Original Investigation Exercise Capacity in Polycystic Kidney Disease Nata´lia Lopes Reinecke, MSc,1 Thulio Marquez Cunha, MD, PhD,2 Ita Pfeferman Heilberg, MD, PhD,1 Elisa Mieko Suemitsu Higa, MD, PhD,1 Jose´ Luiz Nishiura, MD, PhD,1 Jose´ Alberto Neder, MD, PhD,2 Waldemar Silva Almeida, MD, PhD,1 and Nestor Schor, MD, PhD1 Background: Reports about exercise performance in autosomal dominant polycystic kidney disease (ADPKD) are scarce. We aimed to evaluate exercise capacity and levels of nitric oxide and asymmetric dimethylarginine (ADMA) in normotensive patients with ADPKD. Study Design: Prospective controlled cohort study. Setting & Participants: 26 patients with ADPKD and 30 non-ADPKD control participants (estimated glomerular filtration rate . 60 mL/min/1.73 m2, aged 19-39 years, and blood pressure [BP] , 140/ 85 mm Hg). We excluded smokers, obese people, and individuals with associated diseases. Predictor: ADPKD versus control. Outcomes: Exercise capacity and nitric oxide and ADMA levels in response to exercise. Measurements: Cardiopulmonary exercise testing and serum and urinary nitric oxide, plasma ADMA, and BP levels before and after exercise. Results: Mean basal systolic and diastolic BP, estimated glomerular filtration rate, and age did not differ between the ADPKD and control groups (116 6 12 vs 110 6 11 mm Hg, 76 6 11 vs 71 6 9 mm Hg, 113 6 17 vs 112 6 9.6 mL/min/1.73 m2, and 30 6 8 vs 28.9 6 7.3 years, respectively). Peak oxygen uptake and anaerobic threshold were significantly lower in the ADPKD group than in controls (22.2 6 3.3 vs 31 6 4.8 mL/ kg/min [P , 0.001] and 743.6 6 221 vs 957.4 6 301 L/min [P 5 0.01], respectively). Postexercise serum and urinary nitric oxide levels in patients with ADPKD were not significantly different from baseline (45 6 5.1 vs 48.3 6 4.6 mmol/L and 34.7 6 6.5 vs 39.8 6 6.8 mmol/mg of creatinine, respectively), contrasting with increased postexercise values in controls (63.1 6 1.9 vs 53.9 6 3.1 mmol/L [P 5 0.01] and 61.4 6 10.6 vs 38.7 6 5.6 mmol/mg of creatinine [P 5 0.01], respectively). Similarly, whereas postexercise ADMA level did not change in the ADPKD group compared to those at rest (0.47 6 0.04 vs 0.45 6 0.02 mmol/L [P 5 0.6]), it decreased in controls (0.39 6 0.02 vs 0.47 6 0.02 mmol/L [P 5 0.006]), as expected. A negative correlation between nitric oxide and ADMA levels after exercise was found in only the control group (r 5 20.60; P , 0.01). Limitations: Absence of measurements of flow-mediated dilatation and oxidative status. Conclusions: We found lower aerobic capacity in young normotensive patients with ADPKD with preserved kidney function and inadequate responses of nitric oxide and ADMA levels to acute exercise, suggesting the presence of early endothelial dysfunction in this disease. Am J Kidney Dis. -(-):---. ª 2014 by the National Kidney Foundation, Inc. INDEX WORDS: Autosomal dominant polycystic kidney disease (ADPKD); exercise; nitric oxide; asymmetric dimethylarginine (ADMA); cystic diseases; polycystic kidney; physical capacity; cardiopulmonary exercise testing.
A
utosomal dominant polycystic kidney disease (ADPKD) is the most common renal genetic disorder; it affects 4-6 million individuals worldwide and accounts for w3% of end-stage renal disease.1,2 It is caused by mutations in the genes PKD1 or PKD2, which code for the transmembrane proteins polycystin 1 and polycystin 2, respectively. Functional deficiency in polycystin predisposes to the development of renal cysts and several renal and extrarenal manifestations, such as intracranial and aortic aneurysms, mitral valve prolapse, endothelial dysfunction, and hypertension.1,3,4 Hypertension, which is an early and common finding in patients with ADPKD, occurs in w60% of the patients before decreases in kidney function and is associated with a swift progression to end-stage renal disease.5,6 In addition to renal cystic expansion and intrarenal ischemia, endothelial dysfunction, found in Am J Kidney Dis. 2014;-(-):---
the early stages of the disease, also may play a role in the development of hypertension in ADPKD.7 The disruption of the normal function of polycystins, also expressed in vascular endothelial cells, results in an impaired shear stress sensitivity mechanism that may affect the synthesis of nitric oxide (NO), which results From the 1Renal Division and 2Pneumology Division, Department of Medicine, Federal University of São Paulo, São Paulo, Brazil. Received November 5, 2013. Accepted in revised form March 18, 2014. Address correspondence to Nestor Schor, MD, PhD, Department of Medicine, Renal Division, Federal University of São Paulo, Rua Botucatu, 740 CEP:04023-900 São Paulo–SP, Brazil. E-mail: [email protected] 2014 by the National Kidney Foundation, Inc. 0272-6386/$36.00 http://dx.doi.org/10.1053/j.ajkd.2014.03.014 1
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in lower systemic NO levels.8-10 Furthermore, levels of plasma asymmetric dimethylarginine (ADMA), an endogenous inhibitor of NO, also have been observed to be increased in patients with ADPKD compared with healthy controls.11 During physical exercise, several cardiovascular adjustments contribute to providing an increased oxygen supply to the muscles of healthy individuals. The higher production of endothelial NO stimulated by increased shear stress on vessel walls represents one such mechanism, which results in decreased total vascular peripheral resistance.12-14 Martinez-Vea et al15 showed an exaggerated diastolic blood pressure (BP) response during acute exercise in young normotensive patients with ADPKD, who presented with greater left ventricular mass index, which is consistent with early diastolic dysfunction. These findings prompted us to investigate the response patterns to acute exercise in an ADPKD sample consisting of young and normotensive patients with preserved kidney function and without left ventricular hypertrophy. Presently, there are scarce data available regarding the physical exercise response in patients with ADPKD. Because endothelial dysfunction might precede clinically detectable hypertension in ADPKD, we aimed to evaluate the physical capacity and the influence of acute aerobic exercise on serum and urinary NO and ADMA levels in the early stages of ADPKD in a group of normotensive patients with preserved kidney function.
METHODS Setting and Participants Forty-three adult patients with ADPKD being followed up at the Polycystic Kidney Disease Unit of the Nephrology Division at the Federal University of São Paulo were eligible for the study based on data from their medical records (all records [n 5 263] were analyzed). Inclusion criteria for patients with ADPKD were positive family history of ADPKD and renal cysts detected by ultrasound using the diagnostic criteria established by Pei et al,16 systolic BP , 140 mm Hg and diastolic BP , 85 mm Hg without antihypertensive medication,17 and estimated glomerular filtration rate (eGFR) . 60 mL/min/1.73 m2 in individuals aged 19-39 years. Only those who were sedentary and not involved in a physical training program were selected for the study. Exclusion criteria for patients with ADPKD were history of aneurysms; smoking; the presence of associated diseases such as diabetes, hyperlipidemia, and obesity; and use of medications that could interfere with cardiac or pulmonary function. Patients were informed about the study and invited to participate by personal contact. A total of 26 patients (18 women and 8 men) were enrolled in the study, and reasons for discontinuation are shown in Fig 1. Hemoglobin concentration and hematocrit, as well as echocardiography and ultrasonography results, were collected from patients’ medical records. Renal volume had been measured by ultrasound and calculated using a modified ellipsoid formula.18 The values for both kidneys were combined and divided by the individual’s height to yield total kidney volume.19 eGFR was 2
Medical records analyzed (n=263) Excluded by exclusion criteria (n=220)
Assessed for eligibility (n=43)
Scheduled for CPET
(n=13)
(n=30)
Unable to perform CPET (failed to pedal)
(n=2)
Declined to participate
Scheduled for exercise protocol
(n=28)
Discontinued participation (n=2)
Control group (n=30)
Completed protocol (n=26)
Figure 1. Patient enrollment flow chart. Abbreviation: CPET, cardiopulmonary exercise testing.
calculated using the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) creatinine equation,20 and the stages of chronic kidney disease were defined according to the KDIGO (Kidney Disease: Improving Global Outcomes) guidelines.21 The control group was recruited from laboratory staff and consisted of 30 adult age-matched apparently healthy nonobese sedentary individuals (9 men and 21 women). Exclusion criteria consisted of history of cardiovascular, pulmonary or kidney diseases; diabetes; smoking; hyperlipidemia; or hypertension. A fasting blood sample was collected for measurement of hemoglobin, hematocrit, and creatinine, determined by the alkaline picrate method. The reason that healthy individuals and not patients with chronic kidney disease were chosen as a control group was the exclusion of confounding factors for endothelial dysfunction. We obtained written consent from all participants. The local ethics committee approved the study.
Physical Activity Assessment The Baecke Habitual Physical Activity Questionnaire, which measures qualitative and quantitative indexes and addresses occupational, leisure, and locomotion activities,22 was used to assess physical activity performed during the past 12 months. This questionnaire has been validated in a Brazilian population survey.23
Pulmonary Function Test Spirometric tests24 were performed using the Clinical Pulmonary Function-Spirometry System (Medical Graphics Corp-MGC), and airflow was measured using a calibrated pneumotachograph. Participants completed at least 3 acceptable maximum forced expiratory maneuvers. The following variables were measured: forced vital capacity (FVC), forced expiratory volume in the first second of expiration (FEV1), and FVC:FEV1 ratio. Am J Kidney Dis. 2014;-(-):---
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Cardiopulmonary Exercise Testing
NO Determination
Cardiopulmonary exercise testing was conducted by a pneumologist, according to the protocol described by Neder et al.25 Oxygen uptake (VO2) measured at peak exercise (VO2peak) during incremental cardiopulmonary exercise testing is considered to be the gold standard for evaluation of exercise capacity.26 Exercise tests were conducted using an electromagnetically braked cycle ergometer (CPE 2000; Medical Graphics CorpMGC), with gas exchange and ventilator variables analyzed breath by breath using a calibrated computer-based exercise system (MGC-CPX System; Medical Graphics Corp). Cardiac electrical activity (CardioPerfect; MGC) and oxygen saturation measured by pulse oximetry (Ohmeda Biox 3740) were recorded continuously. The exercise test consisted of: (1) 2 minutes at rest, (2) 2 minutes of real zero-external intensity exercise obtained with the use of an electrical system that moved the ergometer flywheel at 60 rpm, (3) an incremental phase, and (4) a 4-minute recovery period. Power (watts) was increased continuously in a linear ramp pattern (10-20 W$min21) such that the test duration was longer than 8 but shorter than 12 minutes. The individual pedaled to the limit of tolerance with active and standardized encouragement from the same investigator. Average pulmonary VO2 (in liters per minute) for the last 30 seconds of the ramp was considered to be representative of the participant’s VO2peak. VO2 at the anaerobic threshold was estimated by the gas exchange threshold method (V-slope).27 All tests were performed in the morning in the same laboratory with temperature controlled at 21 C-24 C.
Serum and urine nitrate concentrations were measured using an NO analyzer (NOA TM280; Sievers Inc).31 Serum and urinary samples were previously deproteinized in zinc sulfate, 10% (1:2), and sodium hydroxide (1:2) for 15 minutes and centrifuged for 5 minutes. A 10-mL aliquot of the supernatant was injected into the analyzer. A calibration curve was plotted for standards of sodium nitrate. In the NO analyzer, nitrate was reduced to NO with vanadium at 90 C, and the NO formed was detected by gaseousphase chemiluminescence after reaction with ozone. The analyzer remained stable, and linearity was maintained during the test period to #1% with a coefficient of variation of #10%. Serum results are expressed as micromoles per liter. Urinary NO excretion was corrected by urinary creatinine and expressed as micromoles per milligram of creatinine.
Exercise Protocol After a minimum 1-week period, participants underwent a 20-minute single bout of cycle ergometer exercise. Exercise intensity was measured by the anaerobic threshold (AT) according to the following formula: D 5 VO2peak (watts) – AT (watts), and Intensity 5 (D 3 0.3) 1 AT (watts). This intensity corresponds to 60% of VO2peak, which was considered to be moderate.28 During the exercise, patients were advised to ingest 200-300 mL of water according to recommendations of the National Athletic Trainers’ Association.29
BP Measurements All participants arrived at the laboratory after an 8-hour fasting period with no water intake restriction. A light snack (25 g of salt cracker and 200 mL of peach juice) was given to participants, followed by a 30-minute rest in seated position. Before starting the exercise, we measured BP by auscultation (sphygmomanometer and stethoscope; Diasyst LTDA) 3 times according to the American Heart Association procedure30 and took the mean value. We also measured BP during exercise (every 3 minutes) and after exercise (every minute until 5 minutes and subsequently, every 3 minutes until 30 minutes after exercise). All BP measurements were made by a single person in order to minimize possible errors. We measured heart rate continuously before, during, and after exercise using an oximeter.
Biochemical Assays All participants fasted for 8 hours overnight and were instructed to avoid foods that could affect the biochemical assays for at least 48 hours before blood and urine collections. Venous blood samples were obtained from the forearm at rest and at the end of the exercise protocol. Urine samples were collected at rest and after the exercise protocol. Samples were centrifuged at 4 C for 20 minutes at 2,000 rpm. The supernatant was removed with a polypropylene pipette, aliquoted into 0.5-mL samples in polypropylene tubes, and stored at 280 C for NO and ADMA measurements. Am J Kidney Dis. 2014;-(-):---
ADMA Determination We assessed plasma ADMA concentration using an ADMA enzyme-linked immunosorbent assay (ELISA) kit validated by Schulze et al32 (Immundiagnostik AG), according to the manufacturer’s instructions. Standard samples and controls were added and incubated overnight at 2 C-8 C. After several washings with a wash buffer, the conjugate was added and incubated for 1 hour at room temperature. Thereafter, the washing step was repeated and the substrate was added and incubated for 25 minutes at room temperature on an orbital shaker. Stop solution was added and immediately analyzed using an ELISA reader at 450nm.
Statistical Analyses We performed statistical analyses using SPSS 19.0 for Windows (SPSS Inc). Data are expressed as mean 6 standard deviation. The data were verified for normality by descriptive statistics, such as comparison of the mean versus the median, histogram and boxplot inspection, and skewness and kurtosis coefficients. The t test was used to compare clinical, spirometric, and cardiopulmonary exercise testing variables of the studied groups. Repeated-measures analysis of variance with Bonferroni post-test was used to analyze NO, ADMA, BP, and heart rate responses to exercise in both groups. Pearson linear correlation was used to establish relationships between variables. Statistical significance was defined as P , 0.05. Comparison of categorical data was performed by c2 test.
RESULTS Baseline Clinical and Laboratory Characteristics Estimated time from the diagnosis of ADPKD until our study ranged from 1-192 months. Total kidney volume adjusted for the participant’s height averaged 175-1,760 (median, 280) mL/m. Based on eGFR, 23 patients were classified as chronic kidney disease stage 1, and 3 patients, as stage 2; mean eGFR was 113 6 17 [standard deviation] mL/min/1.73 m2. Liver size was described as normal for all patients according to computed tomographic results available in their medical records (data not shown). None of these patients presented with microalbuminuria (mean albumin excretion, 9.8 6 5 mg/min). Echocardiographic parameters retrieved from patients’ medical records, such as mean ejection fraction (66% 6 3%) and interventricular septum (7.9 6 0.8 mm), were within the expected range for healthy individuals. No patient presented with left ventricular mass hypertrophy; mean left ventricular mass index was 74.8 6 17 g/m2. 3
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Table 1 lists clinical characteristics of patients and controls. No statistical difference was found with respect to mean age, sex, body mass index, BP at rest, eGFR, hematocrit, hemoglobin level, level of physical activity, and parameters of pulmonary function (FVC, FEV1, and FEV1:FVC ratio). Cardiopulmonary Exercise Testing Cardiopulmonary exercise testing results are listed in Table 2. VO2peak, VO2 at anaerobic threshold, maximal workload, anaerobic threshold workload, respiratory exchange ratio, and oxygen pulse were significantly lower in patients with ADPKD than in controls. We observed no statistical differences between groups with respect to peak carbon dioxide production, respiratory rate, maximum heart rate, end-tidal oxygen partial pressure, or end-tidal carbon dioxide partial pressure. No significant correlations were found between height-adjusted total kidney volume and the exercise parameters VO2peak (r 5 0.08; P 5 0.7), VO2 at Table 1. Baseline Clinical Characteristics, Physical Activity, and Pulmonary Function Parameters
Variable
Sex Female Male
Control (n 5 30)
ADPKD (n 5 26)
21 9
18 8
P
0.9
Age (y) 28.9 6 7.3 30.0 6 8.0 Weight (kg) 68.1 6 14.2 69.1 6 16.3 Height (cm) 169.0 6 9.0 170.0 6 10.0 BMI (kg/m2) 23.5 6 2.8 23.7 6 4.7 SBP (mm Hg) 110.0 6 11.0 116.0 6 12.0 DBP (mm Hg) 71.0 6 9.0 76.0 6 11.3 eGFR (mL/min/1.73 m2) 112 6 9.6 113 6 17 Hematocrit (%) 41.1 6 3.2 41 6 4 Hemoglobin (g/dL) 13.4 6 0.6 13.9 6 1.4 Microalbuminuria — 9.8 6 5 (mg/min) Height-adjusted TKV — 280 (175; 1,760) (mL/m) HPAQ score 7.3 6 1.0 7.4 6 1.9 FVC (L) 4.1 6 0.8 4.1 6 0.9 FEV1 (L) 3.4 6 0.6 3.4 6 0.7 FEV1:FVC ratio (%) 85.3 6 6.2 83.2 6 6.4 Baseline serum NO 53.9 6 3.1 48.3 6 4.6 (mmol/L) Baseline plasma ADMA 0.47 6 0.02 0.45 6 0.02 (mmol/L)
0.6 0.9 0.6 0.9 0.09 0.2 0.7 0.9 0.4
— —
0.8 0.9 0.9 0.3 0.1 0.8
Note: Values for categorical variables are given as number; values for continuous variables are given as mean 6 standard deviation or median (minimum; maximum). Abbreviations: ADMA, asymmetric dimethylarginine; ADPKD, autosomal dominant polycystic kidney disease; BMI, body mass index; DBP, diastolic blood pressure; eGFR, estimated glomerular filtration rate; FEV1, forced expiratory volume in the first second of expiration; FVC, forced vital capacity; HPAQ, Habitual Physical Activity Questionnaire; NO, nitric oxide; SBP, systolic blood pressure; TKV, total kidney volume. 4
anaerobic threshold (r 5 0.01; P 5 0.9), and maximal workload (r 5 20.07; P 5 0.8) in the ADPKD group. Exercise Protocol BP Responses Exercise systolic and diastolic BP responses are shown in Fig 2A. Systolic and diastolic BP during exercise and recovery were significantly greater in patients with ADPKD versus controls. Heart Rate Heart rates during exercise and recovery, shown in Fig 2B, did not differ between groups. Nitric Oxide At baseline, serum and urinary NO levels did not differ between groups. However, postexercise NO levels increased in controls compared to baseline (serum: 63.1 6 1.9 vs 53.9 6 3.1 mmol/L [P 5 0.01]; urinary excretion: 61.4 6 10.6 vs 38.7 6 5.6 mmol/mg creatinine [P 5 0.01]), but did not increase in patients with ADPKD (serum: 45 6 5.1 vs 48.3 6 4.6 mmol/L [P 5 0.9]; urinary excretion: 39.8 6 6.8 vs 34.7 6 6.5 mmol/mg creatinine [P 5 0.8]; Fig 3A and B). In the control group only, there was a negative correlation between serum NO level after exercise and ADMA level after exercise (r 5 20.60; P 5 0.01). Asymmetric Dimethylarginine Basal ADMA values did not differ between groups. However, ADMA levels decreased in controls after exercise versus baseline (0.39 6 0.02 vs 0.47 6 0.02 mmol/L; P 5 0.006), but did not decrease in patients with ADPKD (0.47 6 0.04 vs 0.45 6 0.02 mmol/L; Table 2. Cardiopulmonary Exercise Testing Parameters
Variable
Control (n 5 30)
ADPKD (n 5 26)
P
VO2peak (mL/kg/min) 31.0 6 4.8 22.2 6 3.3 ,0.001 VCO2peak (mL/min) 2,254.2 6 727.2 1,990.1 6 690.4 0.2 VO2AT (L/min) 957.4 6 300.8 743.6 6 220.8 0.01 MW (W) 156.5 6 50.8 114.4 6 30.2 0.002 ATW (W) 70.9 6 21.2 47.9 6 10.2 ,0.001 RERmax 1.17 6 0.1 1.25 6 0.1 0.007 RR (rpm) 38.7 6 10.8 35.7 6 9.1 0.3 HRmax (beats/min) 174.0 6 18.0 174.9 6 14.0 0.9 11.4 6 3.9 9.0 6 2.7 0.02 VO2/HRmax PETO2 (mm Hg) 102.9 6 5.2 102.9 6 8.0 0.9 PETCO2 (mm Hg) 34.5 6 4.5 34.4 6 6.0 0.9 Note: Values are given as mean 6 standard deviation. Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; ATW, anaerobic threshold workload; HRmax, maximum heart rate; MW, maximal workload; PETCO2, end-tidal carbon dioxide partial pressure; PETO2, end-tidal oxygen partial pressure; RERmax, maximum respiratory exchange ratio; RR, respiratory rate; VCO2peak, peak carbon dioxide production; VO2AT, oxygen uptake at anaerobic threshold; VO2peak, peak oxygen uptake; Vo2/HRmax, maximum oxygen uptake to heart rate ratio. Am J Kidney Dis. 2014;-(-):---
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Figure 2. (A) Blood pressure (BP) and (B) heart rate (HR) at rest and during and after exercise. Values are expressed as mean 6 standard error (SE). *P , 0.05 versus control. Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; DBP, diastolic BP; SBP, systolic BP.
P 5 0.6; Fig 3C). No correlations were found between ADMA level and kidney size or NO level and kidney size.
DISCUSSION The present study showed that young normotensive patients with ADPKD with preserved kidney and cardiac function have impaired physical capacity, evidenced by the lower VO2peak and anaerobic threshold compared with healthy individuals with the same previous level of physical activity assessed by the Baecke Habitual Physical Activity Questionnaire. Considered gold-standard measures of cardiorespiratory fitness, VO2peak and anaerobic threshold also are predictors of all-cause and cardiovascular disease mortality.33,34 The normal spirometry parameters (FVC and FEV1) and liver size and the absence of significant correlations between height-adjusted TKV Am J Kidney Dis. 2014;-(-):---
and exercise parameters (VO2peak, VO2 at anaerobic threshold, and maximal workload) suggest that liver and/or kidney size did not interfere with patients’ physical capacity results. Additionally, the ejection fraction and serum hemoglobin levels within normal range suggested adequate oxygen transport in patients with ADPKD. According to Hambrecht et al,35 systemic vasoconstriction and diminished peripheral perfusion can lead to exercise intolerance. In this context, NO plays a key role in vasomotor tone and perfusion regulation. It has been suggested that higher submaximal and maximal aerobic exercise performance is associated with greater flow-mediated dilatation of peripheral conduit arteries in young healthy men.36 Thus, higher peripheral vascular resistance could impair exercise capacity. In the present series, baseline serum and urinary NO levels in patients with ADPKD were similar to 5
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Figure 3. (A) Serum nitric oxide (NO) levels, (B) urinary NO excretion, and (C) plasma asymmetric dimethylarginine (ADMA) levels. Values are expressed as mean 6 standard error (SE). Abbreviation: ADPKD, autosomal dominant polycystic kidney disease.
levels in controls. Although these findings contrast with those from the study of Wang et al,11 who demonstrated impaired production of NO in normotensive patients with ADPKD, their patients were older and presented with microalbuminuria, a feature not shared by the present sample and that could indicate that their patients already had a higher rate of endothelial dysfunction. However, under the exercise stimulus, serum NO levels did not increase in the ADPKD group as expected and observed in the control group, thus demonstrating disturbed NO regulation. Urinary NO excretion also increased after 6
exercise in only the control group, probably due to the higher production during exercise. Higher NO production during physical exercise is due to exercise-induced increases in endothelial shear stress,37,38 which is necessary to increase blood flow and the influx of ionized calcium in endothelial cells, releasing NO.39,40 In the present study, the lack of increase in NO levels, accompanied by a nondecrease in serum ADMA levels after exercise, strongly suggests impaired capacity for exercise-induced vasodilatation. Our findings agree with data published by Wang et al,11 who found higher serum ADMA levels in patients with ADPKD. These investigators suggested that the increased oxidative stress in patients with ADPKD might contribute to the increased ADMA levels by decreasing its metabolism by dimethylarginine dimethylaminohydrolase (DDAH). However, as described for NO, ADMA levels did not differ from the control group at baseline, but only after exercise. Surdacki et al41 suggested that high ADMA levels would lead to diminished NO bioavailability, thereby increasing vascular resistance and BP. Martinez-Vea et al15 observed an exaggerated diastolic BP response during exercise in young normotensive patients with ADPKD, who showed greater left ventricular mass index, consistent with early diastolic dysfunction. In the present study, the inadequate systolic and diastolic BP responses during exercise evident in patients with ADPKD with preserved cardiac function further reinforce the hypothesis of higher vascular resistance leading to defective vasodilatation. This altered NO response likely may be influenced by the mutated polycystins at endothelial and smooth vascular cells because they are required for an appropriate mechanism of blood shear stress by a complex biochemical cascade involving calcium, calmodulin, Akt (also known as protein kinase B [PKB]), and protein kinase C.8,9 Nevertheless, because NO has various nonvascular sources and its systemic levels are only indirect evidence for its role in skeletal muscle exercise hyperemia,42 more studies are necessary to identify whether these exercise-induced changes are linked to endothelial-derived NO in an ADPKD setting. Limitations of the present study include the lack of direct measurement of endothelial function, such as flow-mediated dilatation, and the measurement of other variables that could influence endothelial function, such as oxidative status. In summary, our results showed lower aerobic capacity in young normotensive patients with ADPKD with preserved kidney function and inadequate responses of NO, ADMA, and BP to acute exercise that further suggests the presence of even earlier endothelial dysfunction in this disease. However, further studies are imperative to investigate the effects of long-term exercise on these parameters and determine whether a training program could attenuate these alterations. Am J Kidney Dis. 2014;-(-):---
Exercise Capacity in ADPKD
ACKNOWLEDGEMENTS We thank Angela Tavares Paes for assistance in statistical analysis. This work was presented in part at the 49th European Renal Association–European Dialysis and Transplant Association Congress, May 24-27, 2012, Paris, France, and at the 45th Annual Meeting of the American Society of Nephrology, October 30November 4, 2012, San Diego, CA. Support: Financial support for this study was provided by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior and Fundação de Amparo a Pesquisa do Estado de São Paulo. Financial Disclosure: The authors declare that they have no other relevant financial interests.
REFERENCES 1. Braun WE. Autosomal dominant polycystic kidney disease: emerging concepts of pathogenesis and new treatments. Cleve Clin J Med. 2009;76(2):97-104. 2. Wilson PD, Goilav B. Cystic disease of the kidney. Annu Rev Pathol. 2007;2:341-368. 3. Torres VE, Harris PC. Autosomal dominant polycystic kidney disease: the last 3 years. Kidney Int. 2009;76(2):149-168. 4. Nishiura JL, Neves RF, Eloi SR, Cintra SM, Ajzen SA, Heilberg IP. Evaluation of nephrolithiasis in autosomal dominant polycystic kidney disease patients. Clin J Am Soc Nephrol. 2009;4(4):838-844. 5. Ecder T, Schrier RW. Cardiovascular abnormalities in autosomal-dominant polycystic kidney disease. Nat Rev Nephrol. 2009;5(4):221-228. 6. Gabow PA, Johnson AM, Kaehny WD, et al. Factors affecting the progression of renal disease in autosomal-dominant polycystic kidney disease. Kidney Int. 1992;41(5):1311-1319. 7. Chapman AB, Stepniakowski K, Rahbari-Oskoui F. Hypertension in autosomal dominant polycystic kidney disease. Adv Chronic Kidney Dis. 2010;17(2):153-163. 8. AbouAlaiwi WA, Takahashi M, Mell BR, et al. Ciliary polycystin-2 is a mechanosensitive calcium channel involved in nitric oxide signaling cascades. Circ Res. 2009;104(7): 860-869. 9. Nauli SM, Kawanabe Y, Kaminski JJ, Pearce WJ, Ingber DE, Zhou J. Endothelial cilia are fluid shear sensors that regulate calcium signaling and nitric oxide production through polycystin-1. Circulation. 2008;117(9):1161-1171. 10. Wang D, Iversen J, Wilcox CS, Strandgaard S. Endothelial dysfunction and reduced nitric oxide in resistance arteries in autosomal-dominant polycystic kidney disease. Kidney Int. 2003;64(4):1381-1388. 11. Wang D, Strandgaard S, Borresen ML, et al. Asymmetric dimethylarginine and lipid peroxidation products in early autosomal dominant polycystic kidney disease. Am J Kidney Dis. 2008;51(2):184-191. 12. Laughlin MH. Cardiovascular response to exercise. Am J Physiol. 1999;277(6, pt 2):S244-S259. 13. Maiorana A, O’Driscoll G, Taylor R, Green D. Exercise and the nitric oxide vasodilator system. Sports Med. 2003;33(14): 1013-1035. 14. Sarelius I, Pohl U. Control of muscle blood flow during exercise: local factors and integrative mechanisms. Acta Physiol (Oxf). 2010;199(4):349-365. 15. Martinez-Vea A, Bardaj A, Gutierrez C, et al. Exercise blood pressure, cardiac structure, and diastolic function in young normotensive patients with polycystic kidney disease: a prehypertensive state. Am J Kidney Dis. 2004;44(2):216-223.
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16. Pei Y, Obaji J, Dupuis A, et al. Unified criteria for ultrasonographic diagnosis of ADPKD. J Am Soc Nephrol. 2009;20(1): 205-212. 17. James PA, Oparil S, Carter BL, et al. 2014 Evidence-based guideline for the management of high blood pressure in adults: report from the panel members appointed to the Eighth Joint National Committee (JNC 8). JAMA. 2014;311(5):507-520. 18. Schrier RW, McFann KK, Johnson AM. Epidemiological study of kidney survival in autosomal dominant polycystic kidney disease. Kidney Int. 2003;63(2):678-685. 19. Grantham JJ, Torres VE, Chapman AB, et al. Volume progression in polycystic kidney disease. N Engl J Med. 2006;354(20):2122-2130. 20. Levey AS, Stevens LA, Schmid CH, et al. A new equation to estimate glomerular filtration rate. Ann Intern Med. 2009;150(9):604-612. 21. Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group. KDIGO 2012 clinical practice guideline for the evaluation and management of chronic kidney disease. Kidney Int Suppl. 2013;3(1):1-150. 22. Baecke JA, Burema J, Frijters JE. A short questionnaire for the measurement of habitual physical activity in epidemiological studies. Am J Clin Nutr. 1982;36(5):936-942. 23. Florindo AA, Latorre MRDO. Validação do questionário de Baecke de avaliação da atividade física habitual em homens adultos. Rev Bras Med Esporte. 2003;9:121-128. 24. Miller MR, Hankinson J, Brusasco V, et al. Standardisation of spirometry. Eur Respir J. 2005;26(2):319-338. 25. Neder JA, Nery LE, Castelo A, et al. Prediction of metabolic and cardiopulmonary responses to maximum cycle ergometry: a randomised study. Eur Respir J. 1999;14(6):1304-1313. 26. Balady GJ, Arena R, Sietsema K, et al. Clinician’s guide to cardiopulmonary exercise testing in adults: a scientific statement from the American Heart Association. Circulation. 2010;122(2):191-225. 27. Beaver WL, Wasserman K, Whipp BJ. A new method for detecting anaerobic threshold by gas exchange. J Appl Physiol. 1986;60(6):2020-2027. 28. Pescatello LS, Franklin BA, Fagard R, Farquhar WB, Kelley GA, Ray CA. American College of Sports Medicine position stand. Exercise and hypertension. Med Sci Sports Exerc. 2004;36(3):533-553. 29. Casa DJ, Armstrong LE, Hillman SK, et al. National Athletic Trainers’ Association position statement: fluid replacement for athletes. J Athl Train. 2000;35(2):212-224. 30. Pickering TG, Hall JE, Appel LJ, et al. Recommendations for blood pressure measurement in humans and experimental animals: part 1: blood pressure measurement in humans: a statement for professionals from the Subcommittee of Professional and Public Education of the American Heart Association Council on High Blood Pressure Research. Circulation. 2005;111(1):697-716. 31. Hampl V, Waters CL, Archer SL. Determination of nitric oxide by chemiluminescence reaction with ozone. In: Feelisch M, Stamler JS, eds. Methods in Nitric Oxide Reserarch. New York: John Wiley & Sons Ltd. 1996:309-318. 32. Schulze F, Wesemann R, Schwedhelm E, et al. Determination of ADMA using a novel ELISA assay. Clin Chem Lab Med. 2004;42:1377-1383. 33. Lee DC, Artero EG, Sui X, Blair SN. Mortality trends in the general population: the importance of cardiorespiratory fitness. J Psychopharmacol. 2010;24(4 suppl):27-35. 34. Svedahl K, MacIntosh BR. Anaerobic threshold: the concept and methods of measurement. Can J Appl Physiol. 2003;28(2):299-323.
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Reinecke et al 35. Hambrecht R, Fiehn E, Weigl C, et al. Regular physical exercise corrects endothelial dysfunction and improves exercise capacity in patients with chronic heart failure. Circulation. 1998;98(24):2709-2715. 36. Palmieri EA, Palmieri V, Innelli P, et al. Aerobic exercise performance correlates with post-ischemic flow-mediated dilation of the brachial artery in young healthy men. Eur J Appl Physiol. 2005;94(1-2):113-117. 37. Mendes-Ribeiro AC, Mann GE, de Meirelles LR, Moss MB, Matsuura C, Brunini TM. The role of exercise on L-arginine nitric oxide pathway in chronic heart failure. Open Biochem J. 2009;3:55-65. 38. Tinken TM, Thijssen DH, Hopkins N, Dawson EA, Cable NT, Green DJ. Shear stress mediates endothelial adaptations to exercise training in humans. Hypertension. 2010;55(2):312-318.
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39. Falcone JC, Kuo L, Meininger GA. Endothelial cell calcium increases during flow-induced dilation in isolated arterioles. Am J Physiol. 1993;264(2, pt 2):H653-H659. 40. Mo M, Eskin SG, Schilling WP. Flow-induced changes in Ca21 signaling of vascular endothelial cells: effect of shear stress and ATP. Am J Physiol. 1991;260(5, pt 2):H1698H1707. 41. Surdacki A, Nowicki M, Sandmann J, et al. Reduced urinary excretion of nitric oxide metabolites and increased plasma levels of asymmetric dimethylarginine in men with essential hypertension. J Cardiovasc Pharmacol. 1999;33(4):652-658. 42. Green DJ, Maiorana A, O’Driscoll G, Taylor R. Effect of exercise training on endothelium-derived nitric oxide function in humans. J Physiol. 2004;561(1):1-25.
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A
utosomal dominant polycystic kidney disease (ADPKD) is the most common renal genetic disorder; it affects 4-6 million individuals worldwide and accounts for w3% of end-stage renal disease.1,2 It is caused by mutations in the genes PKD1 or PKD2, which code for the transmembrane proteins polycystin 1 and polycystin 2, respectively. Functional deficiency in polycystin predisposes to the development of renal cysts and several renal and extrarenal manifestations, such as intracranial and aortic aneurysms, mitral valve prolapse, endothelial dysfunction, and hypertension.1,3,4 Hypertension, which is an early and common finding in patients with ADPKD, occurs in w60% of the patients before decreases in kidney function and is associated with a swift progression to end-stage renal disease.5,6 In addition to renal cystic expansion and intrarenal ischemia, endothelial dysfunction, found in Am J Kidney Dis. 2014;-(-):---
the early stages of the disease, also may play a role in the development of hypertension in ADPKD.7 The disruption of the normal function of polycystins, also expressed in vascular endothelial cells, results in an impaired shear stress sensitivity mechanism that may affect the synthesis of nitric oxide (NO), which results From the 1Renal Division and 2Pneumology Division, Department of Medicine, Federal University of São Paulo, São Paulo, Brazil. Received November 5, 2013. Accepted in revised form March 18, 2014. Address correspondence to Nestor Schor, MD, PhD, Department of Medicine, Renal Division, Federal University of São Paulo, Rua Botucatu, 740 CEP:04023-900 São Paulo–SP, Brazil. E-mail: [email protected] 2014 by the National Kidney Foundation, Inc. 0272-6386/$36.00 http://dx.doi.org/10.1053/j.ajkd.2014.03.014 1
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in lower systemic NO levels.8-10 Furthermore, levels of plasma asymmetric dimethylarginine (ADMA), an endogenous inhibitor of NO, also have been observed to be increased in patients with ADPKD compared with healthy controls.11 During physical exercise, several cardiovascular adjustments contribute to providing an increased oxygen supply to the muscles of healthy individuals. The higher production of endothelial NO stimulated by increased shear stress on vessel walls represents one such mechanism, which results in decreased total vascular peripheral resistance.12-14 Martinez-Vea et al15 showed an exaggerated diastolic blood pressure (BP) response during acute exercise in young normotensive patients with ADPKD, who presented with greater left ventricular mass index, which is consistent with early diastolic dysfunction. These findings prompted us to investigate the response patterns to acute exercise in an ADPKD sample consisting of young and normotensive patients with preserved kidney function and without left ventricular hypertrophy. Presently, there are scarce data available regarding the physical exercise response in patients with ADPKD. Because endothelial dysfunction might precede clinically detectable hypertension in ADPKD, we aimed to evaluate the physical capacity and the influence of acute aerobic exercise on serum and urinary NO and ADMA levels in the early stages of ADPKD in a group of normotensive patients with preserved kidney function.
METHODS Setting and Participants Forty-three adult patients with ADPKD being followed up at the Polycystic Kidney Disease Unit of the Nephrology Division at the Federal University of São Paulo were eligible for the study based on data from their medical records (all records [n 5 263] were analyzed). Inclusion criteria for patients with ADPKD were positive family history of ADPKD and renal cysts detected by ultrasound using the diagnostic criteria established by Pei et al,16 systolic BP , 140 mm Hg and diastolic BP , 85 mm Hg without antihypertensive medication,17 and estimated glomerular filtration rate (eGFR) . 60 mL/min/1.73 m2 in individuals aged 19-39 years. Only those who were sedentary and not involved in a physical training program were selected for the study. Exclusion criteria for patients with ADPKD were history of aneurysms; smoking; the presence of associated diseases such as diabetes, hyperlipidemia, and obesity; and use of medications that could interfere with cardiac or pulmonary function. Patients were informed about the study and invited to participate by personal contact. A total of 26 patients (18 women and 8 men) were enrolled in the study, and reasons for discontinuation are shown in Fig 1. Hemoglobin concentration and hematocrit, as well as echocardiography and ultrasonography results, were collected from patients’ medical records. Renal volume had been measured by ultrasound and calculated using a modified ellipsoid formula.18 The values for both kidneys were combined and divided by the individual’s height to yield total kidney volume.19 eGFR was 2
Medical records analyzed (n=263) Excluded by exclusion criteria (n=220)
Assessed for eligibility (n=43)
Scheduled for CPET
(n=13)
(n=30)
Unable to perform CPET (failed to pedal)
(n=2)
Declined to participate
Scheduled for exercise protocol
(n=28)
Discontinued participation (n=2)
Control group (n=30)
Completed protocol (n=26)
Figure 1. Patient enrollment flow chart. Abbreviation: CPET, cardiopulmonary exercise testing.
calculated using the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) creatinine equation,20 and the stages of chronic kidney disease were defined according to the KDIGO (Kidney Disease: Improving Global Outcomes) guidelines.21 The control group was recruited from laboratory staff and consisted of 30 adult age-matched apparently healthy nonobese sedentary individuals (9 men and 21 women). Exclusion criteria consisted of history of cardiovascular, pulmonary or kidney diseases; diabetes; smoking; hyperlipidemia; or hypertension. A fasting blood sample was collected for measurement of hemoglobin, hematocrit, and creatinine, determined by the alkaline picrate method. The reason that healthy individuals and not patients with chronic kidney disease were chosen as a control group was the exclusion of confounding factors for endothelial dysfunction. We obtained written consent from all participants. The local ethics committee approved the study.
Physical Activity Assessment The Baecke Habitual Physical Activity Questionnaire, which measures qualitative and quantitative indexes and addresses occupational, leisure, and locomotion activities,22 was used to assess physical activity performed during the past 12 months. This questionnaire has been validated in a Brazilian population survey.23
Pulmonary Function Test Spirometric tests24 were performed using the Clinical Pulmonary Function-Spirometry System (Medical Graphics Corp-MGC), and airflow was measured using a calibrated pneumotachograph. Participants completed at least 3 acceptable maximum forced expiratory maneuvers. The following variables were measured: forced vital capacity (FVC), forced expiratory volume in the first second of expiration (FEV1), and FVC:FEV1 ratio. Am J Kidney Dis. 2014;-(-):---
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Cardiopulmonary Exercise Testing
NO Determination
Cardiopulmonary exercise testing was conducted by a pneumologist, according to the protocol described by Neder et al.25 Oxygen uptake (VO2) measured at peak exercise (VO2peak) during incremental cardiopulmonary exercise testing is considered to be the gold standard for evaluation of exercise capacity.26 Exercise tests were conducted using an electromagnetically braked cycle ergometer (CPE 2000; Medical Graphics CorpMGC), with gas exchange and ventilator variables analyzed breath by breath using a calibrated computer-based exercise system (MGC-CPX System; Medical Graphics Corp). Cardiac electrical activity (CardioPerfect; MGC) and oxygen saturation measured by pulse oximetry (Ohmeda Biox 3740) were recorded continuously. The exercise test consisted of: (1) 2 minutes at rest, (2) 2 minutes of real zero-external intensity exercise obtained with the use of an electrical system that moved the ergometer flywheel at 60 rpm, (3) an incremental phase, and (4) a 4-minute recovery period. Power (watts) was increased continuously in a linear ramp pattern (10-20 W$min21) such that the test duration was longer than 8 but shorter than 12 minutes. The individual pedaled to the limit of tolerance with active and standardized encouragement from the same investigator. Average pulmonary VO2 (in liters per minute) for the last 30 seconds of the ramp was considered to be representative of the participant’s VO2peak. VO2 at the anaerobic threshold was estimated by the gas exchange threshold method (V-slope).27 All tests were performed in the morning in the same laboratory with temperature controlled at 21 C-24 C.
Serum and urine nitrate concentrations were measured using an NO analyzer (NOA TM280; Sievers Inc).31 Serum and urinary samples were previously deproteinized in zinc sulfate, 10% (1:2), and sodium hydroxide (1:2) for 15 minutes and centrifuged for 5 minutes. A 10-mL aliquot of the supernatant was injected into the analyzer. A calibration curve was plotted for standards of sodium nitrate. In the NO analyzer, nitrate was reduced to NO with vanadium at 90 C, and the NO formed was detected by gaseousphase chemiluminescence after reaction with ozone. The analyzer remained stable, and linearity was maintained during the test period to #1% with a coefficient of variation of #10%. Serum results are expressed as micromoles per liter. Urinary NO excretion was corrected by urinary creatinine and expressed as micromoles per milligram of creatinine.
Exercise Protocol After a minimum 1-week period, participants underwent a 20-minute single bout of cycle ergometer exercise. Exercise intensity was measured by the anaerobic threshold (AT) according to the following formula: D 5 VO2peak (watts) – AT (watts), and Intensity 5 (D 3 0.3) 1 AT (watts). This intensity corresponds to 60% of VO2peak, which was considered to be moderate.28 During the exercise, patients were advised to ingest 200-300 mL of water according to recommendations of the National Athletic Trainers’ Association.29
BP Measurements All participants arrived at the laboratory after an 8-hour fasting period with no water intake restriction. A light snack (25 g of salt cracker and 200 mL of peach juice) was given to participants, followed by a 30-minute rest in seated position. Before starting the exercise, we measured BP by auscultation (sphygmomanometer and stethoscope; Diasyst LTDA) 3 times according to the American Heart Association procedure30 and took the mean value. We also measured BP during exercise (every 3 minutes) and after exercise (every minute until 5 minutes and subsequently, every 3 minutes until 30 minutes after exercise). All BP measurements were made by a single person in order to minimize possible errors. We measured heart rate continuously before, during, and after exercise using an oximeter.
Biochemical Assays All participants fasted for 8 hours overnight and were instructed to avoid foods that could affect the biochemical assays for at least 48 hours before blood and urine collections. Venous blood samples were obtained from the forearm at rest and at the end of the exercise protocol. Urine samples were collected at rest and after the exercise protocol. Samples were centrifuged at 4 C for 20 minutes at 2,000 rpm. The supernatant was removed with a polypropylene pipette, aliquoted into 0.5-mL samples in polypropylene tubes, and stored at 280 C for NO and ADMA measurements. Am J Kidney Dis. 2014;-(-):---
ADMA Determination We assessed plasma ADMA concentration using an ADMA enzyme-linked immunosorbent assay (ELISA) kit validated by Schulze et al32 (Immundiagnostik AG), according to the manufacturer’s instructions. Standard samples and controls were added and incubated overnight at 2 C-8 C. After several washings with a wash buffer, the conjugate was added and incubated for 1 hour at room temperature. Thereafter, the washing step was repeated and the substrate was added and incubated for 25 minutes at room temperature on an orbital shaker. Stop solution was added and immediately analyzed using an ELISA reader at 450nm.
Statistical Analyses We performed statistical analyses using SPSS 19.0 for Windows (SPSS Inc). Data are expressed as mean 6 standard deviation. The data were verified for normality by descriptive statistics, such as comparison of the mean versus the median, histogram and boxplot inspection, and skewness and kurtosis coefficients. The t test was used to compare clinical, spirometric, and cardiopulmonary exercise testing variables of the studied groups. Repeated-measures analysis of variance with Bonferroni post-test was used to analyze NO, ADMA, BP, and heart rate responses to exercise in both groups. Pearson linear correlation was used to establish relationships between variables. Statistical significance was defined as P , 0.05. Comparison of categorical data was performed by c2 test.
RESULTS Baseline Clinical and Laboratory Characteristics Estimated time from the diagnosis of ADPKD until our study ranged from 1-192 months. Total kidney volume adjusted for the participant’s height averaged 175-1,760 (median, 280) mL/m. Based on eGFR, 23 patients were classified as chronic kidney disease stage 1, and 3 patients, as stage 2; mean eGFR was 113 6 17 [standard deviation] mL/min/1.73 m2. Liver size was described as normal for all patients according to computed tomographic results available in their medical records (data not shown). None of these patients presented with microalbuminuria (mean albumin excretion, 9.8 6 5 mg/min). Echocardiographic parameters retrieved from patients’ medical records, such as mean ejection fraction (66% 6 3%) and interventricular septum (7.9 6 0.8 mm), were within the expected range for healthy individuals. No patient presented with left ventricular mass hypertrophy; mean left ventricular mass index was 74.8 6 17 g/m2. 3
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Table 1 lists clinical characteristics of patients and controls. No statistical difference was found with respect to mean age, sex, body mass index, BP at rest, eGFR, hematocrit, hemoglobin level, level of physical activity, and parameters of pulmonary function (FVC, FEV1, and FEV1:FVC ratio). Cardiopulmonary Exercise Testing Cardiopulmonary exercise testing results are listed in Table 2. VO2peak, VO2 at anaerobic threshold, maximal workload, anaerobic threshold workload, respiratory exchange ratio, and oxygen pulse were significantly lower in patients with ADPKD than in controls. We observed no statistical differences between groups with respect to peak carbon dioxide production, respiratory rate, maximum heart rate, end-tidal oxygen partial pressure, or end-tidal carbon dioxide partial pressure. No significant correlations were found between height-adjusted total kidney volume and the exercise parameters VO2peak (r 5 0.08; P 5 0.7), VO2 at Table 1. Baseline Clinical Characteristics, Physical Activity, and Pulmonary Function Parameters
Variable
Sex Female Male
Control (n 5 30)
ADPKD (n 5 26)
21 9
18 8
P
0.9
Age (y) 28.9 6 7.3 30.0 6 8.0 Weight (kg) 68.1 6 14.2 69.1 6 16.3 Height (cm) 169.0 6 9.0 170.0 6 10.0 BMI (kg/m2) 23.5 6 2.8 23.7 6 4.7 SBP (mm Hg) 110.0 6 11.0 116.0 6 12.0 DBP (mm Hg) 71.0 6 9.0 76.0 6 11.3 eGFR (mL/min/1.73 m2) 112 6 9.6 113 6 17 Hematocrit (%) 41.1 6 3.2 41 6 4 Hemoglobin (g/dL) 13.4 6 0.6 13.9 6 1.4 Microalbuminuria — 9.8 6 5 (mg/min) Height-adjusted TKV — 280 (175; 1,760) (mL/m) HPAQ score 7.3 6 1.0 7.4 6 1.9 FVC (L) 4.1 6 0.8 4.1 6 0.9 FEV1 (L) 3.4 6 0.6 3.4 6 0.7 FEV1:FVC ratio (%) 85.3 6 6.2 83.2 6 6.4 Baseline serum NO 53.9 6 3.1 48.3 6 4.6 (mmol/L) Baseline plasma ADMA 0.47 6 0.02 0.45 6 0.02 (mmol/L)
0.6 0.9 0.6 0.9 0.09 0.2 0.7 0.9 0.4
— —
0.8 0.9 0.9 0.3 0.1 0.8
Note: Values for categorical variables are given as number; values for continuous variables are given as mean 6 standard deviation or median (minimum; maximum). Abbreviations: ADMA, asymmetric dimethylarginine; ADPKD, autosomal dominant polycystic kidney disease; BMI, body mass index; DBP, diastolic blood pressure; eGFR, estimated glomerular filtration rate; FEV1, forced expiratory volume in the first second of expiration; FVC, forced vital capacity; HPAQ, Habitual Physical Activity Questionnaire; NO, nitric oxide; SBP, systolic blood pressure; TKV, total kidney volume. 4
anaerobic threshold (r 5 0.01; P 5 0.9), and maximal workload (r 5 20.07; P 5 0.8) in the ADPKD group. Exercise Protocol BP Responses Exercise systolic and diastolic BP responses are shown in Fig 2A. Systolic and diastolic BP during exercise and recovery were significantly greater in patients with ADPKD versus controls. Heart Rate Heart rates during exercise and recovery, shown in Fig 2B, did not differ between groups. Nitric Oxide At baseline, serum and urinary NO levels did not differ between groups. However, postexercise NO levels increased in controls compared to baseline (serum: 63.1 6 1.9 vs 53.9 6 3.1 mmol/L [P 5 0.01]; urinary excretion: 61.4 6 10.6 vs 38.7 6 5.6 mmol/mg creatinine [P 5 0.01]), but did not increase in patients with ADPKD (serum: 45 6 5.1 vs 48.3 6 4.6 mmol/L [P 5 0.9]; urinary excretion: 39.8 6 6.8 vs 34.7 6 6.5 mmol/mg creatinine [P 5 0.8]; Fig 3A and B). In the control group only, there was a negative correlation between serum NO level after exercise and ADMA level after exercise (r 5 20.60; P 5 0.01). Asymmetric Dimethylarginine Basal ADMA values did not differ between groups. However, ADMA levels decreased in controls after exercise versus baseline (0.39 6 0.02 vs 0.47 6 0.02 mmol/L; P 5 0.006), but did not decrease in patients with ADPKD (0.47 6 0.04 vs 0.45 6 0.02 mmol/L; Table 2. Cardiopulmonary Exercise Testing Parameters
Variable
Control (n 5 30)
ADPKD (n 5 26)
P
VO2peak (mL/kg/min) 31.0 6 4.8 22.2 6 3.3 ,0.001 VCO2peak (mL/min) 2,254.2 6 727.2 1,990.1 6 690.4 0.2 VO2AT (L/min) 957.4 6 300.8 743.6 6 220.8 0.01 MW (W) 156.5 6 50.8 114.4 6 30.2 0.002 ATW (W) 70.9 6 21.2 47.9 6 10.2 ,0.001 RERmax 1.17 6 0.1 1.25 6 0.1 0.007 RR (rpm) 38.7 6 10.8 35.7 6 9.1 0.3 HRmax (beats/min) 174.0 6 18.0 174.9 6 14.0 0.9 11.4 6 3.9 9.0 6 2.7 0.02 VO2/HRmax PETO2 (mm Hg) 102.9 6 5.2 102.9 6 8.0 0.9 PETCO2 (mm Hg) 34.5 6 4.5 34.4 6 6.0 0.9 Note: Values are given as mean 6 standard deviation. Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; ATW, anaerobic threshold workload; HRmax, maximum heart rate; MW, maximal workload; PETCO2, end-tidal carbon dioxide partial pressure; PETO2, end-tidal oxygen partial pressure; RERmax, maximum respiratory exchange ratio; RR, respiratory rate; VCO2peak, peak carbon dioxide production; VO2AT, oxygen uptake at anaerobic threshold; VO2peak, peak oxygen uptake; Vo2/HRmax, maximum oxygen uptake to heart rate ratio. Am J Kidney Dis. 2014;-(-):---
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Figure 2. (A) Blood pressure (BP) and (B) heart rate (HR) at rest and during and after exercise. Values are expressed as mean 6 standard error (SE). *P , 0.05 versus control. Abbreviations: ADPKD, autosomal dominant polycystic kidney disease; DBP, diastolic BP; SBP, systolic BP.
P 5 0.6; Fig 3C). No correlations were found between ADMA level and kidney size or NO level and kidney size.
DISCUSSION The present study showed that young normotensive patients with ADPKD with preserved kidney and cardiac function have impaired physical capacity, evidenced by the lower VO2peak and anaerobic threshold compared with healthy individuals with the same previous level of physical activity assessed by the Baecke Habitual Physical Activity Questionnaire. Considered gold-standard measures of cardiorespiratory fitness, VO2peak and anaerobic threshold also are predictors of all-cause and cardiovascular disease mortality.33,34 The normal spirometry parameters (FVC and FEV1) and liver size and the absence of significant correlations between height-adjusted TKV Am J Kidney Dis. 2014;-(-):---
and exercise parameters (VO2peak, VO2 at anaerobic threshold, and maximal workload) suggest that liver and/or kidney size did not interfere with patients’ physical capacity results. Additionally, the ejection fraction and serum hemoglobin levels within normal range suggested adequate oxygen transport in patients with ADPKD. According to Hambrecht et al,35 systemic vasoconstriction and diminished peripheral perfusion can lead to exercise intolerance. In this context, NO plays a key role in vasomotor tone and perfusion regulation. It has been suggested that higher submaximal and maximal aerobic exercise performance is associated with greater flow-mediated dilatation of peripheral conduit arteries in young healthy men.36 Thus, higher peripheral vascular resistance could impair exercise capacity. In the present series, baseline serum and urinary NO levels in patients with ADPKD were similar to 5
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Figure 3. (A) Serum nitric oxide (NO) levels, (B) urinary NO excretion, and (C) plasma asymmetric dimethylarginine (ADMA) levels. Values are expressed as mean 6 standard error (SE). Abbreviation: ADPKD, autosomal dominant polycystic kidney disease.
levels in controls. Although these findings contrast with those from the study of Wang et al,11 who demonstrated impaired production of NO in normotensive patients with ADPKD, their patients were older and presented with microalbuminuria, a feature not shared by the present sample and that could indicate that their patients already had a higher rate of endothelial dysfunction. However, under the exercise stimulus, serum NO levels did not increase in the ADPKD group as expected and observed in the control group, thus demonstrating disturbed NO regulation. Urinary NO excretion also increased after 6
exercise in only the control group, probably due to the higher production during exercise. Higher NO production during physical exercise is due to exercise-induced increases in endothelial shear stress,37,38 which is necessary to increase blood flow and the influx of ionized calcium in endothelial cells, releasing NO.39,40 In the present study, the lack of increase in NO levels, accompanied by a nondecrease in serum ADMA levels after exercise, strongly suggests impaired capacity for exercise-induced vasodilatation. Our findings agree with data published by Wang et al,11 who found higher serum ADMA levels in patients with ADPKD. These investigators suggested that the increased oxidative stress in patients with ADPKD might contribute to the increased ADMA levels by decreasing its metabolism by dimethylarginine dimethylaminohydrolase (DDAH). However, as described for NO, ADMA levels did not differ from the control group at baseline, but only after exercise. Surdacki et al41 suggested that high ADMA levels would lead to diminished NO bioavailability, thereby increasing vascular resistance and BP. Martinez-Vea et al15 observed an exaggerated diastolic BP response during exercise in young normotensive patients with ADPKD, who showed greater left ventricular mass index, consistent with early diastolic dysfunction. In the present study, the inadequate systolic and diastolic BP responses during exercise evident in patients with ADPKD with preserved cardiac function further reinforce the hypothesis of higher vascular resistance leading to defective vasodilatation. This altered NO response likely may be influenced by the mutated polycystins at endothelial and smooth vascular cells because they are required for an appropriate mechanism of blood shear stress by a complex biochemical cascade involving calcium, calmodulin, Akt (also known as protein kinase B [PKB]), and protein kinase C.8,9 Nevertheless, because NO has various nonvascular sources and its systemic levels are only indirect evidence for its role in skeletal muscle exercise hyperemia,42 more studies are necessary to identify whether these exercise-induced changes are linked to endothelial-derived NO in an ADPKD setting. Limitations of the present study include the lack of direct measurement of endothelial function, such as flow-mediated dilatation, and the measurement of other variables that could influence endothelial function, such as oxidative status. In summary, our results showed lower aerobic capacity in young normotensive patients with ADPKD with preserved kidney function and inadequate responses of NO, ADMA, and BP to acute exercise that further suggests the presence of even earlier endothelial dysfunction in this disease. However, further studies are imperative to investigate the effects of long-term exercise on these parameters and determine whether a training program could attenuate these alterations. Am J Kidney Dis. 2014;-(-):---
Exercise Capacity in ADPKD
ACKNOWLEDGEMENTS We thank Angela Tavares Paes for assistance in statistical analysis. This work was presented in part at the 49th European Renal Association–European Dialysis and Transplant Association Congress, May 24-27, 2012, Paris, France, and at the 45th Annual Meeting of the American Society of Nephrology, October 30November 4, 2012, San Diego, CA. Support: Financial support for this study was provided by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior and Fundação de Amparo a Pesquisa do Estado de São Paulo. Financial Disclosure: The authors declare that they have no other relevant financial interests.
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