C. H. van Aswegen

The effect of essential fatty acids on growth and urokinase-type plasminogen activator production in human prostate DU-145 cells

Abstract Urokinase-type plasminogen activator (uPA) is an important protease enzyme in carcinogenesis, and is involved in both invasion and metastasis of cancer. Increased uPA activity and decreased essential fatty acid (EFA) levels have been reported in cancer. This phenomenon may be explained by the fact that certain EFAs, such as γ-linolenic acid (GLA) and eicosapentaenoic acid (EPA), inhibit uPA activity. The effect of EFA on human prostate DU-145 cell growth and uPA production is still unknown and was investigated in this study. Data obtained from the different unsaturated fatty acids showed that oleic acid (OA) and EPA enhanced DU-145 cell proliferation at 0.004 and 0.04 mM for up to 4 days. However, α-linolenic acid (ALA), linoleic acid (LA), GLA and arachidonic acid (AA) suppressed cell proliferation under the same conditions, possibly as a result of inhibition of DNA and protein synthesis as measured using labelled thymidine and glycine incorporation. In contrast to the cell proliferation, uPA production was inhibited by all the unsaturated fatty acids under investigation. Therefore, the absence of EFAs, as reported, may affect invasion and metastasis of cancer.

The relationship between total urinary testosterone and renal calculi.

SummaryIt is generally known that age and sex are risk factors of urolithiasis. Therefore the total urinary testosterone concentrations of persons with and without renal stones were investigated by means of radioimmunoassay. The total testosterone level of the first morning midstream urine was comparable with 24 h urine samples of 16 healthy persons (rs=0.9618). Investigation of the total urinary testosterone confirmed that the concentration is age dependent. A distinct decrease in total testosterone was observed in elderly persons. Therefore the total testosterone concentrations of the two groups, with and without stones, were studied within the same age interval (P=0.8292). The total testosterone level differed significantly for these two groups (P=0.0006). In general, the testosterone level of the kidney stone patients was lower than that of their healthy counterparts. In order to determine whether this variation in testosterone concentration would affect the urinary urokinase activity, a correlation study was undertaken. A positive correlation was found between the total urinary testosterone concentrations and the activity of urokinase (rs=0.7305). It therefore seems that the total urinary testosterone concentrations may play a role in the pathogenesis of the multifactorial disease, urolithiasis.

In vivo effects of urease-producing bacteria involved with the pathogenesis of infection-induced urolithiasis on renal urokinase and sialidase activity.

Many hypothese have been proposed for renal stone formation. It has been argued that with infection-induced renal stones the hydrolysis of urea by bacterial urease increases urinary pH, with consequent stone formation. Unfortunately, this theory is not applicable to the micro-organisms that do not produce urease (e.g. Escherichia coli). It has been recently reported that E. coli reduces the urinary urokinase activity of male rats, but does not influence the urinary sialidase activity. This study has now been expanded to the urease-producing bacteria Proteus mirabilis, Staphylococcus aureus, S. epidermidis, Pseudomonas aeruginosa and Micrococcus luteus. Subcutaneous injections with these bacteria were found to significantly (P<0.003) reduce the UK activity of extrarenally obstructed kidneys. The urease-producing mammalian skin bacterium, M. luteus, was, however, the exception (P=0.1079). In contrast to S. epidermidis, P. aeruginosa and M. luteus (P<0.0213), P. mirabilis and S. aureus had no effect on renal sialidase activity (P<0.4047). These results may explain why Proteus species are predominant in infection-induced renal stones. According to the urokinase-sialidase hypothesis, a decrease in urinary urokinase activity should increase the uromucoid levels, whilst no effect on the urinary sialidase activity should favour conversion of urinary uromucoid to mineralizable matrix. These conditions may lead to renal stone formation. An increase in urinary pH resulting from urease-producing micro-organisms will increase salt precipitation on the uromucoid. It is thus concluded that urease-producing bacteria may play a double role in renal stone formation.

Does urokinase play a role in renal stone formation

Renal stone formation is a complex multifactorial disease, and it is believed that the initial step in the pathogenesis of urolithiasis must be the precipitation of an organic matrix of mucoproteins followed by precipitation of minerals onto this matrix. An important factor in this process may be the activity and/or concentration of the urinary enzyme, urokinase, which would affect the level of urinary mucoproteins such as uromucoid. In support of this hypothesis, ELISA studies were conducted to investigate the urokinase concentrations in urine obtained from males (22-60 years) with and without renal stones. These results showed a significant decrease in urinary urokinase concentration of renal stone patients which, once again, underlines the possible involvement of urokinase in renal stone formation. Therefore, it seems logical to conclude that urokinase may play an integral role in this multifactorial disease.

The effect of calcium and magnesium ions on urinary urokinase and sialidase activity

SummaryThe effect of a promoter (calcium) and an inhibitior (magnesium) of urolithiasis was spectrophotometrically studied on urokinase (0.45 IU) and sialidase (5 mM). Although these mineral did not affect the sialidase activity, total inhibition of urokinase activity was observed with either 0.05 M calcium chloride or 0.1 M magnesium chloride. This observation might explain why calcium and magnesium respectively function as a promoter and an inhibitor of stone formation.

Sialic acid concentrations in the urine of men with and without renal stones

SummaryIt has been suggested that urinary sialidase may play a role in the formation of renal stones. The present study was therefore undertaken to compare spectrophotometrically the different types of sialic acid concentrations and sialidase activities in fresh first morning urine specimens of men (21–65 years) with (13) and without (9) calcium oxalate renal stones. Although the free urinary sialic acid concentrations of the two groups of men were statistically about the same (P=0.0614), the total (P=0.003) and bound (P=0.0012) urinary sialic acid concentrations differed significantly. Both the total and bound sialic acid concentrations were lower in the urine specimens of the stone patients than in their healthy counterparts. This decrease in urinary sialic acid concentrations was firstly thought to be the result of elevated breakdown enzymes of sialic acid, which would favour the production of pyruvate. However, spectrophotometric determinations of the endogenous pyruvate concentrations of the two types of urine specimens did not differ significantly (P=0.0708). Secondly, the decrease in total urinary total sialic acid concentration of stone patients, could be attributed to less sialic acid synthesis or less renal excretion. Therefore, the same experiments were repeated using serum of 13 patients and 9 healthy men. Conversely, the total (P=0.4425) and bound (P=0.2850) serum sialic acid concentrations were found to be similar in the two types of subjects. However, the free serum sialic acid concentration of stone patients was significantly lower than in the healthy subjects (P=0.0062). This phenomenon is also reflected in the average ratio for serum free: bound sialic acid in healthy and stone patients, 1:7.9 and 1:18.7 respectively (P=0.0009). The lower free serum sialic acid concentration may lead to lower renal excretions of sialic acid. This may explain the decrease in total urinary sialic acid concentration in stone patients. The lower bound urinary sialic acid concentrations in patients was also reflected in the urinary free: bound sialic acid ratio for healthy (1:2.3) and stone patients (1:1.3). The difference between these two groups of men was highly significant (P=0.0001). This phenomenon might be explained by the urinary sialidase activities, which was spectrophotometrically determined at 334 nm at 37°C of 11 patients with stones and 17 healthy men. The ages of both groups of men were the same (P=0.326). An increase in urinary sialidase activity was observed with the stone patients (P=0.00001) when compared to specimens of healthy men. This might explain the decrease in urinary bound sialic acid concentration of the stone group. It seems from these results that the urinary concentration of sialic acid and the activity of urinary sialidase, may play a role in the pathogenesis of the multifactorial disease, urolithiasis.

Renal calculi-urate as a urokinase inhibitor.

SummaryThe formation of renal calculi is one of the most widely studied urinary ailments. Spectrophotometric analysis of urinary inhibition on the urokinase/plasmin system revealed a significant difference between subjects with and without renal calculi (P<0.001). The percentage urokinase/plasmin inhibition in the two groups was 77.1% for those with, and 47.1% for those without renal calculi. Because of the significant (P<0.015) positive correlation (r=0.762) between the percentage inhibition of urokinase/plasmin and urinary urate concentration, the inhibition of urate on urokinase/plasmin and on plasmin and urokinase was investigated. The urokinase/plasmin system was inhibited up to 94.4% with 0.5 mM urate. Inhibition occurred with both low and high molecular weight urokinases. In order to determine which enzyme of the complex was inhibited, each was investigated independently and a 50% inhibition of urokinase activity was obtained with 4 mM urate. It was found that the urine of stone formers contained a higher concentration of urate than the urine of healthy subjects and that at the same time there was a decrease in urinary urokinase activity in these patients.

Cytokine levels affected by gamma-linolenic acid.

This study was undertaken to assess whether gamma-linolenic acid (GLA) in the form of evening primrose oil (EPO) could affect rat serum cytokines, interferon-gamma (IFN-gamma), monocyte chemotactic protein-1 (MCP-1) and tumour necrosis factor-alpha (TNF-alpha). The following diets were administered: control, glucan, Freunds adjuvant and glucan plus Freunds adjuvant with and without GLA. In the presence of GLA, the IFN-gamma and MCP-1 levels were significantly decreased in contrast to the control group of TNF-alpha, which was significantly stimulated. On account of interaction between diets and GLA, the remaining diet groups of TNF-alpha were either not affected or were inhibited in the presence of GLA. The observations indicate that GLA may modulate the level of serum IFN-gamma, MCP-1 and TNF-alpha, which may be a worthwhile line of treatment in certain human diseases.

The effect of gamma-linolenic acid and eicosapentaenoic acid on urokinase activity

Abstract Urokinase (UK) is an important protease enzyme in carcinogenesis, and is involved in the invasion and metastasis of cancer. Thus, regulation of UK activity is likely to be important in healthy cell metabolism. As it has been reported that a decrease in Δ6-desaturated essential fatty acid (EFA) metabolites occurs in malignant cells and that gamma-linolenic acid (GLA) and eicosapentaenoic acid (EPA) exert antimutagenic effects, the effects of GLA and EPA on UK activity have been investigated in this study. Both GLA (n-6) and EPA (n-3) acted as competitive inhibitors of UK with K i values of 120 and 96 μM respectively. No modification of plasmin activity occurred with either 1.4 × 10 −4 M GLA or EPA. These results could explain why malignant cells with decreased EFA concentrations show increased UK activity. The addition of EFAs to available therapeutic regimens may be worth considering in the treatment of cancer.

Pathogenesis of kidney stones

Abstract Although many risk factors and theories exist in the literature for urinary stone formation, a hypothesis is suggested for the pathogenesis of renal stones. According to the matrix theory, a protein such as uromucoid activates the initial crystallisation process by promoting the formation of calcium oxalate and calcium phosphate crystals as well as clumping in whole urine. We put forward a theory whereby one of the most important factors in the matrix theory would be the composition and concentration of the protein. In suport of this hypothesis, emphasis is placed on the activities of urokinase and sialidase.