Alberto Tizianello

Renal metabolism of amino acids and ammonia in subjects with normal renal function and in patients with chronic renal insufficiency.

The net renal metabolism of amino acids and ammonia in the post absorptive state was evaluated in subjects with normal renal function and in patients with chronic renal insufficiency by measuring renal uptake and release, and urinary excretion of free amino acids and ammonia. In normal subjects the kidney extracts glutamine, proline, citrulline, and phenylalanine and releases serine, arginine, taurine, threonine, tyrosine, ornithine, lysine, and perhaps alanine. The renal uptake of amino acids from arterial blood occurs by way of plasma only, whereas approximately a half of amino acid release takes place by way of blood cells. Glycine is taken up from arterial plasma, while similar amounts of this amino acid are released by way of blood cells. In the same subjects total renal ammonia production can be largely accounted for by glutamine extracted. In patients with chronic renal insufficiency (a) the renal uptake of phenylalanine and the release of taurine and ornithine disappear; (b) the uptake of glutamine and proline, and the release of serine and threonine are reduced by 80--90%; (c) the uptake of citrulline and the release of alanine, arginine, tyrosine, and lysine are reduced by 60--70%; (d) no exchange of glycine is detectable either by way of plasma or by way of blood cells; (e) exchange of any other amino acid via blood cells disappears, and (f) total renal ammonia production is reduced and not more than 35% of such production can be accounted for by glutamine extracted, so that alternative precursors must be used. A 140% excess of nitrogen release found in the same patients suggests an intrarenal protein and peptide breakdown, which eventually provides free amino acids for ammonia production.

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.

Enhanced activation of the respiratory burst oxidase in neutrophils from hypertensive patients.

In neutrophils of patients with essential hypertension the NADPH-dependent O2- production elicited by stimulation with f-Met-Leu-Phe is three to four fold higher in comparison with neutrophils of normotensive control subjects. Neutrophils from hypertensive patients are less responsive to priming, by non-stimulating doses of the agonist, as compared to control cells, which following this pretreatment augment superoxide anion production up to levels close to those expressed by neutrophils from hypertensive patients. No difference in NADPH oxidase activity, between neutrophils from the two groups of subjects, was observed when the rate of O2- production was evaluated in a reconstructed cell-free system containing the membrane fraction and the cytosolic cofactors. These results are consistent with the hypothesis that differences in the functional organization of the oxidase at the membrane level in neutrophils of hypertensive are responsible for the enhanced O2- production following agonist stimulation.

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.

Erythrocyte deficiency in calpain inhibitor activity in essential hypertension.

The calpain-calpain inhibitor system was evaluated in erythrocytes of patients with essential hypertension and normotensive controls, either with or without a family history of hypertension. Calpain levels were similar in the controls and hypertensive patients, whereas the inhibitor activity level was significantly reduced in the latter (301.8 ± 26.4 vs 220 ± 14 U/mg hemoglobin, p < 0.001). Borderline hypertensive patients and a few controls with a history of hypertension showed low inhibitor activity. Similar results have recently been reported in genetically hypertensive rats of the Milan strain. A significant inverse correlation (r = −0.43, p < 0.001) was found between mean arterial pressure and calpain inhibitor. Although the pathophysiotogical significance of these observations is not yet dear, they suggest a new area of investigation Into the molecular mechanisms underlying essential hypertension and its complications.

Increased phosphorylation in red cell membranes of subjects affected by essential hypertension

In hemolysates of red cells from hypertensive patients the proteolytic activity of calpain is expressed at a rate approximately three fold higher than in red cells of normotensive subjects. Susceptibility to lysis upon exposure to ionophore A23187 and calcium, conditions that increase intracellular calpain activity, is also significantly enhanced in erythrocytes of hypertensive patients. In inside-out vesicles prepared from erythrocytes of these patients band 3 region undergoes a high extent of phosphorylation which is 1.5 fold higher than that occurring in control red cells from normotensive subjects. This increased phosphorylation can be reproduced in inside-out vesicles from erythrocytes of normal subjects following pretreatment with calpain. Taken together, these results suggest that the presence in erythrocytes of hypertensive subjects of an unregulated calpain dependent proteolytic activity may affect the structure of plasma membranes and determine an increased phosphorylation of intrinsic membrane proteins.

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.

Abnormalities of Amino Acid Metabolism

Restriction of protein (PR) intake for patients with chronic renal failure (CRF) aims at reducing retention of waste metabolites and, at the same time, at preserving lean body mass [1]. An additional target is to slow down the progression of renal failure [1], although this possibility is not yet definitely proven in humans. Nutritional treatment of CRF patients has advanced remarkably over the last few years, although many aspects of nitrogen (N) metabolism in CRF are still obscure.

Renal metabolism of amino acids in early insulin-dependent diabetes mellitus

Renal metabolism of amino acids (AAs) was evaluated in 5 patients with early IDDM, and in 7 controls (C) in the basal state for 80 minutes after the ingestion of an AA mixture simulating an animal protein meal. Insulin was withdrawn 20 hours before the study. Renal metabolism of AAs was evaluated by the arterial-venous difference technique. In the basal state in IDDM, as in C, the kidney takes up large amounts of a few nonessential AAs (NEAAs): it releases many NEAAs and a few essential AAs (EAAs). After AA ingestion in C, renal extraction of most EAAs, mainly BCAAs, Lys, and Thr, occurs; Pro extraction also increases and a significant uptake of Gly, Glu, Asp, Orn, and Tyr takes place. EAA extraction accounts for 30-40% of total AA uptake. In IDDM, after AA ingestion, a) renal uptake of total AAs is significantly lower, owing mainly to a markedly lower uptake of BCAAs, Lys, and also of Pro, Orn, and Ala; b) renal EAA uptake accounts for less than 20% of total AA extraction. These results indicate that in IDDM postprandial renal N repletion is impaired and unbalanced.