Acute effects of intradialytic aerobic exercise on solute removal, blood gases and oxidative stress in patients with chronic kidney disease

Correspondência para: Joseane Böhm. Hospital de Clínicas de Porto Alegre. Rua Ramiro Barcelos, no 2350, Porto Alegre, RS, Brasil. CEP: 90035-003 E-mail: josebohm@bol.com.br Esta pesquisa foi realizada com o financiamento do Fundo de Incentivo à Pesquisa e Eventos do Hospital de Clínicas de Porto Alegre, Porto Alegre, Brasil, e do Fundo de Pesquisa do Instituto de Doenças Renais Ltda., Porto Alegre, Brasil. Efeitos agudos do exercício aeróbio intradialítico sobre a remoção de solutos, gasometria e estresse oxidativo em pacientes com doença renal crônica


Introduction:
Hemodialysis contributes to increased oxidative stress and induces transitory hypoxemia.Compartmentalization decreases the supply of solutes to the dialyzer during treatment.The aim of this study was to investigate the acute effects of intradialytic aerobic exercise on solute removal, blood gases and oxidative stress in patients with chronic kidney disease during a single hemodialysis session.Methods: Thirty patients were randomized to perform aerobic exercise with cycle ergometer for lower limbs during 30 minutes with intensity between 60-70% of maximal heart rate, or control group (CG).Blood samples were collected prior to and immediately after exercise or the equivalent time in CG.Analysis of blood and dialysate biochemistry as well as blood gases were performed.Mass removal and solute clearance were calculated.Oxidative stress was determined by lipid peroxidation and by the total antioxidant capacity.Results: Serum concentrations of solutes increased with exercise, but only phosphorus showed a significant elevation (p = 0.035).There were no significant changes in solute removal and in the acid-base balance.Both oxygen partial pressure and saturation increased with exercise (p = 0.035 and p = 0.024, respectivelly), which did not occur in the CG.The total antioxidant capacity decreased significantly (p = 0.027).Conclusion: The acute intradialytic aerobic exercise increased phosphorus serum concentration and decreased total antioxidant capacity, reversing hypoxemia resulting from hemodialysis.The intradialytic exercise did not change the blood acid-base balance and the removal of solutes.

IntRoductIon
Hemodialysis contributes to increased oxidative stress, producing free radicals and reducing the levels of antioxidant enzymes in patients with end-stage renal disease (ESRD). 1 During hemodialysis, the compartmentalization of solutes reduces its blood levels and dialysis efficiency by decreasing the supply to the dialyzer thus impairing their removal. 2emodialysis also decreases oxygen partial pressure (PaO 2 ), increases minute ventilation due to production and excretion of carbon dioxide (CO 2 ), and increases oxygen consumption as a result of metabolic alkalosis. 3,4The hypoxemia generated during hemodialysis due to the sequestration of intrapulmonary leukocytes decreases cardiac output and induces microatelectasis as a result of the procedure. 36][7][8][9] However, the acute effects of exercise during dialysis have not been adequately studied so far.It is possible that acute exercise could have detrimental effects in the short term, such as increased oxidative stress and decreased production of antioxidant enzymes, aggravating the clinical condition of the patients. 10herefore, the aim of this study was to investigate the acute effects of intradialytic aerobic exercise on solute removal, blood gases and oxidative stress in ESRD using a cycle ergometer for lower limbs during a hemodialysis session.

ParticiPants
Thirty patients with clinically stable ESRD who underwent hemodialysis were randomized by the Random Allocation Software 1.0™ (Isfahan University of Medical Sciences, Isfahan, Iran) into intervention group (IG) or control group (CG).Data from medical records and patient history and physical examination were used.
Inclusion criteria were: adults older than 18 years of age, diagnosis of chronic kidney disease (CKD) for at least six months and having hemodialysis treatment for more than three months, physical and clinical stability for performing aerobic exercise, not maintaining regular or having total absence of physical activity, and not having participated in other research related to physical activity in the last six months.As exclusion criteria, presence of acute myocardial infarction three months before the start of the study, unstable angina, hemodialysis vascular access in the lower limbs, hemoglobin level < 10 g/dL, presence of active systemic infection or inflammatory process, or candidate for imminent kidney transplantation with a living donor.
The study protocol was approved by the Research Ethics Committee of the IPA Methodist University Center and Hospital de Clínicas de Porto Alegre, where the study was conducted and performed according to the 1975 Declaration of Helsinki.All participants provided written informed consent prior to enrollment.

Dialysis settings
All patients were dialysed with bicarbonate dialysis, thrice weekly for 4 h with a low-flux polysulphone hollow-fiber dialyser (Diacap™ LOPS 20, Laboratórios B.Braun S. A., São Gonçalo, RJ, Brazil).Hemodialysis sessions were performed with single use of dialyzers, and all patients had a native arteriovenous fistula as vascular access.Blood flow rates ranged between 300 and 350 mL/min.The dialysate flow rate was 500 mL/min.The blood flow and dialysate flow rates were kept constant throughout the study period in the individual patient.

intervention
The intervention was performed in the beginning of the second hour of the third weekly day of hemodialysis.The CG had two blood collections separated by 30-minute interval controlled by the dialysis machine timer on a single hemodialysis session.The IG was compared to the CG and as its own control.The IG was studied contemporaneous with the CG and in the subsequent week, with a blood collection prior to and following 30 minutes of aerobic exercise.The exercise was conducted with a cycle ergometer for lower limbs (Dream EX 150 FLEX ® , Dream Indústria e Comércio Ltda., Esteio, RS, Brazil) coupled to the dialysis chair.
Exercise intensity was set between 60-70% of maximal heart rate, calculated using the formula proposed by Karvonen et al.,11 or intensity between 13-14 points on the Borg's Rating of Perceived Exertion Scale (Borg's RPE Scale).Heart rate was measured using the monitor HR 102 Oregon Scientific™ (Oregon Scientific, São Paulo, SP, Brazil).The interventions were carried out by the same researcher.

ColleCtions anD blooD samples
Blood samples were drawn from the arterial line of hemodialysis vascular access, after reducing the blood flow pump to 50 mL/minute.A sample (3 mL) was collected in a syringe washed with heparin for blood gases analysis.A second sample (5 mL) was collected in a plain tube for solute measurements.A third sample (5 mL) was collected in a plain tube with EDTA, centrifuged at 1900 rpm for 10 minutes at 4 °C, for plasma separation and storage in conical tubes at -80°C for the analysis of oxidative stress.

ColleCtion of Dialysate
The dialysate eliminated by the dialysis equipment during the 30-minute intervention or equivalent time in CG was collected in a container with storage capacity of 20 liters.After homogenization of the content and measurement of the total volume, a sample was separated and aliquoted into two 5-mL collection tubes for analysis.To calculate the clearance of solutes, we used the formula described by Maher. 12

bioChemiCal analysis
The analysis of serum creatinine, urea, potassium, phosphorus, magnesium, dialysate and blood gases were performed by automation.For analysis of lipid peroxidation (malondialdehyde, MDA) thiobarbituric acid reactive substances (TBARS) and colorimetric assay at 540 nm (TBARS Assay kit Cayman™, Cayman Chemical Company, Ann Arbor, Michigan, USA) were used.Analysis of the total antioxidant capacity (TAC) of plasma was based on the ability of antioxidants sample to inhibit the oxidation of the compound 2,2′-azinobis-(3-ethylbenzothiazoline-6sulfonic acid, ABTS) by metmyoglobin.The amount of ABTS produced was read by a colorimetric assay at 540 nm, and compared with Trolox (Antioxidant Assay kit Cayman™, Cayman Chemical Company, Ann Arbor, Michigan, USA).

sample size CalCulation
The sample estimate was performed using the software G*Power™ version 3.1.9.2 (Department of Psychology, Kiel University, Kiel, Germany).The sample size was calculated based on mean and standard deviation of plasma MDA concentration post-hemodialysis in IG and plasma MDA concentration in CG according to the study of Ozden et al. 13 Using a two-tailed test and an effect size of 2.18 obtained in the reference study, it was defined a total of 15 patients in each group to get a power of 80% and a significance level (a) of 5%.

statistiCal analysis
The Kolmogorov-Smirnov test was used to assess the normality of data distribution.Differences between variables were calculated using nonparametric tests (Wilcoxon or Mann-Whitney) or parametric tests (independent samples t test or paired samples t test), as applicable.Data were expressed as mean ± standard error or median and interquartile range.A p < 0.05 was considered statistically significant.The data were analyzed using the program Statistical Package for the Social Sciences™ (SPSS version 19.0, Chicago, Illinois, USA).

Results
The clinical characteristics of the groups are presented in Table 1.In both groups there is a predominance of male and white patients.There were no significant differences between groups with respect to age, Body Mass Index (BMI), duration of hemodialysis treatment, and to Charlson Comorbidity Index (CCI).Five patients underwent previous renal transplantation (4 CG, 1 IG), 17 were former smokers and three patients were active smokers.The predominant etiology of ESRD was diabetes mellitus, followed by hypertension and chronic glomerulonephritis.Seven patients had ESRD from unknown etiology.No patient reported discomfort when performing the exercise.
Comparisons between the differences in measures of the groups, before and after an interval of 30 minutes, are shown in Table 2.With the implementation of aerobic exercise, there was a significant increase in the oxygen partial pressure and saturation [4.30 ± 2.15 mmHg vs. -6.55± 1.90 mmHg, p = 0.001 and 1.1% (0.1% to 2%) vs. -0.6%(-2.8% to -0.1%), p = 0.003, respectively].
Table 3 shows comparisons between the differences of the measures before and after an interval of 30 minutes in the IG, with and without exercise performance.In the hemodialysis session with exercise patients showed an increase in serum phosphorous concentration [0.2 mg/dL (-0.3 mg/dL to 0.37 mg/dL) vs. -0.2mg/dL (-0.4 mg/dL to 0 mg/ dL), p = 0.035], increased oxygen partial pressure and saturation [2.71 ± 1.81 mmHg vs. -0.76± 1.17 mmHg, p = 0.037 and 0.3% (-0.3% to 1.1%) vs.
-0.2% (-0.8% to 0.5%), p = 0.024, respectively] and decreased TAC (-0.165 ± 0.12 mM vs. 0.355 ± 0.2 mM, p = 0.027).Table 4 presents the total dialysate volume, solute concentration measurements, removed mass and solute clearances in the dialysate of 9 individuals of the IG.Compared to baseline, there were no statistically significant differences in any parameter evaluated after the exercise performance.

dIscussIon
In this study, we found that acute intradialytic aerobic exercise increased serum phosphorus concentration, decreased the total antioxidant capacity, and reversed the hypoxemia resulting from hemodialysis.The oxygen partial pressure and saturation increased with exercise, which did not occur in the control group.However, the intradialytic exercise did not change acutely the blood gas parameters or even increased the removal of solutes.
We showed a significant increase in serum phosphorus after aerobic exercise in the IG.Phosphate is predominantly an intracellular ion, which moves into the cell as the extracellular pH increases. 14][16][17] Hemodialysis performed with bicarbonate dialysate favors alkalemia, which can hinder phosphate removal.This fact emphasizes the potential of proposed exercise of mobilizing phosphorus from body compartments, increasing its blood concentration.Furthermore, this increase in phosphorus concentration came to accomplish the intervention in the third weekly hemodialysis session where the patient has already shows a reduction in the concentration of solutes due to treatment.Modulation of the intensity and duration of exercise in a hemodialysis session may also be necessary to further promote increases in serum concentration of solutes mediated by a decrease in blood pH induced by the exercise.
Kirkman et al., 17 performing 60 minutes of intradialytic exercise with intensity of 90% of the lactate threshold, showed that this exercise protocol was more effective than the increase in dialysis time for phosphate removal, being an adjuvant therapy for serum phosphorus control.Farese et al. 15 also found an increase in the phosphate mass and clearance in dialysate after three exercise sessions lasting 20 minutes at 36 rpm.
In our study, aerobic exercise increased the serum concentration of other solutes (but without reaching statistical significance) but with no absolute  changes in solute clearance.Intradialytic aerobic exercise promotes increased blood flow to the central vasculature and increased vascular permeability, allowing greater area of exchange between the intracellular and intravascular compartments.][20][21] Orcy et al. 16 found an increase in the mass removal and phosphate clearance only in dialysate after three sessions of intradialytic aerobic exercise lasting 40 minutes with an intensity between 13-14 points on the Borg's RPE Scale.Changes in blood concentration of solutes have not been found in this study because the samples were collected prior to and after the hemodialysis procedure.The duration of intradialytic exercise we proposed was sufficient to increase solute blood concentrations, but as the solute removal occurs across the entire time of hemodialysis (four hours in average), it would be necessary to measure them again at the end of dialysis treatment, and not only at the end of the exercise.
Analyzing the results of blood gases post-exercise, we observed a statistically significant increase in the oxygen partial pressure and saturation, reversing hypoxemia induced by hemodialysis.Meyring-Wösten et al. 22 described a significant association between hypoxemia and adverse clinical outcomes, most notably all-cause of hospitalization and mortality.Hemodialysis with bicarbonate lowers the PaO 2 in its initial minutes and reaches the nadir between 30 and 60 minutes, remaining at this level throughout the dialysis procedure and after its completion. 3,23][25] Moore et al. 26 performed five minutes of intradialytic aerobic exercise in each hour of treatment with intensity at 60% of maximal oxygen consumption (VO 2max ) to assess the hourly PaO 2. These authors did not observe changes in the PaO 2 probably because the training volume was low.Burke et al. 27 conducted a moderate intensity exercise protocol during hemodialysis treatment, and Germain et al. 28 prescribed aerobic exercise of low intensity for three hours during dialysis.Both groups found an increase in PaO 2 and increased alveolar oxygen pressure during exercise.The intensity and duration of exercise that we applied generated enough hyperventilation to increase the diffusion of oxygen, but this did not decrease the alkalinization of blood, because the partial pressure of CO 2 remained unchanged.
The ability of the body to resist oxidative changes depends on the levels of antioxidant enzymes such as superoxide dismutase, glutathione peroxidase, and catalase, as well as of nonenzymatic antioxidants.TAC is a valuable tool for understanding the ability of the biological system to oppose oxidative stress. 29n our study, we observed a statistically significant reduction in TAC after the exercise.Previous studies performed without exercise [29][30][31] showed significant reduction in TAC after a hemodialysis session that was attributed in large part to a correlation of their elimination with a decrease in uric acid concentration and antioxidant vitamins removed by dialysis.Furthermore, additional loss of antioxidants is frequent and caused by the removal of water soluble antioxidants due to their low molecular weight. 30The increased blood flow promoted by aerobic exercise may have contributed to the additional loss of antioxidants.
This study has limitations, including the small number of patients in each comparison group.Moreover, for better assessment of maximal heart rate training to adjust exercise prescription it would be appropriate to implement a standardized exercise stress test.The analysis of uric acid concentration could have been carried out in order to verify its parallel relationship with the reduction of TAC.
Finally, to verify the effectiveness of hemodialysis on solute removal and the influence of exercise on the outcome of the dialysis process, it would be necessary to perform blood and dialysate collection at the end of hemodialysis session, in addition to the end of exercise.
In conclusion, moderate intradialytic aerobic exercise acutely reversed hypoxemia induced by hemodialysis and increased serum phosphorus levels.Despite this, no changes occurred on solute removal.The exercise did not promote acute changes in the acid-base balance, but decreased the total antioxidant capacity of these patients.
of Basic Sciences of Health of Universidade Federal do Rio Grande do Sul.We would also like to thank Howard Haimes, Sandro Maciel da Silva, and John Widden for reviewing the manuscript.

tAble 1 the
clinical characteristics of the grouPs Data expressed as mean ± standard error or median (interquartile range).BMI: body mass index; CCI: Charlson comorbidity index; CG: control group; IG: intervention group; ESRD: end-stage renal disease.

tAble 2
comParisons between the diferences (d) in measures of the grouPs, before and after an interval of 30 minutes

tAble 3
comParisons between the diferences (d) in measures before and after an interval of 30 minutes, with and without exercise Performance, in intervention grouP(n = 15)