The Impacts of Aquatic Walking and Land-based Walking Exercise Therapy Programs on Vascular Function, Cardiorespiratory Capacity, Exercise Tolerance, Muscular Strength, and Physical Function in Patients With Peripheral Artery Disease
Phase: NA
Conditions: Peripheral Artery Disease, Vascular Function, Exercise Tolerance, Peripheral Arterial Disease
Sponsor: Pusan National University

Document Date: 2/05/2020 
 
The Impacts of Aquatic vs Land Walking on Vascular Health and Exercise Tolerance in 
Patients with Peripheral Artery Disease 
 
NCT#03849300 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

Full study protocol (Control vs. Aquatic walking exercise group 1) 
 
Participants 
Participants were recruited with flyers and clinic referrals in a metropolitan city in South 
Korea at multiple community health centers where the members were primarily female. Study 
allocation was determined randomly with a computerized random number generator (Figure 1). 
All participants were female and classified as Fontaine stage I or II PAD with an ABI of 0.7 – 0.9. 
All participants were sedentary, defined as participating in under 1 hour of regular exercise 
participation per week within the previous year. Exclusion criteria included current smoker 
(smoking within ≤ 6 months), psychiatric conditions, as well as pulmonary, renal, and thyroid 
diseases. These exclusion criteria for these diseases were included to avoid any secondary disease 
effects on the target population used in the present study. All procedures were conducted in 
accordance with the protocols approved by the Institutional Review Board designated by Ministry 
of Health and Welfare (PNU IRB-2016_92_HR) and carried out in accordance with the 
Declaration of Helsinki, and all subjects provided written, informed consent prior to experimental 
measures. All protocols and lab procedures took place at the community centers. This study was 
registered with clinicaltrials.gov (NCT03849300). 
Study design 
This study used a 2-armed parallel experimental design. Arterial stiffness, resting HR, BP, 
ABI, VO2max, 6MWD, physical function scores, hand grip strength, leg strength, anthropometrics, 
RMR, and flexibility were assessed at baseline and after the 12 weeks. All measurements were 
performed at the same time of day (8:00 AM, ±1 h) after a 12-hour fast. The VO2max test and 6-
minute walk test were separated by a period of ~24 hours to prevent confounding effects of fatigue. 
Following baseline measurements, participants were randomly allocated to either the aquatic 

walking training group (AQ, n = 35) or the control group (CON, n = 37). The AQ group 
participated in a supervised aquatic walking training program for 12 weeks that included a warm-
up, main exercise session, and a cool-down. The CON group did not participate in any exercise 
program or perform any additional exercise. The CON group participants were present and 
supervised in the laboratory at the same frequency and time of day as the AQ group (10:00AM, 
±1h) and performed sedentary activities such as reading books, listening to music, or learning new 
computer skills, which are among some of the activities these participants normally perform when 
attending the community center daily. The AQ and CON groups were advised to refrain from 
changing any dietary habits throughout the study period, and diet logs were given to researchers 
weekly to monitor caloric intake to ensure there were no changes in diet throughout the 
intervention. If researchers noticed differences in caloric intake, they advised participants to 
maintain their normal diet. Lunch and dinner were provided to participants at the community 
centers. All participants were supervised by qualified trainers and measurements were taken by 
experienced researchers, and researchers who performed the laboratory measurements were 
blinded to the randomization of subjects.  
Exercise program  
Participants in the AQ group participated in an aquatic walking exercise training program 
for 12 weeks, 4 days per week, for 60 minutes per day (Table 1), and this was adapted from 
previous literature for older individuals and patients with PAD (3, 47). Sessions were conducted 
in waist-to-chest-deep water (28-30°C) in a group setting (3). Participants were allowed to use 
flotation devices as needed for safety purposes. Participant heart rate (HR) was monitored using a 
wearable Polar HR monitor (Electro, Oy, Kempele, Finland) during all exercise sessions to 
maintain the appropriate training intensity. The aquatic walking exercise training program 

intensity was established using heart rate reserve (HRR) and Borg’s revised rating of perceived 
exertion scale (RPE, 0 to 10 scale). HRR was calculated as (HRR = [% intensity desired * (HRmax 
– HRrest)] + HRrest). The intensity of the program increased every 4 weeks: weeks 1-4 were at 50-
60% HRR and 6-8 RPE, weeks 5-8 were at 60-70% HRR and 6-8 RPE, and weeks 9-12 were at 
70-85% HRR and 6-8 RPE. Each training session was supervised, and each trainer was assigned 
to work with 10 participants during the sessions. HR monitors and RPE were checked every 5 
minutes during the sessions. If the HR or RPE were too low or too high according to the intensity 
level, the participants were encouraged to either increase or decrease their effort during the 
exercise training session. 
The program was divided into 3 sections including a warm-up (10 min), main exercise 
component (40 min), and a cool-down (10 min) (Table 1). Both the warm-up and cool-down 
included underwater stretching that emphasized lower limb muscles (quadriceps, hamstrings, hip 
abductors/adductors, and hip flexors) and low-intensity gait training (forward, backward, and 
lateral movement) (3). The main exercises were performed for a total of 40 minutes. The first 10 
minutes of main exercises included simple movement patterns such as hip flexion-extension, hip 
abduction-adduction, and knee flexion-extension (3). The next 30 minutes of the main exercise 
session consisted of water walking exercises (forward, backward, lateral) (3).  
Arterial stiffness, HR, BP, and ABI 
Participants rested in a supine position for 5 minutes. Femoral-to-ankle pulse wave velocity 
(legPWV), a measurement of peripheral arterial stiffness, was measured using applanation 
tonometry (TU-100 and VP-2000, OMRON Healthcare, Kyoto, Japan). A pulse wave sensor was 
placed on the femoral artery, while the cuffs with pulse wave sensors on the ankles and EKG 
electrodes remained in place from the ABI measurement. BP, EKG, and pulse waveforms were 

simultaneously recorded for 10-30 seconds. Data analysis was performed according to the Clinical 
Application of Arterial Stiffness, Task Force III (69). 
Participants rested in a seated position for 5 minutes. Resting HR, systolic BP, and diastolic 
BP were measured in duplicate using radial artery palpation and an automated sphygmomanometer 
(BP-200, OMRON, Kyoto, Japan), respectively, before and after 12 weeks. The average of the two 
measurements was recorded as the resting values. If the measurements differed by >5 mmHg, an 
additional measurement was taken and the two closest readings (differed by ≤2 mmHg) were 
averaged (50). The interclass correlation coefficient in our laboratory for systolic and diastolic BP 
calculated on two measurements is ≈0.97. 
Participants rested in a supine position for 5 minutes. ABI was measured using the VP-
2000 (OMRON, Kyoto, Japan). Electrocardiogram (EKG) electrodes were placed on the forearms. 
BP cuffs with pulse wave sensors were wrapped around both arms (brachial artery) and ankles 
(posterior tibial artery). Measurements were recorded for 10-30 seconds.  
Cardiorespiratory capacity and 6-minute walking distance 
The Cornell modified Bruce treadmill test was used to determine the maximal volume of 
O2 consumption (VO2max) (32). This Bruce protocol modification has been proven to be valid and 
reproducible (48) and has been used in disease populations (76). The modified Bruce consists of 
11, 2-minute stages that progress from 1.7 mph and 0% grade to 5 mph and 18% grade (48).  The 
treadmill test was ended when the participants reached volitional fatigue. Expired gases were 
measured with a metabolic cart (OxyCon Pro, Viasys Healthcare, Conshohocken, PA).   
The 6-minute walk test was used to determine maximal walking distance, or exercise 
tolerance. Participants were asked to walk as many laps as possible around a 100-meter track for 
6 minutes.  

Physical function 
Physical function was assessed collectively using physical function questionnaires from 
the Medical Outcomes Study Short-Form 36 General Health Survey (MOS SF-36) (73). Other 
parameters (i.e. not physical function domain) were not assessed using the MOS-SF 36. Physical 
function questionnaires of the MOS SF-36 are valid and reliable surveys that are often used for 
assessing physical function in patients with PAD (22, 24, 66, 73).  
Muscular strength 
Upper body muscular strength was determined using maximal isometric handgrip strength 
with a standard handgrip dynamometer (JAMAR, Bolingbrook, IL). Measurements of the 
dominant hand were taken 3 times, and the best of the 3 trials was recorded as the measurement.  
Lower body muscular strength was assessed using one-repetition maximum (1RM) on a 
leg extension machine (Cybex 6000; Lumex, Albertson, NY). The 1RM was measured with the 
dominant leg, and the 1RM was achieved within 5 or fewer attempts. The highest weight lifted 
using the proper form was recorded as the 1RM. 
Anthropometrics 
Anthropometric measurements, including height, total body mass, body mass index (BMI), 
and body composition were measured before and after 12 weeks. Height was measured using a 
standard stadiometer to the nearest 0.1 cm. Body composition was estimated using bioelectrical 
impedance analysis (BIA) (InBody 230, Biospace, Seoul, Korea), which simultaneously recorded 
total body mass (nearest 0.1 kg), and body fat (nearest 0.1%) (60). BMI was calculated as body 
mass divided by the height squared (kg/m2).  
Resting metabolic rate 

 
Resting metabolic rate was estimated using a metabolic cart (OxyCon Pro, Viasys 
Healthcare, Conshohocken, PA). Subjects remained in the supine position and were instructed to 
breathe normally and to not fall asleep during the measurement, and the measurements were 
performed for 30 minutes.  
Flexibility 
 
Flexibility was measured using the YMCA sit-and-reach test. Each participant was given 
3 attempts to reach as far as possible with proper form, and the best of the 3 trials was recorded as 
the score.  
Statistical analysis plan 
 
The Shaprio-Wilk test was used to determine data normality. Independent t tests were used 
to determine any baseline differences between the AQ and CON groups. A two-way analysis of 
variance (ANOVA) with repeated measures [group (AQ and CON) x time (pre- and post-12 
weeks)] was used to determine the difference of changes between pre- and post-aquatic walking 
training program within and between groups on the dependent variables. When a significant 
interaction was noted, paired t tests were used for post-hoc comparisons. All analyses were 
performed using SPSS 24 (SPSS Inc. Chicago, IL, USA). Data are presented as Mean ± SD. 
Statistical significance was set to p < 0.05. Additionally, correlations between variables were 
assessed with Pearson product-moment correlations. A power analysis calculation was used to 
determine a minimum sample size of 64 (32 each group) would allow for the observation of a 
difference of 3-5% between groups (AQ vs. CON) for PWV with a power of 90% (4, 47).  
 
 
 
 
 

Full study protocol (Aquatic walking exercise group 2 vs. Land-based walking exercise 
group) 
 
Participants (mean age 65±10, n = 53) were recruited with flyers, referrals, and newspaper 
advertisements in a metropolitan city in South Korea at multiple community health centers where 
the membership population was primarily female. Study allocation was determined randomly 
using a computerized random number generator (Figure 1) to the HWET group or LET group. All 
participants were postmenopausal females (cessation of menses for at least 12 months), sedentary, 
and classified as Fontaine stage II or III PAD with an ABI of 0.6 – 0.8. Sedentary lifestyle was 
termed as taking part in ˂1 hour of regular exercise per week within the previous year. Exclusion 
criteria included stage IV PAD (tissue loss/gangrene), previous or pending revascularization 
procedure, hormone replacement therapy, current smoker (smoking within the previous 6 months), 
psychiatric conditions, as well as pulmonary, renal, and thyroid diseases. These exclusion criteria 
were incorporated to avoid potential secondary disease effects on the target population examined 
in this study. All exercise and experimental methods were conducted in accordance with approved 
protocols by the Institutional Review Board. All participants provided written, informed consent 
and all procedures were performed in accordance with the Declaration of Helsinki. This study was 
registered with clinicaltrials.gov (NCT03849300). 
Study design 
A 2-armed parallel experimental design was used (Figure 2). Anthropometrics, vascular 
function, resting metabolic rate, walking capacity, COT, and muscular strength were assessed at 
baseline and after the 12 weeks. All lab assessments were performed at the same time of day (8:00 
AM, ±1 h) following an overnight fast. All participants were asked to abstain from medications 
for at least 24 hours prior to the baseline and post-exercise therapy measurements but continued 
taking medications throughout the exercise therapy protocols according to their physician 

instructions. After baseline measurements, participants were allocated randomly to the HWET 
group (n = 28) or the LBET group (n = 25). The HWET group participated in a supervised heated-
water exercise therapy program for 12 weeks. The LBET group participated in supervised 
treadmill therapy program for the same period of time. Both exercise groups were asked to refrain 
from altering dietary habits throughout the study period, and researchers obtained weekly diet logs 
from each participant to audit caloric intake. The exercise therapy sessions and experimental 
measures took place at the community centers. All participants were supervised by qualified 
trainers during exercise therapy sessions, and measurements were taken by experienced 
researchers who were blinded to participant randomization.  
Exercise programs 
The HWET group participated in a supervised heated-water exercise therapy program 
(Table 1) for 12 weeks, 4 days per week, for 60 minutes per day. The 12-week duration, exercise 
frequency, and intensity levels were programmed to be similar with previous water-based and 
land-based studies that demonstrated improvements in vascular function for older individuals and 
patients with PAD (3, 45, 54, 56, 87). Exercise sessions were performed in waist-to-chest-deep 
water (30-31°C). HR was monitored using a standard chest strap Polar HR monitor (Electro, Oy, 
Kempele, Finland) during each exercise session to manage the appropriate training intensity level. 
Intensity was programmed using heart rate reserve (HRR) and Borg’s revised rating of perceived 
exertion scale (RPE, 0 to 10 scale). HRR was calculated as (HRR = [% intensity desired * (HRmax 
– HRrest)] + HRrest). Exercise intensity was increased every 4 weeks: weeks 1-4 were at 50-60% 
HRR and 6-8 RPE, weeks 5-8 were at 60-70% HRR and 6-8 RPE, and weeks 9-12 were at 70-85% 
HRR and 6-8 RPE. Exercise sessions were monitored by qualified trainers. Chest HR monitor 
readings and RPE were checked every 5 minutes during the sessions. To maintain the appropriate 

intensity, if the HR or RPE were too low or too high according to the exercise program, the trainer 
encouraged participants to increase or decrease their effort during the session. The HWET program 
was divided into 3 components including a warm-up, main exercise section, and a cool-down. The 
warm-up and cool-down included underwater stretching that emphasized major muscle groups and 
low-intensity gait training (forward, backward, and lateral movement) (3). The main exercises 
were performed for a total of 40 minutes. For the first 10 minutes of the main section, participants 
performed lower limb movement patterns including hip flexion-extension, hip abduction-
adduction, and knee flexion-extension (3). The next 30 minutes of the main exercise session 
included water walking (3).  
The LBET group performed a supervised walking exercise program on a treadmill. The 
warm-up and cool-down components included stretches similar to those in the HWET group, and 
the therapy program also lasted 12 weeks, 4 times per week, for 60 minutes per session (Table 2). 
The main exercise component consisted of simple movement patterns for 10 minutes that included 
low-intensity forward, backward, and lateral side-stepping movements on flat ground. The next 30 
minutes included treadmill walking at the same intensity (HRR and RPE values) as in the HWET 
group. Target HRR and RPE were achieved by adjusting the treadmill speed and incline grade. 
Participants with leg symptoms were encouraged to exercise to near maximal leg symptoms until 
failure of walking.  
Anthropometrics 
Measurements of height, total body mass, and body composition were measured before 
and after 12 weeks for the LBET and HWET groups. Height was measured using a standard 
stadiometer to the nearest 0.1 cm. Body composition was assessed using bioelectrical impedance 
analysis (BIA) (InBody 230, Biospace, Seoul, Korea). Total body mass (nearest 0.1 kg) and body 

fat (nearest 0.1%) were recorded. Body mass index (BMI) was calculated as body mass divided 
by the square of height (kg/m2).  
Vascular function 
With participants in the supine position, ankle-brachial index (ABI) was recorded using the 
VP-2000 (OMRON, Kyoto, Japan). Electrocardiogram (EKG) electrodes were placed on the 
forearms. BP cuffs with pulse wave sensors were wrapped around both arms (brachial artery) and 
ankles (posterior tibial artery). Measurements were recorded for 10-30 seconds. Brachial-to-ankle 
pulse-wave velocity (baPWV) was also recorded using the VP-2000 as previously described (89).  
Femoral-to-ankle pulse-wave velocity (legPWV), a measurement of peripheral arterial 
stiffness, was measured using applanation tonometry (TU-100 and VP-2000, OMRON Healthcare, 
Kyoto, Japan) (56). A tonometer was placed on the femoral artery, while the pulse-wave sensor 
BP cuffs on the ankles and EKG electrodes remained in place from the ABI measurement. BP, 
EKG, and pulse waveforms were simultaneously recorded for 10-30 seconds. Data analysis was 
performed according to the Clinical Application of Arterial Stiffness, Task Force III (77).  
Participants were then moved to a resting seated position. After 5 minutes of quiet sitting 
the resting HR, systolic BP, and diastolic BP were measured in duplicate using radial artery 
palpation and an automated sphygmomanometer (BP-200, OMRON, Kyoto, Japan), respectively, 
before and after 12 weeks of LBET or HWET. The average of the two measurements was recorded 
as the resting values. If the BP measurements differed by more than 5 mmHg, an additional 
measurement was taken and the two readings that were the closest (differed by ≤2 mmHg) were 
averaged and recorded as the resting BP (58). The interclass correlation coefficient in our 
laboratory for systolic and diastolic BP calculated on two measurements is ≈0.97. 
Resting metabolic rate 

Resting metabolic rate (RMR) was calculated using a metabolic cart (OxyCon Pro, Viasys 
Healthcare, Conshohocken, PA). Subjects rested in the supine position on a padded table and were 
asked to breathe normally and to not fall asleep during the assessment. Measurements with the 
metabolic cart were performed for 30 minutes.  
Exercise tolerance and claudication assessment 
The 6-minute walk test was used to determine 6-minute walking distance, or exercise 
tolerance. Participants were asked to walk as many laps as possible around a 60-meter track for 6 
minutes. COT was measured during the 6-minute walk test. Once the participants experienced any 
leg and/or foot pain during the walking test, they reported the pain to the trainers. 
Muscular strength 
Upper body muscular strength was quantified using a standard handgrip dynamometer to 
assess maximal isometric handgrip (JAMAR, Bolingbrook, IL). Measurements of the dominant 
hand were taken 3 times. The best value of the 3 trials was recorded as the hand grip strength value. 
Lower body muscular strength was quantified using leg extension one-repetition maximum (1RM) 
(Cybex 6000; Lumex, Albertson, NY). The 1RM was assessed using the dominant leg, which was 
attained within 5 or fewer attempts. The highest weight lifted with proper form and full range of 
motion was recorded as the 1RM.   
Physical function 
Physical function was assessed collectively using physical function questionnaires from 
the Medical Outcomes Study Short-Form 36 General Health Survey (MOS SF-36) (83). This is a 
valid and reliable survey that is often used for patients with PAD (27, 30, 75, 83). Other parameters 
(i.e. not physical function domain) were not assessed using the MOS-SF 36.  
 
 

Statistical analysis plan 
 
The Shaprio-Wilk test was used to assess normality of the data. Independent t tests were 
used to determine any baseline differences between the LBET and HWET groups. A two-way 
repeated measure analysis of variance (ANOVA) [group (LBET and HWET) x time (pre- and post-
12 weeks)] was used to assess changes between pre- and post- LBET vs. HWET within and 
between groups on the dependent variables. When a significant interaction was found, paired t 
tests were used for post-hoc comparisons. All analyses were performed using SPSS 24 (SPSS Inc. 
Chicago, IL, USA). Statistical significance was set to p < 0.05. Data are presented as Mean ± SD. 
A power analysis calculation was used to determine the minimum sample size of 50 (25 in each 
group) would allow for the observation of a change of 3-5% between groups (LBET vs. HWET) 
for legPWV with a power of 90% (4, 54).