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Clinical Sciences: Clinically Relevant

Effects of long-term training on the progression of chronic renal failure in rats

BERGAMASCHI, CÁSSIA T.; BOIM, MIRIAN A.; MOURA, LUIS A.; PIÇARRO, IVAN C.; SCHOR, NESTOR

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Medicine & Science in Sports & Exercise: February 1997 - Volume 29 - Issue 2 - p 169-174
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Abstract

The life quality of hemodialysis patients is in many aspects markedly precarious. These patients often have reduced levels of work capacity and motivation for physical activity. Many factors contribute to the reduced work capacity including myopathy, myocardial dysfunction, anemia, arterioscleroses, psychosocial abnormalities, and sedentary life style(9).

Regular exercise elicits alterations in cardiovascular function, metabolism, neurohormonal activity, and coronary risk factors in healthy subjects (22). Endurance exercise training increases maximal oxygen consumption by raising maximal cardiac output as well as increasing the arteriovenous oxygen difference (27). Moreover, it has been effective in the treatment of patients with coronary artery disease, hypertension, hyperlipidemia, diabetes, obesity, and depression(1,13,17,25,27,36), reducing the number of risk factors for cardiovascular disease by increasing HDL cholesterol, reducing total cholesterol and triglycerides, lowering blood pressure, improving cardiac performance, enhancing glucose metabolism, and reducing stress (17,22,25). Training has also improved bone density, increased total red cell mass, increased muscle mass and strength, and reduced depression. Thus, endurance training may have a potential ameliorating effect on the severity of many of the metabolic and cardiovascular abnormalities in patients with chronic renal failure. Some groups have studied physical activity as a therapeutic modality for chronic renal failure. Goldberg et al. (14-16), in a long-term study of the therapeutic effects of endurance exercise in hemodialysis patients, have shown a marked improvement in lipoprotein lipid and glucose profiles, normalized insulin sensitivity, increased red cell mass and hemoglobin concentration, and reduced blood pressure. Training has also improved psychosocial function, contributing to the social rehabilitation of dialysis patients (6).

Despite these satisfactory results, the mechanisms involved in these beneficial effects of exercise training have not been fully established. To better understand this adaptation, as well as to evaluate whether exercise can modify the progression of renal failure, we evaluated renal and glomerular hemodynamics, proteinuria, and glomerular sclerosis in a model of chronic renal failure in rats submitted to long-term exercising.

METHODS

Studies were performed on adult male Munich-Wistar rats weighing 250-300 g. Animals were allowed free access to water and a standard rat pellet diet.

The animals were divided into 4 groups: Control (N = 7), rats with 75 d of chronic renal failure (CRF) and maintained sedentary(CRF,N = 11), SHAM plus exercise (SHAM+EX, N = 9), and rats with 75 d of CRF and submitted to 60 d of exercise (CRF+EX, N = 19).

The rats were placed in individual metabolic cages and 24 h urine was collected to determine urinary protein excretion measured by precipitation with 3% sulfosalicylic acid. Urinary protein excretion was measured at the beginning of the protocol and before the hemodynamic study. CRF was obtained by submitting the animals to 5/6 renal mass ablation by ligation of two or three branches of the left renal artery and total right nephrectomy as previously described (7).

To determine the training level, CRF+EX and SHAM+EX animals were submitted to a maximum progressive work test for determination of maximum oxygen consumption (˙VO2max). We used a rapid-flow, open-circuit system developed by Brooks and White (5) to determine˙VO2max. The O2 and CO2 analyses were performed with Beckman OM-11 and LB-2 analyzers (Beckman Instruments, Inc., IL), respectively. The values for maximum oxygen consumption obtained were used to determine the training level based on relative work load (65-75%˙VO2max). The ˙VO2max test was performed only at the beginning of the protocol and the workload measured at this time was maintained during all training period. The training consisted of running on a motordriven treadmill for 30 min·d-1, 5d·wk-1 for 60 d, starting 15 d after nephrectomy. Rats in the sedentary groups were also submitted to the same test to assure that all animals were in the same basal conditions.

GENERAL PROTOCOL

In the training groups the kidney function was measured at minimum 2 d after the last exercise session to avoid any acute effect of the exercise. After 75 d of CRF induction and/or exercise training, rats were anesthetized with Inactin, 100 mg·kg-1 body wt, i.p. (BYK Gulden, Konstanz, Germany), placed on a temperature-regulated micropuncture table, and monitored with a thermometer to maintain their rectal temperature between 36.5 and 37.5°C. After tracheotomy, the left femoral artery was catheterized with a PE-50 polyethylene tube and approximately 60 μl of arterial blood was collected for baseline hematocrit (Hct) and plasma protein (CA) determinations. This artery was also used for periodic blood sampling and estimation of mean arterial blood pressure (MAP). Polyethylene catheters were also inserted into the right and left jugular veins. Saline solution containing inulin (10%) and p-aminohippuric acid (2%) (PAH) was immediately infused into the right side at a rate of 1.2 ml·h-1 and maintained throughout the experiment to determine GFR and renal plasma flow(RPF). Isoncotic rat serum was infused initially at a rate of 10 ml·kg-1·h-1 into the left jugular vein to replace surgical losses, and at 1.5 ml·kg-1·h-1 to maintain animals under euvolemic conditions. Laparotomy was performed and a PE-10 polyethylene tube was introduced into the left ureter for collection of timed urine samples. To prepare the left kidney for micropuncture, the posterior face of the kidney was dissected free, held on a small plastic shovel, and positioned to avoid interference by diaphragm movements.

The initial study period was started after a 30-45 min equilibration period. Two urine samples and blood samples were collected to determine Hct, CA, inulin, and PAH concentration in plasma and urine.

Samples of fluid from surface proximal tubules were collected for determination of flow rate and inulin concentration, allowing the calculation of single nephron glomerular filtration rate (SNGFR). Hydraulic pressures were measured in surface glomerular capillaries (PGC), proximal tubules(PT), efferent arterioles (PEA), and peritubular capillaries(PC) with a continuous recording servonull micropipette transducer system (IPM Inc., San Diego, CA). To estimate the pre- and post-glomerular colloid osmotic pressures, protein concentration in femoral arterial and efferent arteriolar plasma were measured as described previously(10). These estimations permitted the calculation of single nephron filtration fraction (SNFF), initial glomerular capillary plasma flow rate (QA), afferent (RA), efferent (RE), and total(RT) arteriolar resistance, and glomerular capillary ultrafiltration coefficient (Kf), using equations previously described(10,11).

Analytical procedures. Plasma and urinary inulin concentrations were determined by the macroanthrone method of Fuhr et al.(12). PAH concentration was determined by the Bratton and Marshal method modified by Smith et al. (30). Inulin concentration in tubular fluid was measured by the microfluorescence method(34). Protein concentration in efferent arteriolar and femoral arterial plasma was determined by the fluorimetric method(33). Whole kidney filtration fraction (FF) was calculated as the GFR/RPF ratio.

At the end of the functional studies, the kidneys were perfused with a solution containing 2% glutaraldehyde buffered in phosphate, pH 7.2-7.4, at constant 100 mm Hg pressure, for morphological evaluation. Glomerulosclerosis was defined as the absence of cellular elements in the tuft, collapse of capillary lumens, and folding of the glomerular basement membrane with amorphous material, while mesangial matrix expansion was characterized by the presence of increased amounts of periodic acid-Schiff-positive material in the mesangial region (29). The percentage of sclerotic glomeruli was determined by the ratio (number of damaged glomeruli/total number of glomeruli examined × 100).

Statistical analysis was performed by one-way analysis of variance with the Duncan, Mann Whitney, and Kruskal Wallis tests. Significance was defined asP < 0.05. Data are presented as mean ± SE.

RESULTS

Table 1 summarizes the results of ˙VO2max(measured at the beginning of the protocol) and whole kidney function. Maximum oxygen consumption (˙VO2max), body weight (BW), plasma protein concentration (CA), and hematocrit (Hct) did not differ among groups. Mean arterial pressure (MAP) was higher in the CRF and CRF+EX groups than in the CONTR and SHAM+EX groups (160 ± 5 and 167 ± 9 vs 123± 4 and 126 ± 5 mm Hg, P < 0.05, respectively).

A significant decrease in GFR was observed in the CRF and CRF+EX animals compared with control rats (1.13 ± 0.04 vs 0.46 ± 0.06 and 0.44± 0.05 ml·min-1, P< 0.05). A similar decrease in RPF (3.44 ± 0.30 vs 1.73 ± 0.24 and 1.56 ± 0.13 ml·min-1; P < 0.05) occurred in the nephrectomized groups compared with the control group. Since the decrease in GFR and RPF was proportional, the FF was maintained.

Table 2 summarizes the results of superficial single nephron function for all groups. SNGFR and QA were significantly higher in the CRF group when compared with the CONTR group (47.54 ± 6.79 vs 32.14 ± 2.09 nl·min-1 and 137.13 ± 17.78 vs 94.65± 11.15 nl·min-1, respectively, P < 0.05). In the CRF+EX group there was a nonsignificant elevation in SNGFR (32.14± 2.09 vs 45.51 ± 5.45 nl·min-1) with a significant increase in QA (94.65 ± 11.15 vs 147.90 ± 14.49 nl·min-1, P < 0.05). The SNFF was unchanged. The pre- and postglomerular resistances remained unchanged in the CRF group compared to the control group (Table 2). However, a significant decrease in postglomerular resistance was observed in the CRF+EX group when compared with the CRF group (1.35 ± 0.14 vs 0.88 ± 0.06 1010dyn·s-1·cm-5, P < 0.05).

Trained animals with CRF showed a significant increase in Kf when compared with the control group (0.0675 ± 0.0146 vs 0.1764 ± 0.0246 nl·s·mm Hg-1, P < 0.05).

CRF caused a significant increment in PGC (43 ± 1 vs 48± 1 mm Hg, P < 0.05) and PT (13 ± 1 vs 17± 1 mm Hg, P < 0.05). The association of exercise and CRF induced a decrease in PGC compared with sedentary CRF animals (48± 1 vs 42 ± 1 mm Hg, P < 0.05). PT (15± 1 vs 17 ± 1 mm Hg) did not change.

Table 3 summarizes the results of proteinuria and glomerular sclerosis index for all groups. The CRF and CRF+EX groups presented a marked increase in proteinuria, 3.18 ± 0.38 in control vs 66.88± 41.15 in CRF and 76.73 ± 31.53 mg·24h-1 in CRF+EX (P < 0.05) accompanied by elevation in sclerosis index (3± 1 and 19 ± 3 vs 16 ± 4%, P < 0.05). No difference was observed between the CRF and CRF+EX groups.

DISCUSSION

Physical exercise is recommended as a parallel therapy in many pathologies. In chronic renal failure (CRF) physical training is being used in patients submitted to hemodialysis (14,15,26). However, little information is available about the influence of exercise on the evolution of renal disease.

Despite the tremendous potential of endurance exercise training in the rehabilitation of hemodialysis patients, its effects on renal hemodynamics have not been as well studied in this population as they have been in other pathological states, perhaps because of the difficulty in motivating these patients to exercise. Thus, this study was undertaken to evaluate the potential effect of long-term controlled exercise on glomerular hemodynamics and on the renal microcirculation and also to determine if these maneuvers can modify the progression of renal failure.

It has been reported that in normal man acute exercise induces profound changes in renal hemodynamics and in electrolyte and protein excretion. Effective renal plasma flow is reduced during exercise and produces a concomitant effect on glomerular filtration rate. The combination of sympathetic nervous activity and the release of catecholamine substances is involved in this process (26).

˙VO2max indicated that all animals were under the same aerobic conditions during endurance training. Moreover, the animals also had a similar basal aerobic conditions at the beginning of the protocol.

In the present study no significant alterations in general or whole kidney parameters were observed between the CONTROL and the SHAM+EX groups. This may suggest that, differently from acute exercise, long-term physical exercise did not change the parameters evaluated. It is possible that for this group of rats a longer period is necessary to observe an effect of exercise on normal renal function. Moreover, it has been described that during exercise the GFR is maintained unless the exercise intensity is increased above of 50% of the˙VO2max(27,28). However, in normal conditions this change is transient and the GFR should come back to normal levels after a few hours.

In the CRF group without training, results were similar to those expected for the stage of the renal disease (75d). GFR presented a 59% reduction with also a 50% reduction in RPF, indicating a loss of renal function. On the other hand, SNGFR and QA were significantly increased in the CRF group, indicating an adaptive process with hyperflow and hyperfiltration of the superficial nephrons, as previously described(4,20). The glomerular hypertension also suggests this adaptation, which was accompanied by proteinuria and glomerular sclerosis.

Considering whole kidney function, the CRF animals submitted to physical training showed the same profile as that observed for sedentary animals, i.e., reduction in GFR and RPF with no change in FF. All the remaining hemodynamic parameters showed results similar to those of the CRF sedentary animals. Increases in GFR with exercise have been described by Knochel et al(21) in healthy males who submitted to hard physical work in hot climates. In our study exercise did not improve the GFR in rats with chronic renal failure, but we also did not find any changes in sham operated animals that were submitted to exercise. This result may be related to the training load, which was moderated, as we had planned it to be. An interesting alteration was observed in arteriolar resistance since a reduction in efferent arteriolar resistance (RE) was observed in CRF+EX animals, and thus a normal PGC was achieved in this group. Thus, CRF animals submitted to training did not show glomerular hypertension, a potential factor involved in the progression of renal failure. In spite of the normalization of PGC, proteinuria and glomerular sclerosis were not different from those observed in the sedentary CRF group, indicating no correlation between these parameters in this model. Also it is important to note that, despite the reduction in PGC, no change in SNGFR was observed in the training CRF group, probably owing to a compensatory increase in QA and Kf.

In fact, glomerular hypertension is often considered to be the one of the main causes of the evolution of CRF (20). Anderson et al.(2) demonstrated that the control of glomerular hypertension in nephrectomized rats provoked a delay in the progression of CRF, preserved renal morphology, and reduced proteinuria. It was also suggested that, after renal mass ablation, CRF progresses as a result of an increase in both glomerular perfusion and glomerular pressure. Considering this observation, the remaining nephrons compensate for the loss of renal mass by hyperfiltrating leading to glomerular sclerosis. However, recent data have suggested that dissociation of the correlation between hyperfiltration and glomerular sclerosis occurs in certain experimental models(31,35). Studies of glomerular hemodynamics in different hypertensive models have demonstrated that glomerular hyperperfusion alone was not associated with any significant glomerular structural alteration. In contrast, similar levels of SNGFR and QA were associated with severe glomerular damage when glomerular hypertension was also present, indicating that the critical factor for the development of structural injury is the rise in glomerular pressure (19). These studies suggest that the increase in glomerular capillary wall tension is translated into metabolic alterations which cause excessive deposition of extracellular matrix, mesangial expansion, and sclerosis (8). Despite this evidence, the role of glomerular hypertension as the single causative factor has been questioned since some forms of glomerular sclerosis are not followed by an increase in PGC and, therefore, other mechanisms may be responsible for the progression of renal failure (8).

Heifets et al. (18), studying rats with 1.5 subtotal nephrectomy and subjected to swimming for 2 months, demonstrated that exercise training improves GFR, decreases the degree of proteinuria, and reduces the extent of glomerulosclerosis. However, exercise did not alter the degree of systemic hypertension. In contrast, we were unable to show improvement in whole kidney function, proteinuria, or glomerulosclerosis in CRF animals submitted to exercise. These differences may be a result of differences in age, type, and/or length of exercise training.

On the other hand, interestingly, our results showed that the physical training provoked a vasodilatation in the efferent arteriole, inducing a reduction in PGC. It is suggested that many vasoactive agents take part in this situation such as prostaglandins, the renin-angiotensin system, and the sympathetic nervous system, among others. During physical activity catecholamine levels are elevated acutely to promote compensatory body adjustments and to contribute to metabolic processes such as glycogenolysis, gluconeogenesis, glucose uptake, contractility, and chronotropic and inotropic actions of the heart, as well as circulatory changes(22). In contrast, chronic physical training promotes a reduction in catecholamines levels both in man and in rats(23,24). Also Tidgren et al.(32) demonstrated that during exercise angiotensin II is elevated proportionally to the training intensity and is involved in the elevation of renal vascular resistance. The increase in norepinephrine and renin production suggests a possible interaction between the sympathetic nervous system and the renin angiotensin system, inducing a reduction in plasma renin activity. However, training can cause an increase in prostacyclin production or sensitivity (3). Therefore, this complex interaction of many opposite vasoactive hormones plays a role in the adaptation induced by exercise (28).

The present study suggests that exercise is not able to modify the progression of renal disease. However, training led to the normalization of glomerular hypertension, known to be an important factor involved in decreasing glomerular function. The present data suggest a dissociation between PGC and the progression of renal failure in the model of 5/6 nephrectomy in adult rats during the period of time studied.

On the other hand, although it did not ameliorate CRF, exercise did not cause any injury. Thus, it is reasonable to prescribe exercise in this situation to improve the general clinical condition.

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Keywords:

CHRONIC RENAL FAILURE; EXERCISE TRAINING; RENAL AND GLOMERULAR HEMODYNAMICS

©1997The American College of Sports Medicine