Cecchini C

CLERICO, ALDO

Cruschelli L

Giammattei C

Giampietro O.

Gregori G

Lucchetti A

Penno G

Archivio della ricerca della Scuola Superiore Sant'Anna

CV,%fl’T0 20 40 60 80 100Albumin, mg/LCLIN. CHEM. 36/3, 562-564 (1990)562 CLINICAL CHEMISTRY, Vol.36, No. 3, 1990Exercise-Induced Proteinuria in Well-Trained AthletesAldo Clerico, Carlo Glammattel, Lulgi Cecchinl, Amalla Lucchetti, Lorella Cruscheill, Gluseppe Penno, Giovanna Gregori, andOttavle GiampletroWe studied the rate of urinary excretion of albumin, a1-microglobulin (as an indicator of the renal tubular involve-ment), sodium, potassium, and creatinine in the basal state(overnight urine collection) and after physical exercise (train-ing session) in 10 professional cyclists, to verify whetherproteinexcretionis increased even in well-trained athletesafter physical effort. In addition, we wanted to understandwhether the origin of exercise-induced proteinuria was gb-merular, tubular, or both. Compared with the basal state(overnight coection), exercise significantly (P <0.01) in-creased the excretion rate of albumin (4.2 ± 2.6 g/min vs18.1 ± 10.6 p.g/min, mean ± SD), Na, and K, and also theurinary volume. Creatinine output was not affected by exer-cise. The mean (±SD) overnight excretion rate of albumin byathletes was quite similar to that found for 91 healthynonathletes at rest (4.6 ± 2.7 g/min). The mean exercise-related excretion of a1-microgbobulin by the athletes signifi-cantly exceeded the overnight value (6.6 vs 0.3 mg/L, P =0.037). Our study indicates that (a) albuminuna furnishes thegreater contribution to the increase in exercise-induced pro-teinuna; (b) the exercise proteinuna is both glomerular andtubular in origin, and is reversible; (c) the enhanced proteinrequirementof athletes may in part be due to the recurrentexcretion of proteins in the urine after physical effort.Additional Keyphrases: albumin excretion rate diabetes melli-tus a -microglobulin diabetesIn the last few years, measurement of urinary albuminexcretion has become widely used for early assessment ofrenal glomerular involvement in different pathologicalconditions such as arterial hypertension (1-3) or post-transplant kidney rejection (4). In diabetics the evaluationof the urinary albumin excretion rate (AER) has recentlyassumed a central role in prevention and follow-up of dia-betic nephropathy (5). The screening and follow-up of pro-teinuria and (or) albuminuna in diabetic patients demandsa complete knowledge of the pathophysiological mechanismsthat can affect the excretion of urinary proteins (functionalproteinuria) in healthy and diabetic people.Physical effort causes proteinuria in healthy adults (6,7),children, and adolescents (8), as well as in football, basket-ball, and handball players (9) and long-distance runners(10). Physical exercise as a provocative test for proteinuri.ahas been proposed to unmask early changes of diabeticrenal disease (11). So far, the causes of increased excretionof urinary proteins during physical exercise in healthysubjects and diabetic patients are still unknown. In partic-ular, reports have been conflicting on the type of proteinuriainvolved-i.e., glomerular, tubular, or mixed in origin (12).We measured the urinary excretion of albumin, a1-C.N.R.Institute of Clinical Physiology,Cattedra di Malattie delRicambio, Istituto di Clinics Medics II, University of Pisa, ViaSavi 8, 56100 Pisa, Italy.Received July 14, 1989; acceptedDecember 11, 1989.microglobulin (as an indicator of the renal tubular dysfunc-tion) (12), in the basal state (overnight collection) and afterphysical exercise (training session) in 10 professional cy-clists. Urinary excretions of sodium, potassium, and creat-mine were also measured, substances well known to besubject to glomerular filtration and tubular rearrangement,although in differentdegrees and manners. The aim of ourstudy was therefore to verify whether protein excretion isincreased after exercise, even in the case of well-trainedathletes, and to elucidate whether the exercise-inducedproteinuria is glomerular or tubular (or both) in origin.Materials and MethodsUrinary albumin excretion. We measured urinary albu-min excretion in urine samples with a commercial RIA(Albumin RIA 100; Pharmacia AB, Uppsala, Sweden). ThisRIA method makes use of purified human serum albuminas standard, ‘251-labeled human serum albumin as tracer,and a specific antiserum to human serum albumin. Boundalbumin is separated from free by use of a solid-phasesystem (anti-rabbit IgG, raised in sheep, bound to Sepha-rose). The standard curve is prepared with use of solutionscontaining known amounts of purified human serum albu-min (Figure 1). We performed the assay according to themanufacturer’s instructions. The analytical characteristicsof this RIA method have been reported (13). In brief, weadded to all tubes 50 j.L of the standard solutions or urinesamples, 50 L of the tracer solution (about 30000-40 000counts/mm of ‘251-labeled human serum albumin), and 2.0mL of a mixture of antiserum to human serum albumin;mixed; then incubated the tubes for 1 h at room tempera-ture (18-22 #{176}C).After centrifugation (1500 x g, 10 mm,4#{176}C),the supernates were aspirated and discarded, and the6/Do, %1005060402000 20 40 60 80ALBUMIN ADDED, mg/LFig. 1. Standard curve and precision profile of the RIA method forassay of urinary albuminPrecisionprofile (within-runCV) calculated fromthe errorof replicates(14)#{149}overnightexercise* p<0.01*AER No K Cr UVjig/mmmmol/h mmol/h mg/h mvhCLINICALCHEMISTRY, Vol. 36, No. 3, 1990 563radioactivity in the pellets was counted in a well-typegamma counter. The RIA was not significantly subject tointerference from bovine serum albumin, transferrin, orhuman immunoglobulins.All sample values were obtained and the index of sensi-tivity, the precision profile, and the quality control for theRIA were evaluated according to a previously describedcomputer program (14). The interpolation of the dose-response curve was calculated with use of a four-parameterlogistic function (14).All urine samples from the 10 athletes were assayed inthe same RIA experiment. The standard curve and theimprecision profile (within-run variability) of this RIAexperiment are shown in Figure 1. The sensitivity (lowerdetection limit) of this standard curve was 0.38 mg/L. Thereproducibility (between-run variability) of the RIA wasdetermined by measuring three pooled specimens of urinein several consecutive assays. The CVs were 8.19% (albu-min concentration 4.74 ± 0.39 mg/L, mean ± SD, n = 18),7.38% (18.63 ± 1.37 mgfL, n = 26), and 10.53% (43.49 ±4.58 mg/L, n = 17), respectively.Other methods. The a1-microglobulin was measured by aradial immunodiffusion method (LC-Partigen a1-micro-globulin; Behringwerke AG, Marburg, F.R.G.). The detec-tion limit for this assay was 0.9 mg/L. Creatinine, sodium,and potassium in urine were measured by automatedmethods (Astra 4; Beckman Instruments, Brea, CA 92621).Subjects. We studied 10 male professional cyclists (ages21-26 years) during an intense training period. Eachsubject voided the bladder before going to bed and uponrising, collecting the latter. Collection times were exactlyrecorded (timed overnight specimen). In addition, a timedspecimen of urine was also collected from all athletesduring a training session lasting 5 h (road race of 153 kin,energy expenditure ranging from 60% to 90% of maximum02 expenditure during training). Every subject voidedurine just before the start of training session (this specimenwas discarded) and soon after (within 15 mm) the end of thetraining (exercise specimen). For every subject, after thevolumes of the overnight and exercise specimens had beenaccurately measured, the urines were divided into variousaliquots and stored at -20 #{176}Cuntil assay. Bacterial growthwas routinely excluded from all urines. For each urine fromeach subject we calculated the excretion rate for albumin(AER, g/min), creatinine (mg/h), and the electrolytes Naand K (mmol/h). The excretion rate for a1-microglobulinwas not calculated, because this protein was not detectablein half of the urines collected.We also studied samples of overnight urine specimensfrom 91 healthy adults (15) and from 22 healthy subjectswho had collected their urine for 24 h in several portionswhile maintaining their normal activity. None of thesesubjects was participating in physical training programs.The first urine specimen on the morning of the day of the24-h collection (specimen 0) was put in a separate con-tainer. Subsequently, the complete 24-h urinary collectionconsisted of specimen 1, from waking to lunch (generally 5h, from 0800 to 1300 hours); specimen 2, from lunch todinner (generally 6 h, from 1300 to 1900 hours); specimen3, from dinner to bedtime; and specimen 4, overnight, atrest. The volume of each portion was recorded, as was thetime of collection.Statistical analysis. The statistical analysis for the cal-culation of mean values, Student’s t-test for paired andunpaired data, Mann-Whitney U-test, simple and stepwiseregression analysis, and interpolation of the standard curveof the RIA was carried out by a Macintosh SE personalcomputer with use of the “Stat-View 512” program.ResultsFigure 2 shows the mean (± SEM) urinary excretionrates for albumin, Na, K, and creatinine, and the meanurine volumes, calculated both for the overnight and theexercise specimens from the 10 athletes. Compared withthe basal state (overnight collection), physical effort didprovoke a significant (P <0.01, t-test for paired data)increase in the excretion rate of albumin, Na, and K, and itincreased the urinary volume, whereas urinary creatinineoutput was not significantly affected by exercise. The exer-cise-induced increase in Na and K and in the urinaryvolume was 1.5-2.5 times less than that of albumin.The mean (± SD) AER of 10 athletes, measured in theovernight samples, was 4.2 ± 2.6 pg/min, a value not significantly different (P >0.05) from that found at rest for the 91healthy nonathletic subjects (4.6 ± 2.7 ig/min) (15). For the22 healthy subjects who collected a 24-h urine specimenduring a day with “normal” activity, the mean (± SD) AERwas 7.4 ± 3.9 p.g/min (range 1.8-17.0 g/min), significantlygreater than the AER found at rest for the overnight urinesample (sample 4, 5.54 ± 2.64 gImin, P <0.01).The mean (± SD) AER for the 10 athletes during exercise(18.1± 10.6 g/min) was significantly greater (P <0.01,both by t-test for unpaired data and by the Mann-WhitneyU-test) than those measured in the 22 control subjects inthe “morning” (specimen 1) and “afternoon” (specimen 2)periods (8.7 ± 5.9 and 8.2 ± 5.5 g/min, respectively) andduring the entire 24-h collection (7.4 ± 3.9 pg/min).Table 1 lists the correlation coefficients (r) between thevariables studied, calculated for all the urine specimensfrom the athletes (overnight + exercise collections, n = 20).Because the correlation coefficients for simple linear re-gressions were all highly significant (P 0.01), we per-formed stepwise multiple regression analysis to assess theseparate contribution of each variable to the regression.Only urinary volume significantly contributed to explain-ing the variability of AER in the stepwise multiple regres-605040200(meen±SEII, n 10)Fig. 2. Values for albumin (AER), Na, K, and creatinine (Cr) excretionrate and urine volume (UV) measured fortheovernightand theexercise specimens564 CLINICAL CHEMISTRY, Vol. 36, No. 3, 1990Table 1. Correlation Coetficien ts between the Variables‘aNa KCreatinine, Urine vol,mmol/h mg/h mL/hAER 0.70 0.762 0.528 0.786Na 0.779 0.5940.5520.920.834Creatinine 0.801a All rvalues were highlysignificant(P O.01).sion analysis, whereas Na, K, and creatinine excretionrates did not.The urinary concentration of a1-microglobulin was un-detectable in the overnight urine specimens from nine ofthe athletes and ranged from 0.9 to 25.7 mg/L (mean ±SEM 6.6 ± 2.5 mg/L) during exercise; only in a singleathlete was a1-microglobulin detectable in the overnightspecimen (2.8 mg/L) and undetectable in the exercise sam-ple. The mean value of a1-microglobulin in the exercisespecimen was significantly greater than that in the over-night specimen (6.6 vs 0.3 mg/L, P = 0.037, n = 10, t-testfor paired data).DiscussionOvert diabetic nephropathy, defined as proteinuria >500mg/24 h, is preceded by a silent phase characterized by apersistent increase in AER without clinical proteinuria (5).According to several recent prospective studies, this so-called microalbununuria strongly predicts the future devel-opment of overt diabetic nephropathy (5). Measurement ofAER is also of interest in the management of hypertensivepatients (1-3, 7). In both situations, the screening and (or)follow-up programs involving large groups of patients re-quire accurate standardization of the procedures for AERevaluation.In our study, the mean exercise-induced AER in well-trained athletes was 18 .tg/min, a value that clearly ex-ceedsour upper normal limit (11.5 j.ig/min) (15). Kramer etal. (7) recently reported that the 24-h albumin excretionfrom 30 healthy subjects was not significantly affected by amaximal bicycle ergometer test. However, the mean exer-cise AER value they reported (10.3 ± 0.9 mg/24 h) tended tobe greater than the AER measured for the urines collectedwithout exercise (8.5 ± 0.7 mg/24 h) (7). Use of a longerperiod of exercise might have enhanced the albumin excre-tion in that study. Our well-trained athletes excreted onthe average about 5.4mg of urinary albumin during 5 h oftraining. This corresponds to 50.5% of the mean whole 24-hamount of albumin excreted at rest by control subjects (10.7mg). Diabetic patients excrete more albumin in urine afterexercise than do healthy untrained subjects (16,17). Hence,the physical activity may significantly affect the determi-nation of AER in screening or follow-up programs of dia-betic and (or) hypertensive patients.Although in this study albuminuria furnished thegreater contribution to the exercise-induced proteinuria,which fits with reports of others (9, 18), nine of 10 athletesalso showed an increased excretion of a1-microglobulinduring exercise. Conceivably, physical activity also pro-vokes a tubular proteinuria. Thus, exercise proteinuriamay be tubular in origin, in addition to glomerular, asindicated by significant correlations between AER andurinary excretions of Na, K, and creatimne (Table 1).Our findings indicate that exercise proteinuria is rapidlyreversible, because these athletes, whom we studied at theend of an intense one-month period of training, showed nogreater AER values at rest (i.e., in the overnight collection)than did the normal controls.Finally, our data suggest that the increased proteinrequirements of athletes may be at least partly attributedto the repeated loss of proteins in the urine (19).This work was performed in the context of the Ricerca Finaliz-zata della Regione Toscana. In addition, it was supported in partby a grant (87.00381.56) from the National Research Council andby a grant from the Ministero della Pubblica Istruzione (RicercaScientifica 1987).References1. Parving RH, Jensen HA, Mogensen CE, Evrin PE. Increasedurinary albumin excretion rate in benign essential hypertension.Lancet 1974;i:1190-2.2. Pedersen EB, Mogensen CE. Effect of antihypertensive onurinary albumin excretion, glomerular filtration rate, and renalplasma flow in patients with essential hypertension. Scand J ClinLab Invest 1976;36:231-7.3. Losito A, Zarnpi L, Fortunati L, Del Favero A. Glomerularhyperifitration and albuminuria in essential hypertension. Neph-i-on 1988;49:84-5.4. Woo J, Floyd M, Cannon DC, Kaman B. Radioimmunoasaay forurinary albumin. Clin Chem 1978;24:1464-7.5. Viberti GC, Walker JD. Diabetic nephropathy: etiology andprevention [Review]. DiabetesfMetabolism Rev 1988;4:147-62.6. Fletcher DJ. Athletic pseudonephritis [Letter]. Lancet1977;i:910-1.7. Kramer KB, Kernz M, Ress KM, et al. Influence of strenuousexercise on albumin excretion. Clin Chem 1988;34:2516-8.8. Huttunen NP, Kaar ML, Pietilainen M, Vierikko P, Reimla M.Exercise-induced proteinuria in children and adolescents. ScandJClin Lab Invest 1981;41:583-7.9. Campanacci L, Faccini L, Englaro E, et al. Exercise-inducedproteinuria. Contrib Nephrol 1981;26:31-41.10. Mogensen CE, Vittinghus E. Urinary albumin excretion duringexercise in juvenile diabetes. Scand J Clin Invest 1975;35:295-300.11. Gilli P, De Paoli Vitali E, Tataranni G, Farinelli A. Exercise-induced urinary abnormalities in long-distance runners. mt JSport Med 1984;5:237-40.12. Wailer Ky, Ward KM, Mahan LD, Wilsmatt DK. Currentconcepts in proteinuria [Review]. Clin Chem 1989;35:755-65.13. Giampietro 0, Clerico A, Cruschelli L, et al. Measurement ofurinary albumin excretion rate (AER) in normal and diabeticsubjects: comparison of two recent radioimmunoasaays. J NucIMed Allied Sci 1987;31:321-6.14. Pilo A, Zucchelli GC, Malvano R, et al. Main features ofcomputer algorithms for RIA data reduction. Comparison of somedifferent approaches for the interpolation of dose-response curve. JNucI Med Allied Sci 1982;26:235-48.15. Giampietro 0, Miccoli H, Clerico A, et al. Urinary albuminexcretion in normal subjects and in diabetic patients measured bya radioimmunoasaay: methodological and clinical aspects. ClinBiochem 1988;21:63-8.16. Viberti GC, Jarrett RJ, McCartney M, Keen H. Increasedglomerular permeability to albumin induced by exercise in dia-betic subjects. Diabetologia 1978;14:293-300.17. Feldt-Rasmussen B, Baker L, Deckert T. Exercise as a provoc-ative test in early renal disease in Type 1 (insulin-dependent)diabetes: albuminuric, systemic and renal haemodynamic re-sponses. Diabetologia 1985;28:389-96.18. Poortmans J, Jeanloz RW. Quantitative immunological deter-mination of 12 plasma proteins excreted in human urine collectedbefore and after exercise. J Clin Invest 1968;47:386-93.19. Dohm LG. Protein nutrition for the athlete. Clin Sports Med1984;3:595-604.

Exercise-induced proteinuria in well-trained athletes

Exercise-induced proteinuria in well-trained athletes

Abstract

Similar works

Full text

Available Versions

Archivio della ricerca della Scuola Superiore Sant'Anna