M. Teschner

Renal proteinases and kidney hypertrophy in experimental diabetes

SummaryIDDM is associated with an increase in kidney size, which is due to cellular hypertrophy and progressive matrix accumulation within the glomerulus and throughout the tubulointerstitium. The present study addressed the potential role of cysteine and metalloproteinases in renal hypertrophy of short-term diabetes. Three weeks after induction of streptozotocin diabetes in rats, intraglomerular gelatinase activity (streptozotocin: 23±4 vs control: 44±3 mU/μg DNA) and cathepsin L + B activity (streptozotocin: 6.7±0.8 vs control: 9.3±0.7 U/μg DNA) were significantly decreased. Insulin treatment completely prevented the decline in glomerular proteinase activity (gelatinase: 37±6 mU/μg DNA; cathepsin L + B: 9.6±0.9 U/μg DNA). In isolated proximal tubules a similar pattern of enzyme activity could be observed. Three weeks of diabetes caused a significant decline in cathepsin L + B activity (streptozotocin: 28±2 vs control: 37±3 U/μg DNA). Insulin treatment again prevented the decline in these tubular proteinase activities. In parallel, kidney weight increased by 22% and glomerular protein/DNA ratio rose by 17% in untreated diabetic rats. Diabetic rats receiving insulin displayed a normal glomerular protein/DNA ratio and the kidney weight was increased by only 5%. These results show that renal hypertrophy of early diabetes is closely associated with a decline in both glomerular and tubular proteinase activity. Adequate insulin substitution prevented renal hypertrophy and the reduction in proteinase activity.

Activation of Leukocytes During Prolonged Physical Exercise

Previous studies have demonstrated an increment of circulating leukocytes and enhanced secretion of interleukin-1 by monocytes and macrophages during physical exercise. In the present study the effect of physical exertion on the activity of polymorphonuclear (PMN) leukocytes was investigated. Following both short-term (running 2,000 meters) and long-term (running 10,000 meters) exertion, phorbol-stimulated chemiluminescence, as an indicator of leukocytic oxygen radical formation and release of leukocytic elastase, as a parameter of degranulation, were determined immediately after running. The number of circulating leukocytes increased both after short-term (+21%) and long-term (+61%) exercise. There was a minor release of PMN elastase following short-term activity causing plasma levels of this compound to rise from 100 +/- 4.0 ng/ml to 116 +/- 12.3 ng/ml. Long-term exercise, on the other hand, induced a significant increase of elastase plasma levels from 107 +/- 9.1 ng/ml to 300 +/- 23.4 ng/ml, suggesting a remarkable release of this proteinase from neutrophils. Based on these findings we conclude that during physical exercise degranulation of PMN leukocytes occurs. Moreover, the fact that phorbol-stimulated chemiluminescence is decreased after running demonstrates an impaired capability of white cells to generate oxygen radicals.

Decreased Proteinase Activity in Isolated Glomeruli of Streptozotocin Diabetic Rats

The important initiating process of diabetic nephropathy is the glomerular accumulation of proteins which has been proposed to be due to a synergistic interaction of the disturbed intrarenal hemodynamic and the altered chemical composition of glomerular components. This study was performed to investigate whether there might be a reduced activity of glomerular proteinases in streptozotocin diabetic rats. Three weeks after the induction of diabetes by means of streptozotocin, we found a decreased proteinase activity in ultrasonically destroyed isolated glomeruli obtained by a differential sieving technique in comparison to nondiabetic controls. This held true at acidic (16.59 +/- 1.56 vs. 22.19 +/- 1.94 U/min/mg protein) as well as at neutral pH (7.82 +/- 0.55 vs. 10.67 +/- 0.81 U/min/mg protein) and could be confirmed when proteinase activity was related to the single glomerulus or DNA instead of protein. Treatment with insulin was effective in improving the degradative capacity at both pH levels. We suggest that decreased proteinase activity in diabetic glomeruli may, at least partly, explain the glomerular protein accumulation in diabetic nephropathy.

Intraglomerular proteinase activity in adriamycin-induced nephropathy.

Adriamycin (ADR)-induced nephropathy is characterized by focal and segmental glomerulosclerosis and is supposed to be an ideal model of chronic progressive renal disease. The aim of our study was to investigate whether there might be an altered activity of glomerular proteinases in ADR nephropathy, thereby aggravating glomerular protein accumulation as an important initiating hallmark of glomerulosclerosis. In fact, we could demonstrate significantly enhanced levels of intraglomerular protein and DNA content in the experimental animals at week 7, 12 and 22 after administration of ADR. When relating intraglomerular proteinase activity, which was measured in ultrasonically destroyed isolated glomeruli, obtained by differential sieving techniques, to the intraglomerular protein and DNA content, this enzyme activity was significantly reduced throughout the observation period. Based on these data, we suggest that this relatively decreased proteinase activity in glomeruli exposed to ADR might play a pathogenetic role in the development of glomerular hypertrophy, an important harbinger of glomerulosclerosis.

Role of alcohol in clinical nephrology.

SummaryDifferent nephrological derangements are observed in severe alcoholics. Until now the direct toxicity of ethanol is only shown in the fetal alcohol syndrome with various malformations of the genitourinary tract. In the adult the kidney is often involved in the development, maintenance and counterregulation of complex electrolyte disturbances like phosphate and potassium hypoglycemia etc. The alcohol associated retention of urate, induced by hyperlactatemia and/or increasedβ-hydroxybutyrate concentration is only rarely complicated by urate nephropathy. Alcohol intoxication (acute and chronic) predisposes to rhabdomyolysis with the risk of acute renal failure. There are some hints that chronic alcoholism with myopathy increases the vulnerability of the kidney for further toxic agents. In rats glycerol induced renal failure is enhanced by alcohol pretreatment. Finally, regular alcohol consumption raises the blood pressure, which per se is a risk factor for renal damage.ZusammenfassungNephrologisch wichtige Störungen des schwereren Alkoholismus manifestieren sich auf verschiedenen Ebenen. Eine direkte Schädigung der Nieren und abführenden Harnwege ist bislang ausschließlich bei alkoholischer Embryopathie nachgewiesen. Beim Erwachsenen dominieren unspezifische und komplexe Elektrolytstörungen mit Akzentuierung im Alkohol-Entzugssyndrom. Die Niere ist nicht selten primäre Ursache verschiedener Störungen, sie trägt ferner zur — oft inadäquaten — Kompensation extrarenal entstandener Stoffwechselstörungen (z.B. Phosphatmangel, Hypoglykämie) bei. Der alkoholassoziierten Uratretention, hervorgerufen durch Hyperlaktatämie oder Erhöhung derβ-Hydroxybuttersäure, kommt — wegen meist mäßiger Ausprägung — für die Entwicklung einer hyperurikämischen Nephropathie nur geringe Bedeutung zu. Alkoholexzeß (akut oder chronisch) prädisponiert zur Rhabdomyolyse mit konsekutivem Nierenversagen. Möglicherweise ist bei schwerem Alkoholismus und Myopathie die Vulnerabilität der Nieren für andere Noxen gesteigert. Bei der Ratte wird das Glyzerin-induzierte akute Nierenversagen durch Alkoholvorbehandlung verstärkt. Alkohol begünstigt ferner bei Normotonikern und Hypertonikern einen Blutdruckanstieg, der seinerseits das Risiko einer Nierenschädigung erhöht.

Intraglomerular fibronectin accumulation and degradation in obese Zucker rats

SummaryThe obese Zucker rat is a classic model of non-immune mediated spontaneous focal glomerulosclerosis. An important initiating hallmark of glomerulosclerosis in this model is mesangial matrix expansion. Fibronectin, a highly biologically active glycoprotein, is a normal constituent of mesangial extracellular matrix. Using a quantitative method based on enzyme immunoassay we assessed the intraglomerular fibronectin content and its degradation in obese Zucker rats and their lean littermates. In the obese Zucker rats the glomerular fibronectin content was significantly higher in comparison to the controls (88±6 vs 48±4 ng/103 glomeruli). Furthermore, proteinase activity against fibronectin was significantly reduced in the glomeruli of obese Zucker rats when compared to control animals (at pH 5.4: 186±6 U/mg protein vs 286±14 U/mg protein, at pH 7.4: 152±12 U/mg protein vs 193±12 U/mg protein). These data demonstrate that in obese Zucker rats there is a glomerular accumulation of fibronectin which we propose is at least partly due to diminished proteolytic digestion. Whether accumulation of intraglomerular fibronectin contributes to progressive glomerulosclerosis remains a matter of debate.

Effect of Metabolic Acidosis on Tubular Proteinase Activity

Metabolic acidosis is a well-known mediator of compensatory renal hypertrophy; however, the underlying mechanism is still poorly understood. The aim of the present study was to investigate whether metabolic acidosis can influence the proteolytic activity in the proximal tubule. Metabolic acidosis was induced in rats by 0.28 M NH4Cl which was mixed to drinking water. The development of metabolic acidosis was documented by a significant increase in urinary pH. After 11 days of 0.28 M NH4Cl treatment, the experimental animals developed mild proteinuria (9.52 +/- 0.99 versus 17.65 +/- 1.63 mg/day). The kidney weight increased significantly (1,653.56 +/- 27.84 versus 1,753.33 +/- 56.11 mg) and tubular proteinase activity, measured at pH 5.4, was markedly reduced (60.3 +/- 1.2 versus 52.2 +/- 2.4 U/mg protein, or 2,105.5 +/- 92.0 versus 1,631.0 +/- 97.2 mU/micrograms DNA). In summary, these results suggest that compensatory renal hypertrophy induced by metabolic acidosis might at least partly be due to the suppression of tubular proteinase activity.

Hypercatabolism in Acute Renal Failure - Mechanisms and Therapeutical Approaches

Despite intensive efforts in the nutritional treatment of hypercatabolic acute renal failure (ARF), its prognosis is still deleterious. The most important factor determining the outcome of ARF is the extent of catabolism which is caused by alterations of the hormonal milieu, enhanced proteolytic activity, the hemodialysis process and the patients underlying or superimposed illness like surgical or nonsurgical trauma, rhabdomyolysis and septicemia. Up to now, in randomly assigned studies, hyperalimentation with protein- and nonprotein-derived calories has failed significantly to improve the nutritional status of the patient, although maximal doses of amino acids have been administered. Since there is some evidence from animal experiments that high doses of amino acids might act nephrotoxic, perhaps rather than further increasing the quantity of amino acids, in the future antiproteolytically acting substances may help in the management of hypercatabolic ARF. Possibly the use of amino acid solutions, enriched with branched chain amino acids, might represent a new approach in the management of negative nitrogen balance in ARF.

Obese zucker rat: Potential role of intraglomerular proteolytic enzymes in the development of glomerulosclerosis

SummaryThe obese Zucker rat is a classic model of non-immune-mediated spontaneous focal glomerulosclerosis. An early morphological hallmark of glomerular damage in the obese Zucker rat is a mesangial expansion, which precedes and mediates the development of glomerular damage in these animals. This study was designed to investigate whether there might be a reduced activity of glomerular proteinases in kidneys of obese Zucker rats, thereby being involved in the pathogenesis of mesangial expansion, which is mainly due to protein overloading. In fact, we found a decreased proteinase activity in ultrasonically destroyed isolated glomeruli obtained by a differential sieving technique in obese Zucker rats compared with their lean littermates. This held true at acid as well as at neutral pH and could be confirmed when proteinase activity was related to DNA instead of protein. When investigating the glomerular cathepsin B content —this is a lysosomal enzyme with acid pH optimum, which is involved in the degradation of glomerular structural as well as filtered plasma proteins—we found a significantly increased level in the kidneys of obese Zucker rats. Hence, the intraglomerular proteinase activity is reduced in the face of enhanced glomerular content of at least lysosomal proteinases. The underlying reason for this depressed activity remains to be established. We propose that reduced activities of intraglomerular proteinases may be important in the pathogenesis of mesangial matrix expansion in obese Zucker rats, an important initiating hallmark of glomerulosclerosis in this model.

Lovastatin ameliorates depressed intraglomerular proteolytic activities in experimental nephrotic syndrome.

Lipid abnormalities have been implicated in the pathogenesis of glomerulosclerosis in experimental models of kidney disease. In previous studies it has been shown that Adriamycin-induced nephropathy is associated with reduced activities of glomerular proteinases. This observation led to the hypothesis that reduced proteolytic activities may be responsible for mesangial protein accumulation, which ultimately leads to global sclerosis of the glomerular tuft. The aim of the present study was to investigate whether lovastatin treatment, which prevents progressive glomerulosclerosis in experimental nephrotic syndrome, would also have an effect on glomerular proteinase activities. Adriamycin administration resulted in a persistent nephrotic syndrome with gross proteinuria (377±26 mg/24 h), hypoalbuminemia (2.1±0.12 vs. 2.8±0.02 g/dl), hypercholesterolemia (575±74 vs. 68±1.5 mg/dl) and elevated triglyceride levels (1,155±78 vs. 57±8 mg/dl). Glomerular azocaseinolytic activities both at pH 5.4 (−21%) and 7.4 (−37%) were significantly reduced. In contrast to human subjects, nephrotic rats that were treated with lovastatin displayed reduced triglyceride levels (767±134 mg/dl); their serum cholesterol, however, remained unchanged. In terms of glomerular proteolytic enzyme activities, the decline in azocaseinolysis at both pH values was, at least partly, prevented by lovastatin. On the basis of these data, it appears that the beneficial effect of lovastatin on the evolution of glomerulosclerosis in the nephrotic rat is associated with the conservation of glomerular proteolytic activities.