Renal parenchymal thickness and urinary protein levels in patients with ureteropelvic junction obstruction after nephrostomy placement.

OBJECTIVE: To assess recovery of renal parenchymal thickness and urinary protein levels in patients with severely hydronephrotic kidneys after nephrostomy placement. METHODS: Fourteen patients (median age 1 year, range 6 months-7 years) who underwent nephrostomy placement for unilateral ureteropelvic junction obstruction at our hospital between May 2007 and January 2009 were included in a retrospective analysis. All patients had severe hydronephrosis, with a median parenchymal thickness of 1.8 mm (range 1-2.5 mm). Kidney morphology was examined by ultrasound before the procedure and 1, 2, 3, 4, 6 and 8 weeks after. Urinary proteins (including albumin, immunoglobulin [IgG], alpha2-macroglobulin, alpha1-microglobulin, beta2-microglobulin [beta2-MG] and kappa chain) and creatinine levels were also tested during these follow-up visits. Fifteen healthy children were assessed for urinary protein levels as well and made up the control group. RESULTS: Parenchymal thickness increased within 4 weeks of nephrostomy placement. Kidney volumes were significantly decreased within 2 weeks. No further changes in morphology were detected after 4 weeks. Urinary alpha1-microglobulin and beta2-MG levels decreased to baseline within 1 and 4 weeks, respectively. Urinary albumin, IgG, alpha2-macroglobulin and kappa chain levels decreased gradually after nephrostomy, but did not return to baseline within 8 weeks. CONCLUSIONS: After nephrostomy placement, parenchymal thickness increases within 4 weeks, tubular function returns to normal earlier than glomerular function and glomerular membrane repair is inversely correlated with the severity of damage.

Beta-2-microglobulin (B2M) expression in the urinary sediment correlates with clinical markers of kidney disease in patients with type 1 diabetes.

PURPOSE: After observing variation in the expression of the housekeeping gene B2M in cells of the urinary sediment during a study of candidate genes potentially involved in diabetic kidney disease (DKD), we hypothesized that B2M mRNA expression in the urinary sediment could reflect the presence of DKD. METHODS: qPCR was used to quantify B2M mRNA expression in cells of the urinary sediment of 51 type 1 diabetes (T1D) patients (61% women, 33.5 [27.0-39.7] years old, with diabetes duration of 21.0 [15.0-28.0] years and HbA1c of 8.2% [7.3-8.9]; median [interquartile interval]) sorted according to the diabetic nephropathy (DN) stages; 8 focal segmental glomerulosclerosis (FSGS) patients and 10 healthy controls. B2M mRNA expression was also evaluated in human embryonic kidney epithelium-like (HEK-293) cells exposed to 25mM glucose and to albumin in order to mimic, respectively, a diabetic and a proteinuric milieu. RESULTS: No differences were found in B2M mRNA expression among healthy controls, FSGS and T1D patients. Nonetheless B2M mRNA expression was higher in the group composed by T1D patients with incipient or overt DN combined with FSGS patients versus T1D patients without DN combined with healthy controls (P=0.0007). B2M mRNA expression was higher in T1D patients with incipient or overt DN versus without DN (P=0.03). B2M mRNA expression positively correlated with albuminuria in the overall T1D population (r=0.43; P=0.01) and negatively correlated with estimated glomerular filtration rate in male T1D patients (r=- 0.57; P=0.01). Increased B2M expression was observed in HEK-293 cells exposed to 25mM glucose and to albumin. CONCLUSIONS: Β2M mRNA expression in cells of the urinary sediment is higher in T1D patients with DKD and in patients with FSGS in comparison to healthy subjects, maybe reflecting a tubulointerstitial injury promoted by albumin. Given the proinflammatory nature of B2M, we suggest that this protein contributes to diabetic (and possibly, to non-diabetic) tubulopathy.

The effect of dietary protein on the excretion of alpha2u, the sex-dependent protein of the adult male rat.

Adult male rats were maintained on normal (20% casein), protein-free (0% casein), high protein (50% casein), decicient protein (20% zein), and a supplemented, deficient protein (20% zein plus L-lysine and L-tryptophan) diets. Rats on a protein-free diet excreted approximately 1 mg alpha2u/24 h compared with a normal of 10-15 mg/24 h. Depleted rats placed on a 20% casein diet showed a rapid restoration of the normal alpha2u excretion as well as total urinary proteins. Accumulation of alpha2u in the blood serum was measured in nep-rectomized rats. Rats on a 0% casein diet accumulated only 30% of the alpha2u compared to normals. On a 50% casein diet, rats excreted 30-50 mg alpha2u/24 h. However, the accumulation was normal in the serum of nephrectomized rats. A high protein diet did not stimulate alpha2u synthesis but probably increased the renal loss of all urinary proteins. The excretion of alpha2u on a zein diet was reduced to the same degree as with the protein-free diet. Supplementation with lysine and tryptophan restored the capacity to eliminate alpha21 to near normal levels. Accumulation of alpha2u in the serum of nephrectomized rats kept on the zein diets showed that the effect to suppress the synthesis of the ahpha2u. Supplementation restored the biosynthesis of alpha2u. We conclude that the effect of dietary protein on the excretion of urinary proteins in the adult male rat is caused in large part by an influence on the hepatic biosynthesis of alphay2u. The biosynthesis of this protein, which represents approximately 30% of the total urinary proteins, is dependent on an adequate supply of dietary protein.

The different electrophoretic forms of post gamma-globulin their antigenic identity and their structural variability.

Post gamma-globulin, first described as a constant component of protein of cerebrospinal fluid and urine from patients with tubular disorders has also been found in other biological fluids. Post gamma-globulin from a single individual always migrated as several bands after storage. Three electrophoretic forms, immunochemically identical, have been isolated by gel chromatography, preparative continuous flow electrophoresis and ion-exchange chromatography. Their molecular weight was found to be approximately between 11 000 to 12 000. No difference between the three forms could be detected. The N-terminal amino acids were found to be Lys, Arg and Leu respectively for the three forms of post gamma-globulin. The "slow" and "fast" forms of post gamma-globulin seemed to differ by elimination of small basic peptides or amino acids from the N-terminal end of the protein. No enzymatic activity of post gamma-globulin was found, but this requires further investigations.

Electrophoretic pattern of concentrated urine: comparison between 24-hour collection and random samples.

The electrophoretic pattern of concentrated urine samples can be used to identify the type of proteins leaking into the urine and has diagnostic and prognostic value, providing information about the location (glomerular or tubular) and degree of renal injury. This test usually requires a 24-hour urine collection, which can be inconvenient because of its heavy dependence on patient compliance and frequently is unreliable because of errors in collecting a complete 24-hour urine sample. In this study, we compared the electrophoretic pattern in 24-hour urine collections and random samples among patients with glomerular diseases and a wide range of proteinuria. Forty adult patients were evaluated; 24-hour urine collections and random urine samples were analyzed. Protein concentrations were determined using the sulfosalicylic acid method standardized with human serum. Electrophoresis was performed with concentrated urine samples (Ultrafree, PF/Millipore Corporation, Bedford, MA) using Beckman Paragon Electrophoresis System (agarose gels and blue staining; Beckman Instruments, Inc, Brea, CA). Densitometric scanning of electrophoretic pattern (Appraise Clinical Densitometer; Beckman Instruments, Inc) was performed, and the results were reported in percentages of each observed fraction. Our results revealed that despite the significant difference between protein concentration in 24-hour collections and in random samples, the pattern of protein excretion, in percentage basis, remains the same. There were no differences between the albumin, alpha(1)-globulin, alpha(2)-globulin, beta-globulin, and gamma-globulin fractions in both types of specimens. This study shows that, at least in glomerular proteinuria, the electrophoretic analysis of the urine can be performed accurately in random samples, avoiding the inconveniences and errors of a 24-hour urine collection.

Sensitivity of in vitro diagnostic dipstick tests to urinary protein.

Two commonly used brands of reagent strip (dipsticks) were evlauated for their sensitivity to Bence-Jones and seven other urinary proteins. Both brands showed significant differences in sensitivity to albumin, glycoprotein, ribonuclease and lysozyme; both were most sensitive to albumin and least sensitive to globulin. Furthermore, their comparative sensitivities to these proteins also differed markedly. These differences in sensitivity could lead to underestimation of protein content in urine specimens. Tests on urines from patients with multiple myeloma showed that a negative urinary dipstick test result did not rule out the presence of the disease.

[Autonomic determination of proteins from urine by centrifugal and continuous flow analyses (author's transl)]. / Automatisation en analyse centrifuge et en flux continu du dosage des protéines totales urinaires.

A precipitating reagent comprising partially neutralized trichloroacetic acid with magnesium sulfate, sodium dodecyl sulfate and polyethylene glycol is used for automatic determination of proteins from urine by centrifugal analysis and continuous flow analysis. We compare and discuss the results of both methods, their limits of validity and incorporation in an automated laboratory.

The use of preparative continuous flow electrophoresis to study low molecular weight proteins in urine.

The authors propose a convenient method for the separation of low molecular weight proteins, present in tubular urines. In particular, the method is used for the purification of beta2 microglobulin, after one recycling, and also for the isolation of the various post-gamma globulins which can later be separated on DEAE Sephadex.

Automated continuous flow determination of urine albumin by competition with dye-detergent binding.

A continuous flow automated method for urine albumin was developed based on the ability of albumin to displace bromophenol blue from a bromophenol blue detergent complex. The method is almost specific for albumin, giving a slight response with an alpha-globulin fraction from serum. Results agreed closely with rocket electrophoresis albumin estimates on urine samples, but significantly less closely with "total urine protein" estimates by an acid protein precipitation, followed by biuret colour reaction, procedure. This method is being used to replace the indefinite "total protein" test for most routine purposes.

Dysproteinuria induced in rats by O,O,S-trimethyl phosphorothioate.

Administration of a single oral dose of 60 mg/kg O,O,S-trimethyl phosphorothioate (OOS-Me), a malathion impurity, resulted in a substantial increase in the amounts of amino acids along with a change in the nature of proteins excreted in the urine of treated rats. In contrast to control rats, a small increase in albumin and a small decrease in alpha 1-globulin were observed. However, alpha2-, beta- and gamma 1-globulin, which were not detected in the urine of control rats, were found in substantial amounts in the urine of OOS-Me-treated rats. These findings, coupled with observed increases in urinary glucose levels and consistent specific gravity readings of 1.01 even though treated rats were experiencing oliguria, provide evidence for OOS-Me-induced kidney tubule damage.

Renal follow up of premature infants with and without perinatal indomethacin exposure.

AIMS: To evaluate early childhood renal growth, structure, and function in children born at less than 33 weeks gestation and to investigate possible independent effects of perinatal indomethacin exposure. METHODS: A total of 66 children born at less than 33 weeks gestation, 31 of them with perinatal indomethacin exposure (study group) and 35 without (control group), were examined at 2-4 years of age. Serum cystatin C and protein; plasma creatinine, sodium, and potassium; urine protein, calcium:creatinine ratios, and alpha(1) microglobulin; and glomerular filtration rate (GFR) were determined. Renal sonography examinations were performed. RESULTS: The mean serum cystatin C concentrations were slightly higher in the control group than in the study group. Mean values of serum protein, and plasma creatinine and sodium did not differ between the groups, neither did median plasma potassium concentrations and urine protein:creatinine and calcium:creatinine ratios. None had tubular proteinuria. Abnormal GFR (<89 ml/min/1.73 m(2)) was found in one case in each group and renal structural abnormalities in five in each group. In logistic regression analysis the duration of umbilical artery catheter (UAC) use and furosemide treatment emerged as the significant independent risk factors for renal structural abnormalities. Furosemide treatment and assisted ventilation remained the risk factors associated with renal abnormalities in general-that is, functional and/or structural abnormal findings. CONCLUSION: Perinatal indomethacin does not seem to affect long term renal growth, structure, or function in children born at less than 33 weeks gestation. Duration of UAC use, furosemide treatment, and assisted ventilation may be correlated with later renal structural and functional abnormalities.

Urinary protein excretion in healthy children.

The urinary total protein excretion was determined in 270, 18-24 hr urine samples from 130 healthy children of different age groups using the tannic acid-Fe3+-method of Yatzidis [1977]. The daily protein excretion of premature infants in the first month of life varies between 14-60 mg, with a mean of 29 mg, and that of fullterm newborn infants between 15-68 mg, with a mean of 32 mg. Protein excretion increases with age and amounts to 29-238 mg (mean 83 mg) in 10-16 year old children. Thus, the urinary protein concentration during the neonatal period is high when compared to adult values. This explains the "trace" and "positive" reactions frequently obtained in this period of life with Albustix. In 92 urine samples proteins were fractionated by sodium dodecyl sulphate gel discelectrophoresis. Hemoglobinuria was found during the first weeks of life and tubular type proteinuria was found in newborns and infants. The present data suggest that the proteinuria is due to ineffective proximal tubular reabsorption of low molecular weight microproteins as a result of glomerulo-tubular imblance in early life.

Composition of protein in urine from dogs with pyoderma.

The protein fractions in urine from proteinuric dogs with and without pyoderma were estimated. Fifteen dogs with pyoderma (five with superficial and 10 with deep pyoderma) were compared with 10 dogs with glomerulopathy and 27 dogs with diseases other than pyoderma or urinary tract problems. Agarose gel electrophoresis was used to fractionate the proteins. Three types of electrophoretogram were obtained with albuminuria, globulinuria and serum-like profiles. An albuminuria profile was found in eight of the 27 dogs with other diseases, in three of the five dogs with superficial pyoderma, in eight of the 10 dogs with deep pyoderma and in all 10 dogs with glomerulopathy. The albuminuria profile (mean [sem] albumin/globulin ratio 1.98 [0.10]) was also characterised by alpha 1b, alpha 2a and beta 2 globulin peaks in all 29 dogs with this profile, which was therefore thought to indicate that albuminuria (glomerular proteinuria) was a result of glomerular damage and inflammation because alpha 1b, alpha 2a, and beta 2 globulins are considered to be acute phase proteins. The serum-like profile (mean [sem] albumin/globulin ratio 0.72 [0.01]) was observed in 13 per cent of the proteinuric dogs examined and contained all the protein fractions normally detected by electrophoresis of serum. The profile was considered to be a variant from of the albuminuria profile, probably indicating advanced glomerular lesions and inflammation. The globulinuria profile (mean [sem] albumin/globulin ratio 0.33 [0.08]) was significantly different from the other two in that it was characterised by a low albumin peak and the presence of globulin fractions not clearly distinguishable from each other because of their confluency and absence of individual peaks. This profile could indicate severe glomerulotubular lesions and degradation of certain protein fractions. It could also be a result of increased secretion of tissue and other proteins by damaged tubules. It was concluded that glomerular damage leads to glomerular proteinuria characterised by high proportions of albumin together with alpha 1b, alpha 2a and beta 2 globulins in lower but significantly diagnostic proportions.

Protein excretion patterns in cadmium-exposed individuals: high resolution electrophoresis.

High resolution electrophoresis was used to evaluate protein excretion patterns in six cadmium-exposed individuals with proteinuria, seven subjects with nonspecific nephropathies, and four normal unexposed subjects. The aim of the investigation was to determine (a) the type of excretion pattern (i.e., glomerular, tubular, or mixed) associated with cadmium exposure and (b) if the pattern in the cadmium-exposed individuals was distinctly different from subjects with nonspecific nephropathies. The electrophoretic results were consistent with the quantitative results for the cadmium-exposed workers. The results suggest that the pattern associated with cadmium exposure can be glomerular or mixed and that it is different (i.e., no gamma band) from nonspecific nephropathies.

[Automatic dosage of total urinary proteins with centrifugal analyser]. / Dosage automatique des protéines totales urinaires par analyse centrifuge.

Automatic assay of urinary total proteins on a centrifugal analyser. The assay of urinary proteins is achieved opacimetrically after precipitation by trichloracetic acid. The addition of tensio-active agents lates the initiation of precipitation, helps the formation of a microprecipitate thus hold in suspension. The technique is linear from 0 to 2,5 g/l and for every albumin/globulins ratio between pure albumin and pure globulin.

[Development and clinical evaluation of a new urinary protein test paper (URINE-TP)].

Because standard urinary protein test paper reacts more strongly to albumin than to globulin, we attempted to develop a test paper sensitive to globulin by applying chromatic analysis principles used in quantitative analysis of urinary protein. We developed a new urinary protein test paper (URINE-TP) containing a reagent of ACID VIOLET 17. It was found that URINE-TP reacts with the same strength to gamma-globulin as to albumin. This new test paper therefore allows accurate detection of Bence Jones protein, which was formerly detectable with standard test paper. There was a correlation between values for urinary protein obtained with URINE-TP and those obtained by quantitative analysis. As with standard test paper, URINE-TP indicates urinary protein values in the normal range by a lack of reaction, but on URINE-TP positive and negative results are more clearly distinguishable than on standard test paper. From these results, we conclude that URINE-TP enables detection of both gamma-globulin and albumin in the urine, and that is as sensitive as the analytical method in the screening of urinary proteins.

High-resolution gel electrophoresis and sodium dodecyl sulphate-agarose gel electrophoresis on urine samples for qualitative analysis of proteinuria in dogs.

The aims of the current study were to assess whether sodium dodecyl sulphate-agarose gel electrophoresis (SDS-AGE) and high-resolution electrophoresis (HRE) can identify dogs with a urinary protein-to-creatinine ratio (UPC ratio) >0.2 and whether HRE can provide preliminary information about the type of proteinuria, using SDS-AGE as a reference method. HRE and SDS-AGE were conducted on 87 urine samples classified according to the International Renal Interest Society as non-proteinuric (NP; UPC ratio: <0.20; 32/87), borderline proteinuric (BP; UPC ratio: 0.21-0.50; 15/87), or proteinuric (P; UPC ratio: >0.51; 40/87). SDS-AGE and HRE were positive in 14 out of 32 and 3 out of 32 NP samples and in 52 out of 55 and 40 out of 55 samples with a UPC ratio >0.20, respectively. The concordance between HRE or SDS and UPC ratio was comparable (κ = 0.59; κ = 0.55). However, specificity (90%) and positive likelihood ratio (7.76) were higher for HRE than for SDS-AGE (56% and 2.16) while sensitivity was lower (73% vs. 94%). The analysis of HRE results revealed that a percentage of albumin >41.4% and an albumin/α(1)-globulin ratio (alb/α(1) ratio) >1.46 can identify samples classified by SDS-AGE as affected by glomerular proteinuria while a percentage of α(1)-globulin >40.8% and an alb/α(1) ratio <0.84 can identify samples classified by SDS-AGE as affected by tubular proteinuria. In conclusion, both SDS-AGE and HRE could misclassify samples with a UPC ratio higher or lower than 0.20. Therefore, UPC ratio must always be determined before conducting these tests. The percentage of albumin and α(1)-globulin or the alb/α(1) ratio determined by HRE can provide preliminary information about the origin of proteinuria.