Kasahara-variant alkaline phosphatase in a renal cell carcinoma.

Electrophoretically Kasahara-variant alkaline phosphatase we found in a renal cell carcinoma tissue. This enzyme electrophoresed more quickly than liver alkaline phosphatase but more slowly than Kasahara isoenzyme. Neuraminidase treatment of the enzyme caused retardation of electrophoretic mobility which was the same as that of neuraminidase-treated Kasahara isoenzyme. The enzymic properties of this variant enzyme such as inhibition by L-phenylalanine, L-homoarginine, L-leucine, EDTA and urea are consistent with those of Kasahara isoenzyme. On Ouchterlony double diffusion, the precipitin lines of Kasahara and Kasahara-variant enzymes produced by antibody to Kasahara isoenzyme fused completely. These facts may mean that Kasahara-variant isoenzyme is different from the Kasahara one in terminal sialic acid content.     

Placental form of membrane-bound neutral arylamidase found in renal cell carcinoma

A placental form of membrane-bound neutral arylamidase was found in the tissue of renal cell carcinoma. Membrane-bound neutral arylamidases from renal cell carcinoma, an intact part of the same kidney and placenta, had the same molecular weight (240 000) and were identical with respect to k_m value and effect of inhibition by chelators or amino acids. Membrane-bound neutral arylamidase from renal carcinoma had the same electrophoretic mobility and heat stability as placental membrane-bound neutral arylamidase, but differed from kidney membrane-bound neutral arylamidase with respect to electrophoretic mobility, heat stability and susceptibility to urea inactivation. Results suggest the carcinoplacental alterations in membrane-bound neutral arylamidase isoenzyme in renal cell carcinoma.     

Comparison of soluble and membrane-bound neutral arylamidases from renal cell carcinoma

Soluble and membrane-bound neutral arylamidases from renal cell carcinoma were partially purified and their properties were compared. Soluble neutral arylamidase was a heat labile and SH-dependent metalloenzyme. Membranebound neutral arylamidase was a rather heat stable metalloenzyme and was activated by Co²⁺ with changing K_M and V_max. K_M values for soluble and membrane-bound neutral arylamidases were different. L-Methionine (5 mM) did not affect the enzymic activity of soluble neutral arylamidase, but inhibited 82% of the enzymic activity of membrane-bound neutral arylamidase. Molecular weights of soluble and membrane-bound neutral arylamidases by Sephadex G-200 gel filtration were 140 000 and 240 000, respectively.

Soluble and membrane-bound neutral arylamidases from renal cell carcinoma appear to be distinct enzymes.     

Immunological characterization of human membrane-bound arylamidases from small intestine, lung, kidney, liver, placenta and renal cell carcinoma

Anti-placental and anti-kidney membrane-bound arylamidase antibodies were prepared in rabbits. Both antibodies showed similar titration curves against both the original antigens, and the membrane-bound arylamidases partially purified from small intestine, lung, liver and renal cell carcinoma. Double immunidiffusion tests showed that six membrane-bound arylamidases were immunochemically indistinguishable.     

Separation and quantification of urinary di- and polyamines by gas chromatography with electron capture detection

A sensitive and specific gas chromatographic assay procedure employing electron capture detection has been developed for the assay of free and total di- and polyamines in human urine. Urine samples, hydrolysed with hydrochloric acid where necessary for the measurement of total amine output, were evaporated to dryness and, after the residues had been taken up in water, purified successively on Porapak Q and Dowex 50 X2 columns. Following evaporation of eluate, pentafluoropropionyl derivatives were made and analysed gas chromatographically using temperature programming. Di- and polyamines can be measured accurately at the picomole level and normal urinary output values calculated using this method agree well with those noted by other workers.     

Plasma catecholamines and dopamine-beta-hydroxylase activity in chronic renal failure

Plasma and urinary catecholamines (CA) and plasma dopamine β-hydroxylase (DBH) activity were studied serially in 17 hypertensive patients with chronic renal failure (CRF) and in 15 age-matched patients with benign essential hypertension (EH). Resting levels of plasma epinephrine (E) plus norepinephrine (NE) in CRF patients were significantly greater than those in EH patients (P < 0.05), whereas plasma DBH activities in CRF patients tended to be lower than those in EH patients (P < 0.1). However, DBH activities were found to be similar for the two groups, when they were expressed in units per litre of blood instead of per litre of plasma. Urinary free E + NE and dopamine were significantly less in CRF than in EH, whereas no significant difference was noted in urinary excretion of conjugated E + NE and vanillylmandelic acid between the two groups. The ratio of conjugated E + NE/free E + NE and of vanillylmandelic acid/free E + NE were significantly greater in CRF than in EH patients (P < 0.05). Glomerular filtration rate correlated significantly with free E + NE, free and conjugated dopamine, and inversely with the ratio of conjugated E + NE/ free E + NE in the whole subjects. These findings suggest that raised plasma CA concentration associated with the relative enhancement of extraneuronal inactivations may be relevant to the retarded clearance of circulating CA rather than increased CA release in CRF patients. It is likely that many factors unrelated to sympathetic nerve discharge have a considerable influence on both plasma CA concentration and plasma DBH activity in CRF patients, making them unreliable for studying the role of sympathetic nerve activity in these patients.     

Determination of erythrocyte uroporphyrinogen I synthetase activity in chronic renal failure

To obtain some information on porphyrin metabolism in uraemic patients, the activity of erythrocyte uroporphyrinogen I synthetase was measured in patients with chronic renal failure. The results indicate a decreased enzymatic activity which is not due to urea interference, in the hemolysates of these patients.     

Study of the activity of lymphocyte adenosine deaminase in chronic renal failure

The activity of lymphocyte adenosine deaminase was studied in 28 undialysed uremic patients, 19 patients on maintenance hemodialysis and 32 normal subjects. A statistically significant decrease of the enzyme activity was found both for the undialysed and dialysed patients.     

Interaction of calcium ions with serum albumin in chronic renal failure.

In order to study the effect of the uremic environment on serum calcium-albumin interaction, ten adult subjects with chronic renal failure (creatinine clearances below 20 ml/min) and five adult normal subjects, had their serum ionic calcium (Orion SS-20) determined at three different serum pH values. At similar serum protein levels, the correlations between serum ionic calcium pH were: (a) normals: number of pairs = 15, r = -0.90, p less than 0.01, Ca2+ = 4.54-0.484 pH. (b) uremics: number of pairs=15, r=-0.97, p less than 0.01, Ca2+=4.02-0.429 pH. These regressions were similar as far as the slopes (Fslope=0.62, p greater than 0.05) but had different intercepts (Fintercepts=64.34, p less than 0.01). These results confirm that uremics, when compared to normal subjects, present an ionic hypocalcemia at any level of serum pH. They also show that serum calcium-albumin interaction is normal in uremia. In view of the above results, it can be suggested that, among several possible causes, alterations in the binding-affinity of calcium ions to serum albumin can be excluded as an additional factor of secondary hyperparathyroidism in uremia.     

The relationship between the renal clearance of creatinine and the apparent renal clearance of beta-2-microglobulin in patients with normal and impaired kidney function

The renal clearances of creatinine and beta 2-microglobulin of patients with either normal or impaired kidney function were measured. The renal clearance of beta 2-microglobulin depends on the urinary pH and must be considered as an apparent renal clearance because after tubular reabsorption the compound is metabolized in the kidney. Impaired kidney function reduces the percentage of tubular reabsorption of beta 2-microglobulin.     

Fat clearances and hyperlipidaemia in renal allograft recipients — the role of insulin resistance

The mechanism of hyperlipidaemia in renal allograft recipients was investigated in 19 patients randomly selected from a cohort of 54 patients with functioning renal allografts. Serum cholesterol, triglyceride and high-density-lipoprotein cholesterol concentrations as well as plasma immunoreactive insulin levels were measured in fasting blood samples. Intravenous fat tolerance tests were performed before and 15 min after heparin administration. Renal allograft recipients had reduced fractional clearance rates of Intralipid and a positive correlation was demonstrated between plasma immunoreactive insulin levels and serum triglyceride concentrations. Plasma immunoreactive insulins also correlated inversely with fractional clearance rates of Intralipid. It was concluded that both increased production and decreased removal of lipoproteins contribute to the hyperlipidaemia and that insulin resistance due to corticosteroids was the centre of the problem.     

Investigation of pregnancy-related changes in red cell 2,3-diphosphoglycerate.

Significant gestational changes in red cell 2,3-DPG were found. No significant correlation was found between these pregnancy-related changes and alterations in the parameters of iron metabolism or red cell enzymes during the same gestational period. Possible reasons for the high concentration found at 21--24 weeks gestation are discussed.     

Lysosomal enzyme activities in cultured lymphoid cell lines

Several lysosomal enzyme activities in cultured lymphoid cell lines were studied during 3 phases of cell culture; logarithmic growth phase, stationary phase and decline phase.Enzyme induction during cell growth was found in N-acetyl-hexosaminidase, β-galactosidase and α-l-fucosidase, but no induction in a-d-mannosidase, α-glucosidase and β-glucuronidase. The latter two enzymes were unchanged during all cell culture phases.A drop in α-l-fucosidase and α-d-mannosidase activity was found during the stationary and decline phases of cell culture.     

A quantitative ELISA for IgG in cell culture supernatants

The measurement of IgG, using a sandwich-type enzyme-linked immunosorbent assay (ELISA), is described. The assay is specific, sensitive, and simple to perform. Purification of the antibody, prior to attachment to the plastic solid phase, results in a more sensitive test. Optimum standardisation of the reagents and conditions is described and discussed. The assay is capable of detecting IgG to 2.5 X 10(-5) IU/ml (approximately equal to 3 ng/ml) and is useful for measuring IgG produced by cells in culture. The coefficient of variation within assays is 6.0% and between assays is 7.7%.     

Urinary cyclic AMP and post-traumatic acute renal failure.

Consecutive daily urinary excretion of cyclic AMP has been investigated in 16 patients with severe trauma or illness, five of whom developed acute renal failure (ARF). Fluctuations in the nucleotide excretion exceeded the range found in 20 healthy volunteers (1.26-14.74, mean 7.13+/-1.18 vs. 2.04-10.10, mean 5.07+/-2.21 micronmol/24 h). This resulted in a 41% increase of cAMP excretion in the group with normal renal function (P less than 0.003) with the highest individual increase of 87%. The excretion usually reached its peak by 24 h after trauma and its lowest value by the third day, (first day vs. third day; 7.82+/-4.23 vs. 3.96+/-2.58 micronmol/24 h, P less than 0.05 for a group of 11 patients), while creatinine clearance remained normal. In four patients with severe ARF, the mean urine volume was above control value but the cAMP excretion was reduced to 3.9 to 14.4% and in one patient with a mild ARF to 60.6%. Creatinine excretion of the group was less reduced than that of cAMP (41.2% vs. 19.6%, resp.). cAMP excretion declined proportionally with diminishing creatinine clearance. In the category of 33-65 ml/min it decreased by 33.4% to 3.39 micronmol+/-1.16 micronmol/24 h. cAMP/creatinine ratio proved to be a less sensitive indicator than cAMP/24 h. Daily output of cAMP and creatinine correlated highly with diuresis in ARF patients, controls (always P less than 0.001) and less in patients with normal renal function (P less than 0.02). Urinary cAMP appears to be a very sensitive and early indicator of the onset of ARF and subsequent recovery. This warrants its further study.     

The effect of surgery on the renal excretion of β2-microglobulin

Surgical trauma causes an increase in the renal excretion rate of β₂-microglobulin whilst creatinine excretion is not influenced.The increase in the renal excretion rate of β₂-microglobulin is probably the result of an increased release of β₂-microglobulin by the cells which exceeds a maximum in the active tubular reabsorption of the compound by the proximal tubule cell.The renal excretion of β₂-microglobulin is proportional to the relative clinical trauma score.     

4-nitrophenyl phosphatase activity of the red blood cell membrane in essential hypertension

In this paper we report the levels of the 4-nitrophenyl phosphate hydrolysing activity of the red blood cell membrane in 46 hypertensive patients as compared to 41 normal controls and eight secondary hypertensives. This activity has at least two components; one of them is dependent on the presence of magnesium and potassium ions, and more sensitive to sodium, ATP, heat and -SH blockers than the cation-independent activity. This component appears increased in membranes from essential hypertension patients, correlating to the clinical seriousness of the condition, while remaining at control level in the secondary hypertension patients. The cation-independent component of this activity does not differ significantly in any of the groups studied.     

Serum cholesterol and squalene in hepatocellular carcinoma.

In 15 patients with hepatocellular carcinoma, the serum squalene level is raised significantly in parallel with the serum cholesterol level, though significantly decreased in patients with cholestatic hepatobiliary disease. Serum levels of cholesterol and/or squalene, therefore, could be regarded as a laboratory index reflecting a peculiar sterol metabolism in hepatocellular carcinoma.