Clinical Study Am J Nephrol 19 9 2 :1 2 :6 1 -6 7
Department of Internal Medicine. Divisions of Nephrology and Nutrition and Dietetics, University of Freiburg. FRG
Effect of Amino Acid Based Dialysis Solution on Peritoneal Permeability and Prostanoid Generation in Patients Undergoing Continuous Ambulatory Peritoneal Dialysis
K e y W o rd s
A b stract
Continuous ambulatory peritoneal dialysis Osmotic agents Amino acid solution Protein loss Prostanoids
The acute effect of amino acid based dialysis solution on peritoneal kinetics of amino acids and plasma proteins in comparison to conventional glucosebased dialvsate was studied in 9 patients with end-stage renal failure on con tinuous ambulatory peritoneal dialysis. Instillation of 2.6% amino acid solu tion resulted in raised plasma concentrations of all essential amino acids included in the dialysis fluid (p < 0.005). The amino acid solution induced an augmented leakage of plasma proteins into the dialysate at all dwell times investigated (1-8 h). After a dwell time at 8 h. the dialysate total protein increased from 2.62 ± 0.45 g with glucose dialysate to 3.85 ± 0.42 g with amino acid solution (p < 0.05). Corresponding results were obtained for pymicroglobulin. albumin, transferrin. IgG, and for the non-essential amino acids alanine, citrulline, and glutamine (p < 0.025) not included in the initial amino acid composition of the dialysis fluid. During the use o f amino acid based dialysis fluid, the effluent prostaglandin Ey concentration increased by more than 80% in comparison to glucose dialysate (p < 0.025). The aug mented loss of proteins induced by the amino acid solution was positively correlated with increased dialysate prostaglandin Ej (r = 0.8894; p < 0.001). Peritoneal ultrafiltration was not affected by the use of amino acid based dial ysate fluid. The present results indicate that amino acid based dialysis fluid enhances the peritoneal permeability for plasma proteins and amino acids, probably mediated by locally generated prostanoids.
Introduction
Malnutrition is a common complication in patients on chronic renal replacement therapy [1-4], Among patients undergoing continuous ambulatory peritoneal dialysis
Received: November 9, 1990 Accepted: March I. 1992
(CAPD). caloric malnutrition is usually not a problem because o f continuous glucose absorption from the dialy sis solution [5-7], By contrast, several factors contribute to impaired protein metabolism in CAPD. By itself urae mia may promote protein degradation by decreased de
Dr. H.B. Steinhauer Medizinische Universitätsklinik Abteilung Nephrologie Hugstctter Strasse 55 D-W-78ÖO Freiburg (FRG)
0 1992 S. Karger AG, Basel 0250-8095/92/012 2 -0 0 6 1 $2.75/0
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Hjalmar B. Steinhauer a Iris Lubrich-Birkner a Reinliold Klutheb Gieselci Baw nannb Peter Schollmeyer a
P atients and M eth o d s Patients Nine CAPD patients. 2 females and 7 males, aged 2 4 -7 3 years (mean 51.2 years) were enrolled in the study. Informed consent was obtained from all patients prior to the study. The causes of renal failure were chronic glomerulonephritis in 4 cases, diabetic nephrop athy in 3, chronic interstitial nephritis in 1. and Alport's syndrome in 1 case. All patients were stable on CAPD and free from peritonitis for at least 4 weeks. The mean time on CAPD treatment was 15 months (range 1 -72 months). During the study the patients were hospitalized and maintained on their usual protein-rich diet of 1.2-1.5 g/kg body weight/day.
Melhods The protocol was designed as a sequential cross-over study. Dur ing the study ‘period A" the patients were randomly allocated for treatment with a 4 .2 5 % glucose-based dialysis solution or a 2.6% amino acid containing dialysate (both solutions obtained from Fresenius. Bad Homburg. FR G : see table 1). The pH value o f both dialysis fluids was 5.6. and the osmolality was 525 mosm/kg in the glucose and 518 mosm/kg in the amino acid solution. Both solutions were used for one morning exchange daily on 6 consecutive days with a dwell time o f 1, 2, 3. 4, 6. and 8 h and a volume o f 1.5 litres. In study ‘period B' the patients were switched to
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the alternative dialysis fluid not used during ‘period A'. The further protocol remained unchanged. Between the study dialysate exchanges CAPD was performed according to the clinical requirements to maintain a constant state of hydration. Fasting heparinized blood samples were collected before instilla tion of dialysis solution and shortly after dialysate efflux. Samples of dialysate and blood were cooled to 4 ° C after collection and then centrifuged at 3.000,? for 15 min to remove cellular components. The supernatants were stored at - 3 0 °C for further analysis. Diaivsate glucose was measured within 1 h after sampling by standard lab oratory methods. Plasma and dialysis effluent for amino acid analy sis were deproteinized by sulphosalicylic acid (10% w/v) before freezing. The sulphosalicylic acid extracts were stored at - 7 0 ° C . Amino acids were analyzed by means o f a conventional single-col umn ion-cxchangc method with a lithium buffer system [25]. Dialysate total protein was measured by tannic acid precipita tion and F e u colorimetry [26]. (T-Microglobulin was determined by an enzyme-linked immunoassay (Elias. Freiburg. FRG ). Albumin, transferrin, and IgG were measured by the radial immunodiffusion technique (Behringwerke, Marburg. FRG). Plasma and dialysate cre atinine, urea, potassium, and phosphate were determined by stan dard laboratory methods. The dialysate arachidonic acid metabolites prostaglandin (PG) Ei. 6-keto-PGFi«. and thromboxane B? were determined by specific radioimmunoassays as described by Dray et al. [27] and recently by our group [2 8 .2 9 ]. Prior to the prostanoid analysis, dialysate samples were purified by solid-phase sorbent extraction (Amprcp C2 minico lumns: Amersham Buchler, Braunschweig, FRG ). Interfering com pounds were eluted by water. 10% ethanol, and hexane. Analytic elu tion was performed using methyl formeate. After evaporation the eluates were dissolved in phosphate-buff ered saline ( 0 .1 mol/1, pH 7,4, containing gelatine 1 mg/ml) and ana lyzed by radioimmunoassay. The recovery of the extraction procedure was determined by addition o f known amounts of prostanoids to charcoal-absorbed dialysate [0.2 ml charcoal suspension (20 ng/ml phosphate-buffered saline) added to I ml dialysate. 30 min incubation period at room temperature, separated by centrifugation]. The recovery was 77.6 ± (SD) 6.2% for thromboxane B;. 78.6 ± 8.2 for P G E;. and 82.1 ± 7.5% for 6-kcto-PG Fiu (n = 6). Parallel results were obtained between serial dilutions of dialysate samples with unknown prostanoid content and standard curves in phosphate-buffered saline. The coefficients of intra- and interassay variation were below 6.3 and 8.2%, respectively. Ultrafiltration was calculated by the dialysate efflux volume minus the dialysis solution influx volume divided by the dwell time. The results are expressed as mean values ± SEM. The data were evaluated by paired l test and linear regression analyses. The null hypothesis was rejected at p > 0.05.
R esults
The 2.6% amino acid based dialysate was tolerated without severe side effects. 4 out of 9 patients reported slight abdominal pain during the infusion. Within 30 min the discomfort disappeared in all patients. The 4.25%
Steinhauer/Lubrich-Birkner/Kluthc/ Baumann/Schollmevcr
Effect of Amino Acid Based CAPD Fluid on Peritoneal Permeabilitv
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novo protein synthesis and increased protein catabolism [1. 8]. In addition. CAPD treatment leads to both an increased loss of amino acids and proteins into the dialvsatc [9-12] and decreased appetite, probably due to peri toneal glucose absorption and abdominal distension by the dialysis solution. Even though most patients maintain normal serum total protein levels and albumin homeosta sis during the initial months on CAPD [13-15], long-term CAPD treatment results in decreased total body nitrogen and a reduced protein intake < 1 g/kg body weight/day [13. 16]. Since nitrogen balance studies in CAPD patients rec ommend a dietary protein intake > 1.2 g/kg body weight/ day in order to maintain nitrogen equilibrium [17-19], the replacement of glucose dialysate by an amino acid based solution should improve the nutritional status by providing additional nitrogen to replace losses of proteins and amino acids as well as to compensate for diminished protein intake. Recent clinical studies have demonstrated that significant amounts of amino acids are absorbed fol lowing intraperitoneal administration [20-24]. In the fol lowing study we investigated the acute effects of an amino acid based dialysis solution in comparison to conven tional glucose dialysate on the peritoneal kinetics of amino acids and plasma proteins as well as on the local generation of prostanoids affecting the peritoneal perme ability.
standard glucose solution was well tolerated by all pa tients. The dialysate leucocyte count was not affected by the use of amino acid solution (glucose solution 57 ± 8/ mm3: amino acid solution 49 ± 10/mm3; dwell time 1 h). The plasma amino acid concentration expressed as percentage of fasting concentration after instillation of amino acid based dialysis fluid or glucose solution is shown in figure 1. The use of amino acid solution resulted in significantly increased plasma concentrations of the eight essential amino acids included in the dialysis fluid (table 2). The maximum plasma levels of histidine, isoleu cine, leucine, lysine, methionine, threonine, and valine were ascertained after a dialysis time of I h. The peak plasma concentration o f phenylalanine occurred 2 h after instillation. After 8 h of dialysis the plasma levels of histidine and lysine returned to the basal range. The concentration of the further amino acids included in the dialysis fluid exceeded the fasting plasma concentrations by 60-118% after a dwell time o f 8 h (table 2). The glucose-based dialy sis solution w'as without significant effect on plasma amino acids (fig. 1). The amino acid based dialysis solution induced an augmented peritoneal leakage of plasma proteins and amino acids. The dialysate concentration of 14 non-essen tial amino acids not initially present in the dialysis fluid is shown in figure 2. Alanine, citrulline, and glutamine con centrations in the dialysate increased by more than 40% (p < 0.025) as compared with glucose exchanges, (dwell time 8 h): alanine in glucose solution 242 ± 18 pmol/I. in amino acid solution 351 ± 24: citrulline in glucose solu tion 52 ± 5. in amino acid solution 72 ± 7: glutamine in
Fig. 1. Total plasma concentration of eight essential amino acids included in the amino acid based CAPD fluid during amino acid exchanges (•) and glucose exchanges (o). Amino acid concentration expressed as percentage of basal concentration. Mean values ± SEM are shown ( n = 9).
Table 1. Composition of the amino acid based dialysis solution (mmol/1; all amino acids used were Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Valine
11.75 32.39 54.04 19.57 13.90 11.04 15.03 30.20
L forms) 143.00 Sodium Calcium 1.75 Magnesium 0.50 103.50 Chloride 35.00 Lactate Dextrose monohydrate 27.75
Table 2. Effect o f amino acid based dialysis solution on plasma amino acid concentrations (pmol/1: means ± SEM. n = 9) A m in o a cid s
B as al va lu e1
D w e ll tim e, h 1
2
3
8
6
4
H istid ine
84 + 7
1 5 2 ± 12 b
116+ 1 la
106+10
9 5 ± 10
99 ± 9
Isoleucine
75±4
413 + 26'
338 + 32e
287±21'
226 ± 2 5 '
1 47 ± 9 '
121 ± 1 1 b
L eucin e
105 + 7
638 ± 40e
538 ± 45e
465 ± 3 2 '
386 + 40'
247 ± 1 8 '
205 + 2 2b
L ysin e
183 ± 1 3
397 + 22e
317 ± 21c
283 ± 2 3 b
252±29a
2 0 2 ± 17
2 0 0 ± 17
M eth io n in e
38 ± 2
1 7 9 ± 15 e
163+19'
1 4 9 ± 17 C
129±24b
P henylalan in e
74 + 5
1 7 5 ± 18 h
189 ± 3 0 b
17 3 ± 2 2 b
159 ± 2 3 b
139+21°
1 2 5 ± 12 b
9 5 ± 19 a
98±8
7 8 ± 12a
T h reon in e
1 4 2 ± 15
175 ± 3 1 b
255±25b
1 64 ± 2 7 b
220 ± 2 5 3
229±38a
231 ± 4 2 a
V aline
1 7 4 ± 12
6 1 7 ± 3 5C
611 ± 39c
608±32b
512±45'
433 ± 3 2 '
379±36b
B e f o r e t h e first in flu x o f a m i n o a c i d b a s e d d i a l y s i s s o l u t i o n . ap < 0 . 0 5 : h p < 0 . 0 0 5 : c p < 0 . 0 0 0 5 as c o m p a r e d w ith the basal valu es.
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1
Fig. 2. Total concentration of 14 non-essential amino acids (ala nine. arginine, asparagine, aspartic acid, a-aminobutyric acid, citrulline. cysteine, glutamine, gluatmic acid, glycine, ornithine, serine, taurine, tyrosine) in dialysate at different dwell times during amino acid exchanges (♦ ) and during glucose exchanges (0 ). The values represent mean ± SEM (n = 9). Fig. 3. Effect of amino acid based CAPD fluid (closed symbols) versus glucose based dialysis solution (open symbols) on the perito neal loss of total protein (♦ / 6 h). peritoneal ultrafiltration and clearance rates of small solutes did not differ significantly between amino acid solution and glucose-based dialysis fluid. These data are in accordance with the results of several previous studies [20.21,23]. Furthermore, amino acid based dialy sate was found to enhance the peritoneal loss of plasma proteins into the dialysis fluid to a clinically relevant extent. The CAPD-associated loss o f protein increased from 2.62 ± 0.45 g/8 h using conventional 4.25% glucose solution to 3.85 ± 0.42 g/8 h with amino acid based dial ysate. The augmented peritoneal protein loss was ob served for all individual proteins investigated, indepen dent of their molecular weight (fig. 3). The dialysate PGE2 concentration during the use o f the amino acid solution was found to be positively correlated with the enhanced permeation of total plasma protein into the dialysate
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(fig. 4). Significant positive correlations were also estab lished between dialysate PGE2 and various individual proteins. Since PGE2 is known to possess vasodilative and permeability-increasing properties [36, 37], it is rea sonable to assume that the observed enhanced peritoneal permeability for plasma proteins during the use of amino acid based dialysate is mediated by locally generated p g e 2. The observed stimulation of peritoneal prostanoid gen eration rises further questions. The amino acid induced release of PGE2 may be a sign of local non-bacterial inflam mation. The initial abdominal pain in some of our patients after instillation o f the amino acid solution supports this assumption. Similar clinical symptoms during the use of amino acid based CAPD fluid were observed by Hanning ct al. [23]. Since repeated peritoneal inflammation is sup posed to contribute to impaired ultrafiltration and ana tomic changes of the peritoneum [38-40], amino acid based dialysis solutions should be used with caution. Recently prostaglandins of the E series were established to induce specific morphological changes in connective tissue [41. 42], PGE2 is supposed to promote the extrusion of 'matrix lysosomes' from myofibroblasts into the extracel lular space. The consecutive release of lysosomal enzymes such as proteases and phospholipases leads to the degrada tion of fibrinogen and fibrin to split products which are known to stimulate myofibroblast proliferation [41], By this way locally released PGE2 may contribute to the development of peritoneal fibrosis. In our opinion, the use of an amino acid containing dialysis solution is at present no convincing way to improve the nutrition status of CAPD patients. The observed change in peritoneal permeability with clinically relevant increased losses of proteins and non-essential amino acids is probably caused by amino acid induced locally generated prostanoids. Further long-term studies are necessary to elucidate the effect of continuous stimulation of peritoneal PG syn thesis and o f increased peritoneal leakage for macromole cules by dialysate amino acids on the clinical outcome of CAPD. A ck n o w le d g m e n ts This work was supported in part by a grant from the Fresenius Foundation, FRG . Portions of this work were presented at the 8th National Conference on CAPD. Kansas City, Mo.. USA, 1988. and at the 26th Congress of EDTA. Göteborg. Sweden. 1989. The authors wish to thank Mrs. K. Bappert and Mrs. G. Meier for their excellent technical assistance and Mrs. M. Baur for her careful secretarial assistance.
Stcinhauer/Lubrich-Birkner/Kluthe/ Ba um a nn/Sch oil m ever
Effect of Amino Acid Based CAPD Fluid on Peritoneal Permeability
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D iscussio n
R eferences 15 Lindholm B. Bergstrom J: Nutritional aspects on CAPD: in Gokal R (ed): Continuous Ambu latory Peritoneal Dialysis. London, Churchill Livingstone. 1986. p 228. 16 Schilling H. Wu G. Pettit J. Harrison J. McNeil K, Siccion Z. Oreopoulos DG: Nutritional sta tus of patients on long-term CAPD. Périt Dial Bull 1985:5:12-18. 17 Giordano C. De Santo NG. Pluvio M. Di Leo VA, Capodicasa G, Carillo D. Esposito R, Damiano M: Protein requirement of patients on CAPD: A study on nitrogen balance. Int J ArtifOrgans 1980:3:11-16. 18 Gahl GM. Baeyer HV. Avcrdunk R: Outpa tient evaluation o f dietary intake and nitrogen removal in continuous ambulatory' peritoneal dialysis. Ann Intern Med 1981:94:643-646. 19 Blumenkrantz M J. Kopplc JD . Moran JK . Co burn JW : Metabolic balance studies and di etary protein requirements in patients un dergoing continuous ambulatory peritoneal di alysis. Kidney Int 1982;21:849-861. 20 Williams PF. Marliss EB. Anderson GH. Oren A, Stein AN. Khanna R. Pettit J. Brandes L. Rodclla H. Mupas L. Dombros N. Oreopoulos DG: Amino acid absorption following intra peritoneal administration in CAPD patients. Peril Dial Bull 1982;2:124-130. 21 Oren A. Wu G. Anderson GH. Marliss E, Khanna R. Pettit J. Mupas L, Rodclla H. Roncari DA. Kakis G. Harrison J. McNeil K. Oreo poulos DG: Effective use o f amino acid dialy sate over four weeks in CAPD patients. Périt Dial Bull 1983;3:66-73. 22 Pedersen FB. Dragsholt C. I.aier E. Frifelt JJ. Trostmann AF. Ekelund S. Paaby P: Alternate use of amino acid and glucose solution in CAPD. Périt Dial Bull 1985;5:215-218. 23 Hanning RM. Balfe JW , Zlotkin SH: Effective ness and nutritional consequences on amino acid-based vs. glucose-based dialysis solutions in infants and children receiving CAPD. Am J Clin Nutr 1987:46:22-30. 24 Young GA. Dibble JB . Hobson SM. Tompkins L. Gibson J. Turney JH . Brownjohn AM: The use of an amino-acid-based CAPD fluid over 12 weeks. Nephrol Dial Transplant 1989:4: 285-292. 25 Jontofsohn R. Trivisas G. Katz N. Kluthe R: Amino acid content of erythrocytes in uremia. Am J Clin Nutr 19 7 8 :3 1:1956-1 960. 26 Yatzidis H: New colorimetric method for quantitative determination of protein in urine. ClinChem 1977:23:811-812. 27 Dray F. Charbonnel B. Maclouf J: Radioim munoassay of prostaglandins F. Ei and Ey in human plasma. Eur J Clin Invest 1975:5:311318. 28 Steinhaucr HB. Batsford S. Schollmeyer P. Kluthe R: Studies on thromboxane By and prostaglandin Ey in the course of murine au toimmune disease: Inhibition by oral histidine and zinc supplementation. Clin Nephrol 1985: 24:63-68.
29 Steirhauer HB. Wilms H. Rüther M. Schollmeyer P: Clinical experience with urine TXBy in acute renal allograft rejection. Transplant Proc 1986; 18(suppl ):9 8 -103. 30 Dombros N. Oren A. Marliss EB. Anderson GH. Stein AN, Khanna R. Pettit J. Brandes L. Rodclla H. Leibl SB. Oreopoulos DG: Plasma amino acid profiles and amino acid losses in patients undergoing CAPD. Peril Dial Bull 1982;2:27-32. 31 Katirtzoglou A. Oreopoulos DG. Husdan H. Leung M. Ogilvie R, Dombros N: Reappraisal of protein losses in patients undergoing contin uous ambulatory peritoneal dialysis. Nephron 1980:26:230-233. 32 Steinhauer HB. Schollmeyer P: Prostaglandinmediated loss of proteins during peritonitis in continuous ambulatory peritoneal dialysis. Kidney Int 1986:29:584-590. 33 Bruno M. Bagnis C. Marangella M. Rovcra L. Cantaluppi A. Linari F: CAPD with an amino acid dialyis solution: A long-term, cross-over study. Kidney Int 1989:35:1189-1194. 34 Dombros NV. Prutis K. Tong M, Anderson GH. Harrison J. Sombolos K. Digenis G. Pettit J. Oreopoulos DG: Six-month overnight intra peritoneal amino-acid infusion in continuous ambulatory peritoneal dialysis (CAPD) pa tients: No effect on nutritional status. Périt Dial Int 1990:10:79-84. 35 Dibble JB . Young GA, Hobson SM. Brown john AM: Amino-acid-based continuous am bulatory peritoneal dialysis (CAPD) fluid over twelve weeks: Effects on carbohydrate and lipid metabolism. Peril Dial Int 1990:10:7177. 36 Williams T J. Peck MJ: Role o f prostaglandinmediated vasodilation in inflammation. Na ture 1977:270:530-532. 37 Murota SI. Morita I: Effect of prostaglandin ly and related compounds on vascular permeabil ity responses in granuloma tissue. Prostaglan dins 1978:15:297-301. 38 Steinhaucr HB. Frei A. Drcyling KW. Scholl meyer P: Changes of eicosanoid metabolism in CAPD-associated peritonitis; in Augustin R (ed): Peritonitis in CAPD. Contrib Nephrol. Basel. Karger. 1987, vol 57. pp 45-54. 39 Oreopoulos DG: Peritoneal membrane: Han dle with care. Peril Dial Bull I983;3:l 11-113. 40 Backenroth-Maayan R. Longnecker R. Kambosos D: Failure o f the peritoneal membrane during chronic intermittent peritoneal dialysis; in Gahl GM. Kessel M. Nolph KD (eds): Ad vances in Peritoneal Dialysis. Amsterdam. Excerpta Medica. 19 8 1. pp 2 0 8 -2 13. 4 1 Riede UN: Pathogenesis of shock-induced pul monary fibrosis in man as a model of prolifera tive inflammation: in Deicher H. Schulz LC (eds): Arthritis. Models and Mechanisms. Ber lin, Springer, 19 8 1. pp 88-95. 42 Joh K. Riede UN. Zahradnick HP: The effect of prostaglandins on the lysosomal function in the cervix uteri. Arch Gynecol 1983:234:1-16.
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1 Kopple JD : Abnormal amino acid and protein metabolism in uremia. Kidney Int 1978:14: 340-348. 2 Bansal VK, Popli S. Pickering J, Ing TS. Vertuno LL. Hano JE : Protein-caloric malnutri tion and cutaneous anergy in hemodialysis maintained patients. Am J Clin Nutr 1980:33: 1608-1611. 3 Blumenkrantz MJ. Kopple JD . Gutman RA. Chan YK. Barbour GL. Roberts C. Shen FH. Gandhi VS, Tucker CT. Curtis FK, Coburn JW : Methods of assessing nutritional status of patients with renal failure. Am J Clin Nutr 1980:33:1567-1585. 4 Alvestrand A. Bergström J: Nutritional man agement; in Suki WN. Massry SG (eds): Ther apy of Renal Disease and Related Disorders. The Hague. Nijhoff. 1984. p 459. 5 De Santo NG. Capodicasa G. Senatore R. Cicchctti T. Cirillo D. Damiano M. Torella R. Giugliano D. Improta L. Giordano C: Glucose utilization from dialysate in patients on contin uous ambulatory peritoneal dialysis (CAPD). Int J Artif Organs 1979:2:119-124. 6 Grodstein GP. Blumenkrantz MJ. Kopple JD . Moran JK . Coburn JW : Glucose absorption during continuous ambulatory peritoneal dial ysis. Kidney Jnt 1981:19:564-567. 7 Blumenkrantz MJ. Schmidt RW: Managing the nutritional concerns of the patient undergoing peritoneal dialysis: in Nolph KD (ed): Perito neal Dialysis. The Hague. Nijhoff. 1981. pp 2 7 5 -3 0 8 .' 8 Bergstrom J: Protein catabolic factors in pa tients on renal replacement therapy. Blood Purif 1985:3:215-236. 9 Giordano C. De Santo NG, Capodicasa G. Di Leo VA. Di Serafino A. Carillo D. Esposito R. Fiore R. Damiano M. Buonadonna L, Cocco F. Di Iorio B: Amino acid losses during CAPD. Clin Nephrol 1980:14:230-232. 10 Rubin J. Nolph KD, Arfania D. Prowant B. Frulo L. Brown P. Moore H: Protein losses in continuous ambulatory peritoneal dialysis. Nephron 1981:28:218-221. 11 Blumenkrantz MJ. Gahl GM. Kopple JD . Kamdar AV. Jones MR. Kessel M. Coburn JW: Protein losses during peritoneal dialysis. Kid ney Int 1981:19:593-602. 12 Kopple JD . Blumenkrantz MJ. Jones MR. Mo ran JK . Coburn JW : Plasma antino acid levels and amino acid losses during continuous am bulatory peritoneal dialysis. Am J Clin Nutr 1982:36:395-402. 13 Heide B. Picrratos A. Khanna R. Pettit J . Ogil vie R. Harrison J, McNeil K. Siccion Z. Oreopoulos D: Nutritional status o f patients un dergoing continuous ambulatory peritoneal di alysis (CAPD). Peril Dial Bull 1983:3:138141. 14 Kaysen GA, Schoenfeld PY: Albumin homeo stasis in patients undergoing continuous ambu latory peritoneal dialysis. Kidney Int 1984:25: 107-114.
Department of Internal Medicine. Divisions of Nephrology and Nutrition and Dietetics, University of Freiburg. FRG
Effect of Amino Acid Based Dialysis Solution on Peritoneal Permeability and Prostanoid Generation in Patients Undergoing Continuous Ambulatory Peritoneal Dialysis
K e y W o rd s
A b stract
Continuous ambulatory peritoneal dialysis Osmotic agents Amino acid solution Protein loss Prostanoids
The acute effect of amino acid based dialysis solution on peritoneal kinetics of amino acids and plasma proteins in comparison to conventional glucosebased dialvsate was studied in 9 patients with end-stage renal failure on con tinuous ambulatory peritoneal dialysis. Instillation of 2.6% amino acid solu tion resulted in raised plasma concentrations of all essential amino acids included in the dialysis fluid (p < 0.005). The amino acid solution induced an augmented leakage of plasma proteins into the dialysate at all dwell times investigated (1-8 h). After a dwell time at 8 h. the dialysate total protein increased from 2.62 ± 0.45 g with glucose dialysate to 3.85 ± 0.42 g with amino acid solution (p < 0.05). Corresponding results were obtained for pymicroglobulin. albumin, transferrin. IgG, and for the non-essential amino acids alanine, citrulline, and glutamine (p < 0.025) not included in the initial amino acid composition of the dialysis fluid. During the use o f amino acid based dialysis fluid, the effluent prostaglandin Ey concentration increased by more than 80% in comparison to glucose dialysate (p < 0.025). The aug mented loss of proteins induced by the amino acid solution was positively correlated with increased dialysate prostaglandin Ej (r = 0.8894; p < 0.001). Peritoneal ultrafiltration was not affected by the use of amino acid based dial ysate fluid. The present results indicate that amino acid based dialysis fluid enhances the peritoneal permeability for plasma proteins and amino acids, probably mediated by locally generated prostanoids.
Introduction
Malnutrition is a common complication in patients on chronic renal replacement therapy [1-4], Among patients undergoing continuous ambulatory peritoneal dialysis
Received: November 9, 1990 Accepted: March I. 1992
(CAPD). caloric malnutrition is usually not a problem because o f continuous glucose absorption from the dialy sis solution [5-7], By contrast, several factors contribute to impaired protein metabolism in CAPD. By itself urae mia may promote protein degradation by decreased de
Dr. H.B. Steinhauer Medizinische Universitätsklinik Abteilung Nephrologie Hugstctter Strasse 55 D-W-78ÖO Freiburg (FRG)
0 1992 S. Karger AG, Basel 0250-8095/92/012 2 -0 0 6 1 $2.75/0
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Hjalmar B. Steinhauer a Iris Lubrich-Birkner a Reinliold Klutheb Gieselci Baw nannb Peter Schollmeyer a
P atients and M eth o d s Patients Nine CAPD patients. 2 females and 7 males, aged 2 4 -7 3 years (mean 51.2 years) were enrolled in the study. Informed consent was obtained from all patients prior to the study. The causes of renal failure were chronic glomerulonephritis in 4 cases, diabetic nephrop athy in 3, chronic interstitial nephritis in 1. and Alport's syndrome in 1 case. All patients were stable on CAPD and free from peritonitis for at least 4 weeks. The mean time on CAPD treatment was 15 months (range 1 -72 months). During the study the patients were hospitalized and maintained on their usual protein-rich diet of 1.2-1.5 g/kg body weight/day.
Melhods The protocol was designed as a sequential cross-over study. Dur ing the study ‘period A" the patients were randomly allocated for treatment with a 4 .2 5 % glucose-based dialysis solution or a 2.6% amino acid containing dialysate (both solutions obtained from Fresenius. Bad Homburg. FR G : see table 1). The pH value o f both dialysis fluids was 5.6. and the osmolality was 525 mosm/kg in the glucose and 518 mosm/kg in the amino acid solution. Both solutions were used for one morning exchange daily on 6 consecutive days with a dwell time o f 1, 2, 3. 4, 6. and 8 h and a volume o f 1.5 litres. In study ‘period B' the patients were switched to
62
the alternative dialysis fluid not used during ‘period A'. The further protocol remained unchanged. Between the study dialysate exchanges CAPD was performed according to the clinical requirements to maintain a constant state of hydration. Fasting heparinized blood samples were collected before instilla tion of dialysis solution and shortly after dialysate efflux. Samples of dialysate and blood were cooled to 4 ° C after collection and then centrifuged at 3.000,? for 15 min to remove cellular components. The supernatants were stored at - 3 0 °C for further analysis. Diaivsate glucose was measured within 1 h after sampling by standard lab oratory methods. Plasma and dialysis effluent for amino acid analy sis were deproteinized by sulphosalicylic acid (10% w/v) before freezing. The sulphosalicylic acid extracts were stored at - 7 0 ° C . Amino acids were analyzed by means o f a conventional single-col umn ion-cxchangc method with a lithium buffer system [25]. Dialysate total protein was measured by tannic acid precipita tion and F e u colorimetry [26]. (T-Microglobulin was determined by an enzyme-linked immunoassay (Elias. Freiburg. FRG ). Albumin, transferrin, and IgG were measured by the radial immunodiffusion technique (Behringwerke, Marburg. FRG). Plasma and dialysate cre atinine, urea, potassium, and phosphate were determined by stan dard laboratory methods. The dialysate arachidonic acid metabolites prostaglandin (PG) Ei. 6-keto-PGFi«. and thromboxane B? were determined by specific radioimmunoassays as described by Dray et al. [27] and recently by our group [2 8 .2 9 ]. Prior to the prostanoid analysis, dialysate samples were purified by solid-phase sorbent extraction (Amprcp C2 minico lumns: Amersham Buchler, Braunschweig, FRG ). Interfering com pounds were eluted by water. 10% ethanol, and hexane. Analytic elu tion was performed using methyl formeate. After evaporation the eluates were dissolved in phosphate-buff ered saline ( 0 .1 mol/1, pH 7,4, containing gelatine 1 mg/ml) and ana lyzed by radioimmunoassay. The recovery of the extraction procedure was determined by addition o f known amounts of prostanoids to charcoal-absorbed dialysate [0.2 ml charcoal suspension (20 ng/ml phosphate-buffered saline) added to I ml dialysate. 30 min incubation period at room temperature, separated by centrifugation]. The recovery was 77.6 ± (SD) 6.2% for thromboxane B;. 78.6 ± 8.2 for P G E;. and 82.1 ± 7.5% for 6-kcto-PG Fiu (n = 6). Parallel results were obtained between serial dilutions of dialysate samples with unknown prostanoid content and standard curves in phosphate-buffered saline. The coefficients of intra- and interassay variation were below 6.3 and 8.2%, respectively. Ultrafiltration was calculated by the dialysate efflux volume minus the dialysis solution influx volume divided by the dwell time. The results are expressed as mean values ± SEM. The data were evaluated by paired l test and linear regression analyses. The null hypothesis was rejected at p > 0.05.
R esults
The 2.6% amino acid based dialysate was tolerated without severe side effects. 4 out of 9 patients reported slight abdominal pain during the infusion. Within 30 min the discomfort disappeared in all patients. The 4.25%
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Effect of Amino Acid Based CAPD Fluid on Peritoneal Permeabilitv
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novo protein synthesis and increased protein catabolism [1. 8]. In addition. CAPD treatment leads to both an increased loss of amino acids and proteins into the dialvsatc [9-12] and decreased appetite, probably due to peri toneal glucose absorption and abdominal distension by the dialysis solution. Even though most patients maintain normal serum total protein levels and albumin homeosta sis during the initial months on CAPD [13-15], long-term CAPD treatment results in decreased total body nitrogen and a reduced protein intake < 1 g/kg body weight/day [13. 16]. Since nitrogen balance studies in CAPD patients rec ommend a dietary protein intake > 1.2 g/kg body weight/ day in order to maintain nitrogen equilibrium [17-19], the replacement of glucose dialysate by an amino acid based solution should improve the nutritional status by providing additional nitrogen to replace losses of proteins and amino acids as well as to compensate for diminished protein intake. Recent clinical studies have demonstrated that significant amounts of amino acids are absorbed fol lowing intraperitoneal administration [20-24]. In the fol lowing study we investigated the acute effects of an amino acid based dialysis solution in comparison to conven tional glucose dialysate on the peritoneal kinetics of amino acids and plasma proteins as well as on the local generation of prostanoids affecting the peritoneal perme ability.
standard glucose solution was well tolerated by all pa tients. The dialysate leucocyte count was not affected by the use of amino acid solution (glucose solution 57 ± 8/ mm3: amino acid solution 49 ± 10/mm3; dwell time 1 h). The plasma amino acid concentration expressed as percentage of fasting concentration after instillation of amino acid based dialysis fluid or glucose solution is shown in figure 1. The use of amino acid solution resulted in significantly increased plasma concentrations of the eight essential amino acids included in the dialysis fluid (table 2). The maximum plasma levels of histidine, isoleu cine, leucine, lysine, methionine, threonine, and valine were ascertained after a dialysis time of I h. The peak plasma concentration o f phenylalanine occurred 2 h after instillation. After 8 h of dialysis the plasma levels of histidine and lysine returned to the basal range. The concentration of the further amino acids included in the dialysis fluid exceeded the fasting plasma concentrations by 60-118% after a dwell time o f 8 h (table 2). The glucose-based dialy sis solution w'as without significant effect on plasma amino acids (fig. 1). The amino acid based dialysis solution induced an augmented peritoneal leakage of plasma proteins and amino acids. The dialysate concentration of 14 non-essen tial amino acids not initially present in the dialysis fluid is shown in figure 2. Alanine, citrulline, and glutamine con centrations in the dialysate increased by more than 40% (p < 0.025) as compared with glucose exchanges, (dwell time 8 h): alanine in glucose solution 242 ± 18 pmol/I. in amino acid solution 351 ± 24: citrulline in glucose solu tion 52 ± 5. in amino acid solution 72 ± 7: glutamine in
Fig. 1. Total plasma concentration of eight essential amino acids included in the amino acid based CAPD fluid during amino acid exchanges (•) and glucose exchanges (o). Amino acid concentration expressed as percentage of basal concentration. Mean values ± SEM are shown ( n = 9).
Table 1. Composition of the amino acid based dialysis solution (mmol/1; all amino acids used were Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Valine
11.75 32.39 54.04 19.57 13.90 11.04 15.03 30.20
L forms) 143.00 Sodium Calcium 1.75 Magnesium 0.50 103.50 Chloride 35.00 Lactate Dextrose monohydrate 27.75
Table 2. Effect o f amino acid based dialysis solution on plasma amino acid concentrations (pmol/1: means ± SEM. n = 9) A m in o a cid s
B as al va lu e1
D w e ll tim e, h 1
2
3
8
6
4
H istid ine
84 + 7
1 5 2 ± 12 b
116+ 1 la
106+10
9 5 ± 10
99 ± 9
Isoleucine
75±4
413 + 26'
338 + 32e
287±21'
226 ± 2 5 '
1 47 ± 9 '
121 ± 1 1 b
L eucin e
105 + 7
638 ± 40e
538 ± 45e
465 ± 3 2 '
386 + 40'
247 ± 1 8 '
205 + 2 2b
L ysin e
183 ± 1 3
397 + 22e
317 ± 21c
283 ± 2 3 b
252±29a
2 0 2 ± 17
2 0 0 ± 17
M eth io n in e
38 ± 2
1 7 9 ± 15 e
163+19'
1 4 9 ± 17 C
129±24b
P henylalan in e
74 + 5
1 7 5 ± 18 h
189 ± 3 0 b
17 3 ± 2 2 b
159 ± 2 3 b
139+21°
1 2 5 ± 12 b
9 5 ± 19 a
98±8
7 8 ± 12a
T h reon in e
1 4 2 ± 15
175 ± 3 1 b
255±25b
1 64 ± 2 7 b
220 ± 2 5 3
229±38a
231 ± 4 2 a
V aline
1 7 4 ± 12
6 1 7 ± 3 5C
611 ± 39c
608±32b
512±45'
433 ± 3 2 '
379±36b
B e f o r e t h e first in flu x o f a m i n o a c i d b a s e d d i a l y s i s s o l u t i o n . ap < 0 . 0 5 : h p < 0 . 0 0 5 : c p < 0 . 0 0 0 5 as c o m p a r e d w ith the basal valu es.
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1
Fig. 2. Total concentration of 14 non-essential amino acids (ala nine. arginine, asparagine, aspartic acid, a-aminobutyric acid, citrulline. cysteine, glutamine, gluatmic acid, glycine, ornithine, serine, taurine, tyrosine) in dialysate at different dwell times during amino acid exchanges (♦ ) and during glucose exchanges (0 ). The values represent mean ± SEM (n = 9). Fig. 3. Effect of amino acid based CAPD fluid (closed symbols) versus glucose based dialysis solution (open symbols) on the perito neal loss of total protein (♦ / 6 h). peritoneal ultrafiltration and clearance rates of small solutes did not differ significantly between amino acid solution and glucose-based dialysis fluid. These data are in accordance with the results of several previous studies [20.21,23]. Furthermore, amino acid based dialy sate was found to enhance the peritoneal loss of plasma proteins into the dialysis fluid to a clinically relevant extent. The CAPD-associated loss o f protein increased from 2.62 ± 0.45 g/8 h using conventional 4.25% glucose solution to 3.85 ± 0.42 g/8 h with amino acid based dial ysate. The augmented peritoneal protein loss was ob served for all individual proteins investigated, indepen dent of their molecular weight (fig. 3). The dialysate PGE2 concentration during the use o f the amino acid solution was found to be positively correlated with the enhanced permeation of total plasma protein into the dialysate
66
(fig. 4). Significant positive correlations were also estab lished between dialysate PGE2 and various individual proteins. Since PGE2 is known to possess vasodilative and permeability-increasing properties [36, 37], it is rea sonable to assume that the observed enhanced peritoneal permeability for plasma proteins during the use of amino acid based dialysate is mediated by locally generated p g e 2. The observed stimulation of peritoneal prostanoid gen eration rises further questions. The amino acid induced release of PGE2 may be a sign of local non-bacterial inflam mation. The initial abdominal pain in some of our patients after instillation o f the amino acid solution supports this assumption. Similar clinical symptoms during the use of amino acid based CAPD fluid were observed by Hanning ct al. [23]. Since repeated peritoneal inflammation is sup posed to contribute to impaired ultrafiltration and ana tomic changes of the peritoneum [38-40], amino acid based dialysis solutions should be used with caution. Recently prostaglandins of the E series were established to induce specific morphological changes in connective tissue [41. 42], PGE2 is supposed to promote the extrusion of 'matrix lysosomes' from myofibroblasts into the extracel lular space. The consecutive release of lysosomal enzymes such as proteases and phospholipases leads to the degrada tion of fibrinogen and fibrin to split products which are known to stimulate myofibroblast proliferation [41], By this way locally released PGE2 may contribute to the development of peritoneal fibrosis. In our opinion, the use of an amino acid containing dialysis solution is at present no convincing way to improve the nutrition status of CAPD patients. The observed change in peritoneal permeability with clinically relevant increased losses of proteins and non-essential amino acids is probably caused by amino acid induced locally generated prostanoids. Further long-term studies are necessary to elucidate the effect of continuous stimulation of peritoneal PG syn thesis and o f increased peritoneal leakage for macromole cules by dialysate amino acids on the clinical outcome of CAPD. A ck n o w le d g m e n ts This work was supported in part by a grant from the Fresenius Foundation, FRG . Portions of this work were presented at the 8th National Conference on CAPD. Kansas City, Mo.. USA, 1988. and at the 26th Congress of EDTA. Göteborg. Sweden. 1989. The authors wish to thank Mrs. K. Bappert and Mrs. G. Meier for their excellent technical assistance and Mrs. M. Baur for her careful secretarial assistance.
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D iscussio n
R eferences 15 Lindholm B. Bergstrom J: Nutritional aspects on CAPD: in Gokal R (ed): Continuous Ambu latory Peritoneal Dialysis. London, Churchill Livingstone. 1986. p 228. 16 Schilling H. Wu G. Pettit J. Harrison J. McNeil K, Siccion Z. Oreopoulos DG: Nutritional sta tus of patients on long-term CAPD. Périt Dial Bull 1985:5:12-18. 17 Giordano C. De Santo NG. Pluvio M. Di Leo VA, Capodicasa G, Carillo D. Esposito R, Damiano M: Protein requirement of patients on CAPD: A study on nitrogen balance. Int J ArtifOrgans 1980:3:11-16. 18 Gahl GM. Baeyer HV. Avcrdunk R: Outpa tient evaluation o f dietary intake and nitrogen removal in continuous ambulatory' peritoneal dialysis. Ann Intern Med 1981:94:643-646. 19 Blumenkrantz M J. Kopplc JD . Moran JK . Co burn JW : Metabolic balance studies and di etary protein requirements in patients un dergoing continuous ambulatory peritoneal di alysis. Kidney Int 1982;21:849-861. 20 Williams PF. Marliss EB. Anderson GH. Oren A, Stein AN. Khanna R. Pettit J. Brandes L. Rodclla H. Mupas L. Dombros N. Oreopoulos DG: Amino acid absorption following intra peritoneal administration in CAPD patients. Peril Dial Bull 1982;2:124-130. 21 Oren A. Wu G. Anderson GH. Marliss E, Khanna R. Pettit J. Mupas L, Rodclla H. Roncari DA. Kakis G. Harrison J. McNeil K. Oreo poulos DG: Effective use o f amino acid dialy sate over four weeks in CAPD patients. Périt Dial Bull 1983;3:66-73. 22 Pedersen FB. Dragsholt C. I.aier E. Frifelt JJ. Trostmann AF. Ekelund S. Paaby P: Alternate use of amino acid and glucose solution in CAPD. Périt Dial Bull 1985;5:215-218. 23 Hanning RM. Balfe JW , Zlotkin SH: Effective ness and nutritional consequences on amino acid-based vs. glucose-based dialysis solutions in infants and children receiving CAPD. Am J Clin Nutr 1987:46:22-30. 24 Young GA. Dibble JB . Hobson SM. Tompkins L. Gibson J. Turney JH . Brownjohn AM: The use of an amino-acid-based CAPD fluid over 12 weeks. Nephrol Dial Transplant 1989:4: 285-292. 25 Jontofsohn R. Trivisas G. Katz N. Kluthe R: Amino acid content of erythrocytes in uremia. Am J Clin Nutr 19 7 8 :3 1:1956-1 960. 26 Yatzidis H: New colorimetric method for quantitative determination of protein in urine. ClinChem 1977:23:811-812. 27 Dray F. Charbonnel B. Maclouf J: Radioim munoassay of prostaglandins F. Ei and Ey in human plasma. Eur J Clin Invest 1975:5:311318. 28 Steinhaucr HB. Batsford S. Schollmeyer P. Kluthe R: Studies on thromboxane By and prostaglandin Ey in the course of murine au toimmune disease: Inhibition by oral histidine and zinc supplementation. Clin Nephrol 1985: 24:63-68.
29 Steirhauer HB. Wilms H. Rüther M. Schollmeyer P: Clinical experience with urine TXBy in acute renal allograft rejection. Transplant Proc 1986; 18(suppl ):9 8 -103. 30 Dombros N. Oren A. Marliss EB. Anderson GH. Stein AN, Khanna R. Pettit J. Brandes L. Rodclla H. Leibl SB. Oreopoulos DG: Plasma amino acid profiles and amino acid losses in patients undergoing CAPD. Peril Dial Bull 1982;2:27-32. 31 Katirtzoglou A. Oreopoulos DG. Husdan H. Leung M. Ogilvie R, Dombros N: Reappraisal of protein losses in patients undergoing contin uous ambulatory peritoneal dialysis. Nephron 1980:26:230-233. 32 Steinhauer HB. Schollmeyer P: Prostaglandinmediated loss of proteins during peritonitis in continuous ambulatory peritoneal dialysis. Kidney Int 1986:29:584-590. 33 Bruno M. Bagnis C. Marangella M. Rovcra L. Cantaluppi A. Linari F: CAPD with an amino acid dialyis solution: A long-term, cross-over study. Kidney Int 1989:35:1189-1194. 34 Dombros NV. Prutis K. Tong M, Anderson GH. Harrison J. Sombolos K. Digenis G. Pettit J. Oreopoulos DG: Six-month overnight intra peritoneal amino-acid infusion in continuous ambulatory peritoneal dialysis (CAPD) pa tients: No effect on nutritional status. Périt Dial Int 1990:10:79-84. 35 Dibble JB . Young GA, Hobson SM. Brown john AM: Amino-acid-based continuous am bulatory peritoneal dialysis (CAPD) fluid over twelve weeks: Effects on carbohydrate and lipid metabolism. Peril Dial Int 1990:10:7177. 36 Williams T J. Peck MJ: Role o f prostaglandinmediated vasodilation in inflammation. Na ture 1977:270:530-532. 37 Murota SI. Morita I: Effect of prostaglandin ly and related compounds on vascular permeabil ity responses in granuloma tissue. Prostaglan dins 1978:15:297-301. 38 Steinhaucr HB. Frei A. Drcyling KW. Scholl meyer P: Changes of eicosanoid metabolism in CAPD-associated peritonitis; in Augustin R (ed): Peritonitis in CAPD. Contrib Nephrol. Basel. Karger. 1987, vol 57. pp 45-54. 39 Oreopoulos DG: Peritoneal membrane: Han dle with care. Peril Dial Bull I983;3:l 11-113. 40 Backenroth-Maayan R. Longnecker R. Kambosos D: Failure o f the peritoneal membrane during chronic intermittent peritoneal dialysis; in Gahl GM. Kessel M. Nolph KD (eds): Ad vances in Peritoneal Dialysis. Amsterdam. Excerpta Medica. 19 8 1. pp 2 0 8 -2 13. 4 1 Riede UN: Pathogenesis of shock-induced pul monary fibrosis in man as a model of prolifera tive inflammation: in Deicher H. Schulz LC (eds): Arthritis. Models and Mechanisms. Ber lin, Springer, 19 8 1. pp 88-95. 42 Joh K. Riede UN. Zahradnick HP: The effect of prostaglandins on the lysosomal function in the cervix uteri. Arch Gynecol 1983:234:1-16.
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1 Kopple JD : Abnormal amino acid and protein metabolism in uremia. Kidney Int 1978:14: 340-348. 2 Bansal VK, Popli S. Pickering J, Ing TS. Vertuno LL. Hano JE : Protein-caloric malnutri tion and cutaneous anergy in hemodialysis maintained patients. Am J Clin Nutr 1980:33: 1608-1611. 3 Blumenkrantz MJ. Kopple JD . Gutman RA. Chan YK. Barbour GL. Roberts C. Shen FH. Gandhi VS, Tucker CT. Curtis FK, Coburn JW : Methods of assessing nutritional status of patients with renal failure. Am J Clin Nutr 1980:33:1567-1585. 4 Alvestrand A. Bergström J: Nutritional man agement; in Suki WN. Massry SG (eds): Ther apy of Renal Disease and Related Disorders. The Hague. Nijhoff. 1984. p 459. 5 De Santo NG. Capodicasa G. Senatore R. Cicchctti T. Cirillo D. Damiano M. Torella R. Giugliano D. Improta L. Giordano C: Glucose utilization from dialysate in patients on contin uous ambulatory peritoneal dialysis (CAPD). Int J Artif Organs 1979:2:119-124. 6 Grodstein GP. Blumenkrantz MJ. Kopple JD . Moran JK . Coburn JW : Glucose absorption during continuous ambulatory peritoneal dial ysis. Kidney Jnt 1981:19:564-567. 7 Blumenkrantz MJ. Schmidt RW: Managing the nutritional concerns of the patient undergoing peritoneal dialysis: in Nolph KD (ed): Perito neal Dialysis. The Hague. Nijhoff. 1981. pp 2 7 5 -3 0 8 .' 8 Bergstrom J: Protein catabolic factors in pa tients on renal replacement therapy. Blood Purif 1985:3:215-236. 9 Giordano C. De Santo NG, Capodicasa G. Di Leo VA. Di Serafino A. Carillo D. Esposito R. Fiore R. Damiano M. Buonadonna L, Cocco F. Di Iorio B: Amino acid losses during CAPD. Clin Nephrol 1980:14:230-232. 10 Rubin J. Nolph KD, Arfania D. Prowant B. Frulo L. Brown P. Moore H: Protein losses in continuous ambulatory peritoneal dialysis. Nephron 1981:28:218-221. 11 Blumenkrantz MJ. Gahl GM. Kopple JD . Kamdar AV. Jones MR. Kessel M. Coburn JW: Protein losses during peritoneal dialysis. Kid ney Int 1981:19:593-602. 12 Kopple JD . Blumenkrantz MJ. Jones MR. Mo ran JK . Coburn JW : Plasma antino acid levels and amino acid losses during continuous am bulatory peritoneal dialysis. Am J Clin Nutr 1982:36:395-402. 13 Heide B. Picrratos A. Khanna R. Pettit J . Ogil vie R. Harrison J, McNeil K. Siccion Z. Oreopoulos D: Nutritional status o f patients un dergoing continuous ambulatory peritoneal di alysis (CAPD). Peril Dial Bull 1983:3:138141. 14 Kaysen GA, Schoenfeld PY: Albumin homeo stasis in patients undergoing continuous ambu latory peritoneal dialysis. Kidney Int 1984:25: 107-114.
