Light chain polymerism in normal individuals in patients with severe proteinuria and in normals with inhibited tubular protein reabsorption by lysine

The polymeric forms of free kappa and lambda light chains were estimated in serum and urine from twelve normal individuals, from seven patients with nephrotic syndrome associated with normal GFR, and from eleven normal persons after inhibition of renal tubular protein reabsorption by lysine. The investigation showed that the serum concentrations of the monomeric and dimeric forms of light chains were similar in the three groups. The urinary excretion was increased in patients with nephrotic syndrome, the mean excretion of kappa chains being 9.2 mg/24 h and of lambda 6.6 mg/24 h compared with normal excretion of 2.2 mg/24 h for kappa and 1.0 mg/24 h for lambda. After inhibition of tubular protein reabsorption, the urinary light chains excretion increased to 101.3 mg/24 h for kappa and 40.0 mg/24 h for lambda chains, i.e. approximately 40 times normal level. The relative concentrations in the urine of the monomeric and dimeric forms of the two chain types were varied widely in the three groups investigated, particularly for lambda monomers where a low excretion rate and clearance value was found during inhibition of the protein reabsorption. The reason for the variations were found to be differences in polymerization and consequently differences in glomerular filtration. Evidence for a preferential reabsorption of any of the chain types or polymers was not seen.     

Tubular reabsorption of pepsinogen isozymogens in man studied by the inhibition of tubular protein reabsorption with dibasic amino acids.

1. The fractional clearances of pepsinogen A (PGA), pepsinogen C (PGC) and the main PGA isozymogens, i.e. PGA-3, PGA-4 and PGA-5, were measured in 13 healthy male volunteers before and during blockade of tubular protein reabsorption by intravenous infusion of either L-arginine hydrochloride (n = 8; 0.5 g h-1 kg-1 body weight) or an equimolar amount of L-lysine hydrochloride (n = 5; 0.44 g h-1 kg-1 body weight). Glomerular filtration rate was measured by a radioisotope method. 2. The fractional baseline clearance of PGC (1 +/- 1%) was lower than that of PGA (20 +/- 10%). In addition, the fractional clearance of the PGA isozymogens appeared to be different: the fractional clearance of PGA-5 (7 +/- 3%) was lower than that of PGA-4 (18 +/- 9%), and the fractional clearance of PGA-4 was lower than that of PGA-3 (30 +/- 10%). These differences in fractional clearance between PGA isozymogens decreased during infusion of both arginine and lysine. 3. Pepsinogens are freely filtered proteins. It can therefore be concluded that the differences in fractional clearance between PGA isozymogens imply differences in tubular reabsorption. This is remarkable as PGA isozymogens are proteins with an almost identical amino acid sequence and electric charge. The disappearance of the differences in tubular reabsorption during arginine and lysine infusion suggests that PGA isozymogens differ in affinity for negatively charged binding sites in the tubular cell membrane. In order to explain the low fractional clearance of PGC compared with that of PGA and the less marked effect of arginine or lysine infusion on the fractional clearance of PGC, an additional PGC-specific binding site has to be postulated.     

Cubilin is an albumin binding protein important for renal tubular albumin reabsorption

Using affinity chromatography and surface plasmon resonance analysis, we have identified cubilin, a 460-kDa receptor heavily expressed in kidney proximal tubule epithelial cells, as an albumin binding protein. Dogs with a functional defect in cubilin excrete large amounts of albumin in combination with virtually abolished proximal tubule reabsorption, showing the critical role for cubilin in the uptake of albumin by the proximal tubule. Also, by immunoblotting and immunocytochemistry we show that previously identified low-molecular-weight renal albumin binding proteins are fragments of cubilin. In addition, we find that mice lacking the endocytic receptor megalin show altered urinary excretion, and reduced tubular reabsorption, of albumin. Because cubilin has been shown to colocalize and interact with megalin, we propose a mechanism of albumin reabsorption mediated by both of these proteins. This process may prove important for understanding interstitial renal inflammation and fibrosis caused by proximal tubule uptake of an increased load of filtered albumin.     

Effects on renal tubular protein reabsorption of the infusion of free haemoglobin

Abstract. Hydroxy-ethyl-starch cryopreserved blood with a mean free haemoglobin content of 260 μmol/l (SD 57 μmol/l) was infused into five normal volunteers. The renal clearance of six proteins of differing molecular weight was measured together with an analysis of the qualitative changes in protein excretion during the infusion and over the following 48 h. Other aspects of renal tubular function were assessed by the measurement of urinary N -acetyl-β- D -glucosaminidase, phosphate and amino acids. Increased clearance of low molecular weight proteins and N -acetyl-β- D -glucosaminidase occurred during the 12 h following infusion. β₂ microglobulin clearance × 10³ creatinine clearance rose from a mean of 0·4 (SD 0·1) to a mean of 74·5 (SD 32·3) and N -acetyl-β- D -glucosaminidase from 54·2 units/mmol creatinine (SD 18·0) to 1525 units/mmol creatinine (SD 2318). Increased amounts of low molecular weight proteins were detected in the urine by crossed immunoelectrophoresis. Creatinine clearance remained unaltered. It is argued that these changes may be caused by the competitive inhibition of low molecular weight protein reabsorption in the renal tubule by filtered haemoglobin.     

Ultrafiltration of renin in the mouse kidney studied by inhibition of tubular protein reabsorption with lysine

1. In order to study the role of the kidney in the elimination of endogenous plasma renin, renin was measured in the plasma and urine of female mice. 2. The renin concentration was two orders of magnitude lower in urine than in plasma, but it increased after intraperitoneal injection of submandibular mouse renin. No correlation between the plasma renin concentration and the urinary renin concentration and renin excretion rate could be demonstrated. 3. Blockade of the tubular reabsorption of proteins by intraperitoneal injection of the basic amino acid lysine increased the urinary renin concentration, renin excretion rate and renin clearance two to three orders of magnitude, without affecting the plasma renin concentration. 4. This finding demonstrates that ultrafiltered renin is reabsorbed almost completely in the renal tubules and that the mechanism most likely is the same as for other filtered proteins. 5. The large renal renin clearance obtained after intraperitoneal lysine is in accordance with a major role of the kidneys in the elimination of renin from the circulation, by a glomerular ultrafiltration and tubular reabsorption and metabolization of renin.     

Electrophoretic examination of proteinuria in Lowe's syndrome and other causes of renal tubular Fanconi syndrome

Urine samples from 26 patients with five different causes of renal tubular Fanconi syndrome were examined by zone electrophoresis on agarose gel and immunofixation. The tubular disorders associated with Lowe's syndrome, cystinosis, and idiopathic Fanconi syndrome exhibited urine protein electrophoretic characteristics that differentiated them from normal and from each other. In particular, Lowe's syndrome urine exhibited four discrete bands in the gamma globulin zone. Electrophoresis of urinary proteins may be useful in distinguishing among the different metabolic disorders causing renal tubular Fanconi syndrome.     

The Coomassie Brilliant Blue method underestimates drug-induced tubular proteinuria

The sensitivity of the Coomassie Brilliant Blue (CBB) procedure differs for different proteins. To investigate the impact of this variability on clinical measurements of proteinuria, we collected urine specimens from patients with glomerular disease or drug-induced tubular nephrotoxicity and compared urinary protein concentrations as determined by the CBB method with those obtained by the biuret method. The CBB method underestimated the protein concentration predicted by the biuret method by as much as eightfold in specimens with low proportions of albumin. Determinations of protein by the CBB method in serially diluted urine specimens from patients with tubular damage also deviated from linearity by as much as twofold for protein concentrations less than 1 g/L. We conclude that CBB binding is relatively insensitive for measuring proteins specifically associated with tubular damage and hence may be inappropriate for clinical use.     

Studies on renal tubular protein reabsorption: partial and near complete inhibition by certain amino acids.

Urinary excretion of albumin, free light chains of immunoglobulins and beta-2-microglobulin was measured after injection of certain amino acids and derivatives. Substances with a positively charged group located terminally in the molecule (ornithine, lysine, arginine, and the almost not metabolized epsilon-amino-caproic acid, and Cyclocaprone) proved to inhibit instantaneously the tubular protein reabsorption. Lysine is the most effective molecule tested. We hypothesize that the initial event in the normal tubular protein reabsorption is a binding between a free positive amino- or guanidino-group in the protein molecule and a negative site on the tubular cell surface. In an attempt to achieve complete inhibition of tubular protein reabsorption, large doses of lysine were injected intravenously. The maximal protein clearance rose from the exceedingly low baseline excretion to values probably representing near complete inhibition of protein reabsorption. The minimal value of albumin concentration in the glomerular filtrate was calculated to 2.6 mg/l.     

Measurement of urinary retinol-binding protein by enzyme-linked immunosorbent assay, and its application to detection of tubular proteinuria

An enzyme-linked immunosorbent assay (ELISA) for urinary retinol-binding protein (RBP) has been developed and compared with urinary beta 2-microglobulin for the detection of tubular proteinuria. The assay has a working range of 10 to 250 micrograms of RBP per liter of urine. The within-assay CV was 3.2-7.1%, the between-assay CV 12.5%. A control population of 118 male subjects gave a geometric mean urinary RBP concentration of 7.7 micrograms per millimole of creatinine and a 95th centile of 22 micrograms per millimole of creatinine. Comparison of urinary RBP and beta 2-microglobulin concentrations in 80 control subjects and 117 subjects exposed to cadmium fumes gave correlations of r = 0.59 and 0.91, respectively. Of the 117 subjects exposed to cadmium fumes, 103 gave both RBP and beta 2-microglobulin concentrations on the same side of the upper 95th centile values of 22 and 38 micrograms per millimole of creatinine for RBP and beta 2-microglobulin respectively (Chi-square analysis p less than 0.001), demonstrating that RBP and beta 2-microglobulin detect tubular proteinuria with equal sensitivity and specificity. ELISA and an established latex immunoassay gave well-correlated results.     

Filtration and reabsorption of protein by the kidney

Plasma proteins of the rat have been labelled by the in vivo injection of the dye T-1824. From a study of the rate of disappearance of T-1824 from the circulating blood, and the total T-1824 content of the perfused kidney the rate of protein reabsorption from the glomerular fluid by the cells of the renal tubule has been calculated. It is concluded that protein reabsorption by the cells lining the proximal convoluted tubule of the rat kidney proceeds at a rate of at least 5 mg. per hour, equivalent to a daily filtration and reabsorption of 33 per cent of the circulating plasma protein.     

The effect of sodium taurocholate on proximal tubular reabsorption in the rat kidney

Microperfusion of tubules in situ was used to study the direct effect of sodium taurocholate on reabsorption of fluid by the proximal tubule of the rat. Sodium taurocholate (0.1 mmol/l) in the tubular perfusate reduced proximal tubular fluid reabsorption by approximately 30%. Thus, the proximal tubule appears to be a major site at which bile salts cause a natriuresis in the rat, and possibly in obstructive jaundice in man.     

Changes in proximal and distal tubular reabsorption produced by rapid expansion of extracellular fluid

Acute infusions of isotonic saline in the rat cause an increase in glomerular filtration rate and in the excretion of salt and water. The kidney swells, due to expansion of tubular and interstitial volume. Despite the increase in tubular diameter, transit time through the proximal tubules and loops of Henle is decreased, presumably owing to a greatly accelerated rate of tubular flow. Proximal tubular reabsorption, measured in blocked tubules, is inhibited in a way that cannot be ascribed to changes in tubular diameter. The prolongation of proximal reabsorptive half-time is not affected by the administration of aldosterone. It occurs equally in rats chronically loaded with or deprived of salt, and it is therefore not likely that it is influenced by the renal content of renin. In contrast, reabsorption from the distal convoluted tubule is enhanced by saline infusion. This change is observed in segments of tubules blocked with oil and isolated from their glomeruli and thus appears to occur independently of changes in glomerular filtration or tubular flow.