., P.S., V.P., and E.N. interpreted results of experiments; M.S. and F.S. prepared figures; M.S. and E.N. drafted manuscript; M.S., H.S., J.J., G.M., A.S., L.P., F.S., J.L., P.S., V.P., and E.N. edited and revised manuscript; M.S., H.S., J.J., G.M., A.S., L.P., F.S., J.L., P.S., V.P., and E.N. approved final version of manuscript.