Cristina Robaudo

Kidney, splanchnic, and leg protein turnover in humans. Insight from leucine and phenylalanine kinetics.

The rate of kidney protein turnover in humans is not known. To this aim, we have measured kidney protein synthesis and degradation in postabsorptive humans using the arterio-venous catheterization technique combined with 14C-leucine, 15N-leucine, and 3H-phenylalanine tracer infusions. These measurements were compared with those obtained across the splanchnic bed, the legs (approximately muscle) and in the whole body. In the kidneys, protein balance was negative, as the rate of leucine release from protein degradation (16.8 +/- 5.1 mumol/min.1.73 m2) was greater (P < 0.02) than its uptake into protein synthesis (11.6 +/- 5.1 mumol/min. 1.73 m2). Splanchnic net protein balance was approximately 0 since leucine from protein degradation (32.1 +/- 9.9 mumol/min. 1.73 m2) and leucine into protein synthesis (30.8 +/- 11.5 mumol/min. 1.73 m2) were not different. In the legs, degradation exceeded synthesis (27.4 +/- 6.6 vs. 20.3 +/- 6.5 mumol/min. 1.73 m2, P < 0.02). The kidneys extracted alpha-ketoisocaproic acid, accounting for approximately 70% of net splanchnic alpha-ketoisocaproic acid release. The contributions by the kidneys to whole-body leucine rate of appearance, utilization for protein synthesis, and oxidation were approximately 11%, approximately 10%, and approximately 26%, respectively; those by the splanchnic area approximately 22%, approximately 27%, and approximately 18%; those from estimated total skeletal muscle approximately 37%, approximately 34%, and approximately 48%. Estimated fractional protein synthetic rates were approximately 42%/d in the kidneys, approximately 12% in the splanchnic area, and approximately 1.5% in muscle. This study reports the first estimates of kidney protein synthesis and degradation in humans, also in comparison with those measured in the splanchnic area, the legs, and the whole-body.

Renal ammoniagenesis in an early stage of metabolic acidosis in man.

Total renal ammonia production and ammonia precursor utilization were evaluated in patients under normal acid-base balance and in patients with 24-h NH4Cl acidosis by measuring (a) ammonia excreted with urine and that added to renal venous blood, and (b) amino acid exchange across the kidney. In 24-h acidosis not only urinary ammonia excretion is increased, but also total ammonia production is augmented (P less than 0.005) in comparison with controls. By evaluating the individual role of acid-base parameters, urine pH and urine flow in influencing renal ammonia production, it was shown that the degree of acidosis and urine flow are likely major factors stimulating ammoniagenesis. Both urine pH and urine flow are determinant in the preferential shift of ammonia into urine. In 1-d acidosis, renal extraction of glutamine was not increased and the total ammonia produced/glutamine N extracted ratio was higher than in controls (P less than 0.005) and was inversely correlated with the log of arterial bicarbonate concentration (P less than 0.001). In the same condition, renal glycine and ornithine uptake took place; the more severe the acidosis, the greater was the renal extraction of these amino acids (P less than 0.001). These data indicate that at the early stages of metabolic acidosis, in spite of a brisk increase in ammonia production, the mechanisms responsible for the increased glutamine use, which are operative in chronic acidosis, are not activated and other ammonia precursors, besides glutamine, are probably used for ammonia production.

Treatment of diabetic nephropathy in its early stages

Diabetic nephropathy is one of the most frequent causes of end‐stage renal disease (ESRD), and, in recent years, the number of diabetic patients entering renal replacement therapy has dramatically increased. The magnitude of the problem has led to numerous efforts to identify preventive and therapeutic strategies. In normoalbuminuric patients, optimal glycemic control (HbA1c lower than 7.5%) plays a fundamental role in the primary prevention of ESRD [weighted mean relative risk reduction (RRR) ∼37% for metabolic control versus trivial renoprotection for intensive anti‐hypertensive therapy or ACE‐inhibitors (ACE‐I)]. In the microalbuminuric stage, strict glycemic control probably reduces the incidence of overt nephropathy (weighted mean RRR ∼50%), while blood pressure levels below 130/80 mmHg are recommended according to the average blood pressure levels obtained in various studies. In normotensive patients, ACE‐I markedly reduce the development of overt nephropathy almost regardless of blood pressure levels; in hypertensive patients, ACE‐I are less clearly active (weighted mean RRR ∼23% versus other drugs), whereas angiotensin‐receptor blockers (ARB) appear strikingly renoprotective. Once overt proteinuria appears, it is uncertain whether glycemic control affects the progression of nephropathy. In type 1 diabetes, various anti‐hypertensive treatments, mainly ACE‐I, are effective in slowing down the progression of nephropathy; in type 2 diabetes, two recent studies demonstrate that ARB are superior to conventional therapy or calcium channel blockers (CCB). In clinical practice, pharmacological tools are not always used to the best benefit of the patients. Therefore, clinicians and patients need to be educated regarding the renoprotection of drugs inhibiting the renin‐angiotensin system (RAS) and the overwhelming importance of achieving target blood pressure. Copyright

Effects of a Protein Meal on Blood Amino Acid Profile in Patients with Chronic Renal Failure

Arterial whole blood levels of amino acids were determined in patients with chronic renal failure and in healthy subjects before and after 270 min after the ingestion of a grilled beefsteak (4 g/kg). In patients, total nonessential amino acids increased significantly more (+46%) than in controls owing to an exaggerated rise of serine, glutamine, proline, glycine, cyst(e)ine and alanine. Total essential amino acids increased as much as in controls; however, threonine, histidine and phenylalanine showed greater increases, while tryptophan had a smaller increment. Abnormalities in amino acid levels were even more evident in the postprandial period than in the postabsorptive state owing to reduced levels of valine, leucine, tryptophan, tyrosine, aspartate and glutamate and higher levels of glutamine, proline, glycine, cyst(e)ine, threonine, histidine and phenylalanine. Moreover, after the meal, the ratios total essential amino acids/total nonessential amino acids, valine/glycine, and branched-chain amino acids/total amino acids rose but persisted to be reduced whereas tryptophan/total amino acids and tyrosine/phenylalanine ratios increased in controls, but did not change in patients. In conclusion, in chronic renal failure, protein ingestion enhances the imbalance in amino acid levels already present in the postabsorptive state. The all data indicate that in patients with chronic renal failure, the metabolism of exogenous protein is impaired and the flow of amino acids to the organs is altered during the phase of body nitrogen replenishment.

Kidney Protein Dynamics and Ammoniagenesis in Humans with Chronic Metabolic Acidosis

To evaluate the effects of chronic metabolic acidosis on protein dynamics and amino acid oxidation in the human kidney, a combination of organ isotopic ((14)C-leucine) and mass-balance techniques in 11 subjects with normal renal function undergoing venous catheterizations was used. Five of 11 studies were performed in the presence of metabolic acidosis. In subjects with normal acid-base balance, kidney protein degradation was 35% to 130% higher than protein synthesis, so net protein leucine balance was markedly negative. In acidemic subjects, kidney protein degradation was no different from protein synthesis and was significantly lower (P < 0.05) than in controls. Kidney leucine oxidation was similar in both groups. Urinary ammonia excretion and total ammonia production were 186% and 110% higher, respectively, and more of the ammonia that was produced was shifted into urine (82% versus 65% in acidemic subjects versus controls). In all studies, protein degradation and net protein balance across the kidney were inversely related to urinary ammonia excretion and to the partition of ammonia into urine, but not to total ammonia production, arterial pH, [HCO(-)(3)], urinary flow, the uptake of glutamine by the kidney, or the ammonia released into the renal veins. The data show that response of the human kidney to metabolic acidosis includes both changes in amino acid uptake and suppression of protein degradation. The latter effect, which is likely induced by the increase in ammonia excretion and partition into the urine, is potentially responsible for kidney hypertrophy.

Vitamin E-coated filter decreases levels of free 4-hydroxyl-2-nonenal during haemodialysis sessions

Uraemic subjects undergoing chronic haemodialysis show increased oxidative stress. The use of non-biocompatible filters and reduced antioxidative defences are important sources of reactive oxygen species (ROS) release. The highly oxidative environment accelerates the onset and progression of tissue damage and atherosclerotic cardiovascular disease. The aldehyde 4-hydroxyl-2-nonenal (HNE) is probably the best marker of oxidative stress. In this study, the concentration of plasma HNE was evaluated in eight uremic subjects during two sessions of haemodialysis: the first using a standard biocompatible filter and the second using a filter coated with vitamin E. Baseline plasma levels of HNE were elevated, and dropped during haemodialysis. At the end of the session, however, low levels were maintained only when the vitamin E-modified filter was used. By contrast, a marked increase in HNE was recorded at the end of the session in all subjects who underwent haemodialysis with the conventional filter. This study provides evidence that the vitamin E-coated filter plays a role in counteracting oxidative stress. The chronic use of vitamin E-modified filters in haemodialysed subjects might help to counterbalance oxidative attack and, consequently, contribute to preventing cardiovascular disease.

Renal Metabolism of C-Peptide in Patients with Early Insulin-Dependent Diabetes mellitus

Renal metabolism of C-peptide was studied in 6 patients with early insulin-dependent diabetes mellitus (IDDM) with residual beta cell activity and in 11 nondiabetic subjects by the arterial-venous difference technique both in the postabsorptive state and for 80 min after ingestion of an amino acid mixture (0.8 g/kg). Urinary C-peptide (Cp) excretion, glomerular filtration rate and renal plasma flow were also measured. In the postabsorptive state in IDDM, renal uptake of Cp is reduced, while its urinary excretion and clearance are significantly increased. As a result, net renal extraction is markedly reduced. In contrast to controls, renal uptake and net extraction of C-peptide after amino acid ingestion do not increase in patients; the peritubular uptake evident in normal subjects is not detectable. Urinary excretion and clearance of Cp remain significantly higher in IDDM patients. In both groups, renal uptake of C-peptide is directly related to its renal load: however, in IDDM, the increase in Cp uptake for each increment in renal load is 35% lower than in controls (p < 0.001). Furthermore, as opposed to controls, urinary Cp excretion is not correlated with its arterial levels. Therefore IDDM patients have marked defects in renal handling of endogenous Cp, regarding both the amount metabolized by renal tissue and that reabsorbed by tubular cells. These data indicate an early alteration in the diabetic kidney that also impairs the reliability of urinary Cp evaluation as an index of residual beta cell activity in IDDM patients.

Effects of a new amino acid supplement on blood AA pools in patients with chronic renal failure

SummaryWe designed a new formula for AA supplement in order to correct blood pools of AA in chronic renal failure (CRF). This supplement was given to 5 patients with CRF and its effectiveness was tested during long term (12–24 weeks) administration. The patients had previously been on a diet providing 0.6 g of protein and 34–36 kcal/kg/day. The diet was then modified to one providing the same caloric content but only 0.3 g/kg high biological value protein per day with the addition of the AA supplement (0.3 g/kg). The new diet corrected most of the abnormalities in blood AA pools. After 1 month of treatment Val, Leu, Thr, Ser and Tyr levels rose and became normal throughout the study. Ratios Tyr/Phe, Ser/Gly and Val/Gly also improved. During the treatment no side effect or toxicity was observed, and serum albumin, transferrin and nutritional anthropometric parameters persisted to be normal. It is concluded that this specially designed AA supplement added to a hypoproteic diet is an acceptable regimen which can quite completely correct the imbalance in blood AA pools in CRF.

Effects of hemodialysis on guanidinopropionic acid metabolism.

Blood levels of guanidinopropionic acid (GPA), a putative uremic toxin, have been evaluated in 5 uremic patients before a dialytic session, at the end of it and during the following 68 h. GPA levels are markedly higher in uremic patients than in controls and are significantly reduced at the end of dialysis even if still higher than in controls. The clearance of GPA is similar to those of urea and creatinine, even if at the end of the dialysis session the percent decrease of GPA is significantly lower than that of urea. During the first 8 h after the end of dialysis GPA levels increase steeply; subsequently, the rate of accumulation of GPA in blood declines markedly remaining constant until the 68th hour. In conclusion GPA is markedly increased in blood of uremic patients and is significantly removed by dialysis. The evaluation of GPA increase per hour after the end of dialysis may provide an estimation of GPA production in uremic patients.

Erythropoietin Treatment and Amino Acid Metabolism in Hemodialysis Patients

A previous report suggests that treatment with recombinant human erythropoietin (rH-EPO) significantly improves many abnormalities in circulating amino acids (AA) in hemodialysis patients. We evaluated the effects of a 12-month treatment with rH-EPO (150-250 U/kg/week) on blood AA levels in 10 patients with chronic renal failure under regular dialytic treatment. During treatment, hemoglobin levels increased from 7.0 +/- 0.3 to 10.1 +/- 0.3 g/dl at 3 months remaining steady thereafter. Before the treatment, patients showed reduced levels of essential AA (EAA), mainly valine, leucine and threonine (p < 0.05-0.01); among non-EAA (NEAA), aspartate and serine were reduced, whereas glycine, alanine, proline, citrulline and cyst(e)ine were increased (p < 0.05-0.001). Val/Gly, Ser/Gly and Tyr/Phe ratios were low (p < 0.05-0.01). Total EAA and total NEAA (619 +/- 21 and 1,382 +/- 75 mumol/l, respectively, before the study) were unchanged (639 +/- 22 and 1,410 +/- 89 mumol/l, respectively) at 12 months. Abnormalities in AA levels observed before the treatment persisted throughout the study. Only serine increased at the end of the study (p < 0.05). In conclusion, contrary to what has been reported, treatment with rH-EPO is not associated with an amelioration of AA metabolism in hemodialysis patients.