Renal tissue injury and proliferative response after successive treatments with anticancer platinum derivatives and tobramycin

The administration of anticancer platinum derivatives such as cisplatin, or aminoglycoside antibiotics is frequently associated with tubular necrosis which can eventually lead to acute renal failure. Previously, we have shown that renal tissue injury induced by these drugs elicits a process of tissue repair involving the stimulation of cell proliferation. The present study was undertaken to examine the morphological alterations and the proliferative response resulting from tobramycin administration to animals previously challenged with the platinum derivatives cisplatin and carboplatin. Female Sprague-Dawley rats were treated i.p. with cisplatin (8 mg/kg delivered in four daily injections) or carboplatin (40 mg/kg given in one injection) and sacrificed 21 or 60 days after drug administration. Tobramycin was administered i.p. twice a day at a daily dose of 10 mg/kg over the ten days preceding sacrifice. At 1 h before sacrifice, each animal received i.p. 200 microCi of [3H] thymidine for the measurement of DNA synthesis and cell proliferation (determined by histoautoradiography). Successive treatments with cisplatin and tobramycin appeared to produce an increase in the severity of histopathological alterations such as tubular necrosis and cystic degeneration. Moreover, cisplatin pretreatment dramatically increased the severity of tobramycin-induced lysosomal phospholipidosis. Histopathological alterations were followed by an important proliferative response partly associated with tubular regeneration but also due to fibroblastic proliferation which led to peritubular fibrosis. Surprisingly, the additive effect of cisplatin and tobramycin on renal injury became particularly striking with increasing time intervals between treatments. In contrast, successive treatments with carboplatin and tobramycin did not cause significative changes of the degree of renal injury, compared with either drug given alone.(ABSTRACT TRUNCATED AT 250 WORDS)     

Renal tissue injury and proliferative response after successive treatments with anticancer platinum derivatives and tobramycin

The administration of anticancer platinum derivatives such as cisplatin, or aminoglycoside antibiotics is frequently associated with tubular necrosis which can eventually lead to acute renal failure. Previously, we have shown that renal tissue injury induced by these drugs elicits a process of tissue repair involving the stimulation of cell proliferation. The present study was undertaken to examine the morphological alterations and the proliferative response resulting from tobramycin administration to animals previously challenged with the platinum derivatives cisplatin and carboplatin. Female Sprague-Dawley rats were treated i.p. with cisplatin (8 mg/kg delivered in four daily injections) or carboplatin (40 mg/kg given in one injection) and sacrificed 21 or 60 days after drug administration. Tobramycin was administered i.p. twice a day at a daily dose of 10 mg/kg over the ten days preceding sacrifice. At 1 h before sacrifice, each animal received i.p. 200 μCi of [³H] thymidine for the measurement of DNA synthesis and cell proliferation (determined by histoautoradiography). Successive treatments with cisplatin and tobramycin appeared to produce an increase in the severity of histopathological alterations such as tubular necrosis and cystic degeneration. Moreover, cisplatin pretreatment dramatically increased the severity of tobramycin-induced lysosomal phospholipidosis. Histopathological alterations were followed by an important proliferative response partly associated with tubular regeneration but also due to fibroblastic proliferation which led to peritubular fibrosis. Surprisingly, the additive effect of cisplatin and tobramycin on renal injury became particularly striking with increasing time intervals between treatments. In contrast, successive treatments with carboplatin and tobramycin did not cause significative changes of the degree of renal injury, compared with either drug given alone. In these groups we observed a moderate increase in cell proliferation which decreased with increasing intervals of time between treatments. In conclusion, the administration of tobramycin, even 2 months after cisplatin exposure, may still be hazardous. In contrast, successive treatments with carboplatin and tobramycin only seems to involve some, albeit moderate, nephrotoxic risk when the time interval between treatments is short.     

Morphological alterations in distal and collecting tubules of the rat renal cortex after aminoglycoside administration at low doses

Ultrastructural alterations in the cortical, distal and collecting tubules have been examined in female Sprague-Dawley rats treated with various aminoglycosides in clinical use. Gentamicin, dibekacin (10 mg/kg X day), netilmicin, tobramycin (4 or 10 mg/kg X day) or amikacin (37.5 mg/kg X day) were administered intraperitoneally twice a day over different periods of time, extending from 4 to 14 days. The kidney cortex was examined after 4, 7, 10 or 14 days of aminoglycoside administration by light (semithin sections) and electron microscopy. After 7 or more days of treatment, lysosomes in collecting tubular cells (and to a lesser extent in distal tubular cells) contained concentric lamellar material (myeloid bodies), an ultrastructural alteration typical of drug-induced lysosomal phospholipidosis. Although this alteration appeared qualitatively similar to that observed in proximal tubular cells, it was less conspicuous and occurred later during treatment. In addition, distal tubular cells occasionally showed marked vacuolization and disruption of the basal cell architecture. The possible relationship between these alterations and the urine hypo-osmolality characteristic of aminoglycoside-induced renal dysfunction is discussed.     

Distribution and excretion of a phosphorothioate oligonucleotide in rats with experimentally induced renal injury

The effects of renal injury on the urinary excretion and tissue distribution of a 20-mer phosphorothioate oligonucleotide were investigated in male Sprague-Dawley rats. Renal injury was produced by treating the rats with either 5.0 mg/kg cisplatin or 2.5 mg/kg of a monoclonal antibody (mAb) directed toward Thy1.1. Controls received saline. Three days after cisplatin treatment or 2 days after anti- Thy1.1 treatment, the rats received 10 mg/kg ISIS 3521. Blood was collected at various times to assess the plasma concentrations of ISIS 3521, and rats were killed at various times from 6 to 48 hours after intravenous (i.v.) infusion of oligonucleotide to assess tissue concentrations by capillary gel electrophoresis (CGE). Cisplatin and anti-Thy1.1 antibody produced histologic and biochemical changes consistent with proximal tubular damage and glomerular damage, respectively. Urinary excretion of oligonucleotides was increased 2- to 4-fold of control; however, this amount accounted for only 1% to 2% of dose compared to 0.5% in controls. Proximal tubular damage reduced renal accumulations of ISIS 3521 and other oligonucleotide metabolites, but there were no obvious compensatory increases in concentrations in other organs except for a slight increase in spleen levels of total oligonucleotide. Glomerular damage was not associated with any change in oligonucleotide disposition. Immunohistochemical studies showed no evidence of alterations in the pattern of distribution within the injured kidney. The data suggest that acute renal dysfunction, either renal tubular or glomerular, does not markedly alter the urinary elimination and tissue deposition of a phosphorothioate oligonucleotide.     

Microdissection studies of the structural alterations induced in rat kidneys by experimental postischemic acute renal failure

A unique opportunity presented itself for a morphologic study of experimental unilateral acute renal failure (ARF) in male rats. The ARF had been induced in the rats by temporary occlusion (1h) of the left renal artery. Twenty-nine rats were divided into subsets as follows: 2-3 h, 24 h, 1 week, 2, 4, 8, and 12 weeks following release of occlusion. Microdissection showed a heterogeneous population of abnormally structured proximal tubules in which the regressive lesions of tubular necrosis were combined with the progressive reaction of repair. The lesions demonstrated are reminiscent of those which have been described in ARF in the human and in experimental animals. Many proximal tubules in the 2- to 3-hour subset presented 1-3 disruptive lesions (DLs) while greater numbers of proximal tubules from the 24-hour group presented 1-5 DLs. Many proximal tubules presented no DLs, but nearly all from the 24-hour subset (97-100%) displayed a squamate appearance which paralleled and was caused by acute tubular necrosis. At 1 week, a dilated pars recta was common, but by this time, the squamate pattern had disappeared. Many casts were present. At 2 weeks, many fewer casts were present in proximal tubules and none were seen at 4, 8 or 12 weeks. The nephrons, particularly the proximal tubules, presented a variety of structural alterations at 2, 4, 8 and 12 weeks. Changes of special interest include (1) the presence of swan-necks; (2) a distinctive squamate appearance of the proximal tubules in the animals killed at 24 h; (3) a spiral, curled appearance caused by differential hyperplasia in animals at 4, 8 and 12 weeks, and (4) a tendency for ischemic lesions to involve all layers of the renal cortex.     

Mercuric chloride-induced alterations in stress protein distribution in rat kidney

Mercuric chloride (HgCl2) induces acute renal failure associated to tubular impairment in experimental animals and humans. Stress proteins are a superfamily of proteins, comprising heat- shock proteins (HSP) and glucose-regulated proteins (GRP), enhanced or induced in the kidney in response to stress. They act as molecular chaperones that protect organelles and repair essential proteins which have been denatured during adverse conditions. The involvement of stress proteins in mercury-nephrotoxicity has not yet been well clarified. This study was undertaken to detect the tubular distribution of four stress proteins (HSP25, HSP60, GRP75, HSP72) in the rat kidney injected with HgCl2 and to quantify lysosomal and mitochondrial changes in straight proximal tubules, the main mercury target. Sprague-Dawley rats were administered i.p. with progressive sublethal doses of HgCl2 (0.25 mg/kg, 0.5 mg/kg, 1 mg/kg and 3.5 mg/kg) or saline (as controls) and sacrificed after 24 h. In dosages over 0.50 mg/kg, stress proteins increased and changed localization in a dose-dependent manner. HSP25 was focally expressed in altered proximal tubules at 1 mg/kg but in the macula densa it was at 3.5 mg/kg. HSP60 and GRP75 were intense in the nucleus and cytoplasm of proximal tubules but moderate in distal tubules. HSP72 was induced in distal tubules after low exposures but in proximal tubules it happened at the highest dose. Moreover, a significant increase in lysosomal and total mitochondria (normal and with broken cristae) area and density were progressively found after HgCl2 treatments. Stress proteins could represent sensitive biomarkers that strongly correlate with the degree of oxidative injury induced by HgCl2 in the rat proximal tubules.     

Expression of metallothionein in renal tubules of rats exposed to acute and endurance exercise

The induction of exercise-induced apoptosis in not actively involved in exercise organs, such as kidney could be a result of oxidative stress. Metallothionein (MT) exerts a protective effect in the cell against oxidative stress and apoptosis. We have previously demonstrated an increased incidence of apoptosis in distal tubular cells and collecting ducts in rat kidney after acute exercise. The present study was designed to test the hypothesis that MT may play a protective role in rat renal tubules against exercise-induced apoptosis after the acute exercise and regular training. Male Wistar rats were divided into control, acute exercised and 8-wk regularly trained groups. The kidneys were removed after a rest period of 6 h and 96 h. The ultrastructure of renal tubular cells was examined by electron microscopy. Apoptosis was detected in paraffin sections by the TUNEL technique. Expression of MT was examined by immunohistochemistry. The level of lipid peroxidation (thiobarbituric acid reactive substances - TBARS) was assayed in renal tissue homogenates. After acute exercise, the occurrence of apoptosis was restricted to distal tubules and collecting ducts of rat kidney, whereas the proximal tubules remained unaffected. The 8-wk training did not result in increased apoptosis in tubular cell. MT expression was confined exclusively to proximal tubules in all groups. However, it was significantly increased in acutely exercised animals, as compared to control and trained rats. After the 8-wk training, MT expression remained unaltered as compared to the control group. TBARS levels were significantly increased after acute exercise, while after regular training they remained unchanged. A significant correlation between TBARS level and MT expression was demonstrated. The findings could suggest a protective role of MT against oxidative stress and apoptosis in proximal tubular cells.     

Edaravone inhibits acute renal injury and cyst formation in cisplatin-treated rat kidney

Background: Although cis-diamminedichloroplatinum (II) (cisplatin) is an effective anticancer agent, its clinical use is highly limited predominantly due to its adverse effects on renal functions. The present work examined the therapeutic potential of edaravone, a free radical scavenger, for inhibiting cisplatin-induced renal injury.

Methods: Edaravone, 3-methyl-1-phenyl-pyrazolin-5-one, was administrated intravenously at a dose of 30 mg/kg of body weight to male Wistar rats (200-220 g). After 30 min, cisplatin was injected intraperitoneally at a dose of 5 mg/kg of body weight. At the indicated times after the treatment, functions and histological changes of the kidney were analyzed. To test the therapeutic potential of edaravone in chemotherapy, its effect on the anticancer action of cisplatin was examined in ascites cancer-bearing rats.

Results: We found that cisplatin rapidly impaired the respiratory function and DNA of mitochondria in renal proximal tubules, thereby inducing apoptosis of tubular epithelial cells within a few days and chronic renal dysfunction associated with multiple cysts one-year after the administration. Administration of edaravone inhibited the cisplatin-induced acute injury of mitochondria and their DNA and renal epithelial cell apoptosis as well as the occurrence of chronic renal dysfunction and multiple cyst formation. The anticancer effect of cisplatin remained unaffected by intravenous administrating of edaravone.

Conclusions: These results indicate that edaravone may have therapeutic potential for inhibiting the acute and chronic injury of the kidney induced by cisplatin.     

Rat renal carcinogenesis after chronic simultaneous exposure to lead acetate and N-nitrosodiethylamine

Chronic oral administration of lead acetate and/or N-nitrosodiethylamine to rats produces three different types of renal cell tumors composed of basophilic, chromophobic or oncocytic cells. The most frequent tumor, often visible macroscopically, is made up of basophilic cells and forms tubular, cystic, pseudo-papillary or solid structures; it may show considerable cellular atypia but does not metastasize or invade the surrounding parenchyma. Chromophobic and oncocytic tumors are rare and can only be detected with the microscope; they usually form cystic or solid structures. Basophilic and chromophobic tumors arise from specific segments of the proximal tubules, characteristic for each carcinogen: P2, P3C and P3M for lead acetate; P2 and P3C for N-nitrosodiethylamine. Karyomegalia in proximal tubule cells appears to be irrelevant in renal carcinogenesis. However, the appearance of basophilic and chromophobic cells in P2, P3C and P3M segments is considered to be an early change in tumor development. Oncocytic microadenomas originate from collecting ducts showing focal oncocytic transformation. Synergistic or inhibitory effects are not observed after chronic simultaneous administration of lead acetate and N-nitrosodiethylamine, although both carcinogens act in common on P2 and P3C proximal tubule segments.     

Anti-VEGF antibody treatment accelerates polycystic kidney disease

Polycystic kidney growth implies expansion of the vasculature, suggesting that vascular endothelial growth factor (VEGF)-dependent processes play a critical role and that VEGF is a putative therapeutic target. Whether an anti-VEGF antibody improves renal cystic disease has not been determined. We administrated 5 mg/kg B20.4.1, an anti-VEGF-A antibody, or vehicle intraperitoneally twice weekly to 4-wk-old male normal (+/+) and cystic (Cy/+) Han:SPRD rats for 6 wk. Renal function, urinary protein excretion, organ/body weight ratios, cyst volume, tubular epithelial cell (TEC) proliferation, renal VEGF, hypoxia-inducible factor (HIF)-1α and -2α expression, renal histology, and kidney hypoxia visualized by [(18)F]fluoromisonidazole positron emission tomography were assessed. The treated compared with untreated +/+ rats had lower TEC proliferation rates, whereas Cy/+ rats receiving B20.4.1 displayed an increased proximal TEC proliferation rate, causing enhanced cyst and kidney growth. The +/+ and Cy/+ rats receiving B20.4.1 had severe renal failure and extensive glomerular damage. Proteinuria, which was highest in anti-VEGF-treated Cy/+ and lowest in untreated normal littermates, was positively correlated with renal HIF-1α and negatively correlated with VEGF expression. The untreated Cy/+ vs. +/+ rats had higher overall [(18)F]fluoromisonidazole uptake. The +/+ rats receiving B20.4.1 vs. untreated had increased [(18)F]fluoromisonidazole uptake, whereas the uptake was unchanged among treated vs. untreated Cy/+ animals. In conclusion, B20.4.1 caused an exaggerated cystic response of the proximal tubules in cystic rats and severe kidney injury that was associated with low renal VEGF and high HIF-1α levels. Anti-VEGF drug therapy may therefore not be a treatment option for polycystic kidney disease.     

Development of resistance against hexachlorobutadiene in the proximal tubules of young male rat

Hexachlorobutadiene (HCBD) is a potent nephrotoxin in rodents that can cause degeneration, necrosis and regeneration in renal tubular epithelial cells. Its toxicity is due to its conjugation by glutathione (GSH) to form glutathione S-conjugate, by the enzyme glutathione S-transferase and finally to the related cysteine-conjugate. This metabolite is then actively taken up by kidney and cleared in the renal tubular epithelial cells, rich in beta-lyase, to a reactive thiol derivative that covalently binds to the macromolecules. In this study, different groups of 28-day male Wistar albino (W/A) rats were dosed daily with 25 mg/kg HCBD for 2, 3, 4 and 7 days; control group dosed with corn oil. Data showed that in the 2- and 3-day treated groups there was substantial necrosis to the straight portion of the proximal tubules (pars recta or S3 segment), rich in glutamine transaminase K (GTK/beta-lyase). In the 4-day treated group, the renal proximal tubules had regenerated and showed a basophilic appearance. In animals treated for 7 days, it was observed that the kidney appeared to have returned to normal and had become resistant to further doses of HCBD. To define the time for the kidney to regain susceptibility to HCBD, 18- and 25-day studies with both low (25 mg/kg) and high (100 mg/kg) doses of HCBD (following two initial doses of 25 mg/kg) were performed. In the 18-day study, histopathological examination of the kidneys in animals of this group and also animals in the 25-day study, which received two further doses of HCBD, showed that the severity of kidney damage is much less than in the 2-day treated animals, a clear indication that the tubular cells were still resistant to the low dose of HCBD. Concentration of blood urea nitrogen, as a marker of kidney damage, in these two groups also confirmed the results. In animals re-exposed to the high dose of HCBD, data showed that the susceptibility to HCBD was starting to return.     

Gentamicin nephrotoxicity. II. Physiological, biochemical and morphological effects of prolonged administration to rats.

The purpose of this investigation was to determine the morphological, physiological and biochemical effects of gentamicin upon the rat kidney following prolonged administration of the antibiotic. Sprague-Dawley and Fischer 344 strain rats were given 3, 10, 20 or 40 mg gentamicin per kg body weight per day for 28 days. Morphologic alterations were evaluated by light and electron microscopy. Functional parameters included glomerular filtration rate, PAH secretion, renal plasma flow, sodium reabsorption, potassium excretion, urine volume and protein, and serum urea nitrogen. Oxidative metabolism of mitochondrial fractions from renal cortical homogenates was evaluated by oxygen uptake and P:O ratios. The results indicate focal proximal tubular injury, decreased tubular maximum secretion of PAH, and altered oxidative metabolism at the higher dose levels of gentamicin. Neither elevations of serum urea nitrogen nor alterations in glomerular filtration rate, renal plasma flow, or sodium or potassium excretion were observed. Thus, it appears that high dose levels (40 mg per kg per day) alter the structure and function of some proximal tubular segments when administered over prolonged periods. The alterations appear reversible. Although nephro-toxicity is identified under these conditions in rats, extrapolation to human patients usually receiving much lower doses must be guarded.     

Renal function and arterial blood pressure alterations after exposure to acetaminophen with a potential role of Nigella sativa oil in adult male rats

Hepatic injury by acetaminophen (APAP) has been extensively studied, although the alterations of renal functions and arterial blood pressure (ABP) after APAP exposure are still uncertain, and the impact of Nigella sativa oil (NSO) in this case is poorly defined. Sixty adult male albino rats were involved in two sets of experiments. The first was exposed to a single high dose of APAP (2.5 g/kg) orally preceded by 4 ml NSO/kg orally, while the second received 750 mg APAP/kg/day orally for seven consecutive days and was pretreated with 2 ml NSO/kg/day. Proximal tubular injury was assessed by laboratory and histological studies, and arterial blood pressure was recorded in all animals. In both experiments, urinary α-glutathione S-transferase and neutral endopeptidase, and microproteinuria were dramatically increased early indicating glomerulus and proximal tubule dysfunction that was mediated by raising 8-isoprostanes. Concomitantly, urinary albumin, total protein, creatinine, urea, glomerular filtration rate, Na and K levels, plasma creatinine, and urea were all changed significantly after APAP administration. Currently, ABP increased significantly after APAP which was mostly mediated by renal impairment and increased both renin activity and aldosterone secretion. Pretreatment with NSO produced significant normalization of physiological parameters as well as suppression of structural changes. In conclusion, measurement of urinary biomarkers can be considered a powerful tool for early screening of renal injury and alteration of ABP after APAP treatment. Concomitant administration of NSO can counterbalance these detrimental effects.     

Effect of cisplatin on H+ transport by H+ -ATPase and Na+/H+ exchanger in rat renal brush-border membrane

The effect of the potent anticancer drug cisplatin, cis-diamminedichloroplatinum (II) (CDDP), on H+ -ATPase and Na+/H+ exchanger in rat renal brush-border membrane was examined. To measure H+ transport by vacuolar H+ -ATPase in renal brush-border membrane vesicles, we employed a detergent-dilution procedure, which can reorientate the catalytic domain of H+ -ATPase from an inward-facing configuration to outward-facing one. ATP-driven H+ pump activity decreased markedly in brush-border membrane prepared from rats two days after CDDP administration (5 mg/kg, i.p.). In addition, N-ethylmaleimide and bafilomycin A1 (inhibitors of vacuolar H+ -ATPase)-sensitive ATPase activity also decreased in these rats. The decrease in ATP-driven H+ pump activity was observed even at day 7 after the administration of CDDP. Suppression of ATP-driven H+ pump activity was also observed when brush-border membrane vesicles prepared from normal rats were pretreated with CDDP in vitro. In contrast with H+ -ATPase, the activity of Na+/H+ exchanger, which was determined by measuring acridine orange fluorescence quenching, was not affected by the administration of CDDP. These results provide new insights into CDDP-induced renal tubular dysfunctions, especially such as proximal tubular acidosis and proteinuria.     

Phagocytosis of bacteria by proximal tubular epithelium in experimental pyelonephritis

Acute pyelonephritis was induced in rats by temporary unilateral ureteric obstruction and the intravenous injection of Escherichia coli. Animals were sacrificed 48 h after infection and changes in renal cortical tubules due to the presence of bacteria were studied. Bacteria appeared and multiplied in the tubular lumina and proximal tubular epithelial cells endocytosed the microorganisms in large numbers. Coalescence of phagosomes with lysosomes resulted in the surrounding of engulfed bacteria with acid phosphatase. However, the lysosomal apparatus of the cells did not eliminate Escherichia coli since the bacteria multiplied within phagosomes and destroyed the normal cell architecture. The peritubular interstitial inflammatory infiltrate caused ischemia of tubules, enhancing bacterial damage to the proximal tubules. The cytoplasm of the injured tubular cells was sometimes detached from the basement membrane. Cells of the distal tubules and collecting ducts did not show significant endocytosis or bacterial tubular damage.     

Interleukin-6 plays a protective role in development of cisplatin-induced acute renal failure through upregulation of anti-oxidative stress factors

AimsCisplatin, a major chemotherapeutic agent, accumulates in proximal tubules of the kidneys and causes acute renal failure dose-dependently. We previously reported that cisplatin induced more severe renal dysfunction in interleukin-6 (IL-6) knockout (IL-6^?/?) mice than in wild-type (WT) mice. Expression of a pro-apoptotic protein was significantly increased with cisplatin in IL-6^?/? mice compared to that in WT mice. IL-6, locally expressed in renal tubular cells after cisplatin administration, prevents the development of renal dysfunction at an early stage. In the present study, we focused on downstream signals of IL-6 and oxidative stress induced by cisplatin in order to evaluate the protective role of IL-6 in the development of acute renal failure.Main methodsWT and IL-6^?/? mice were given either cisplatin (30 mg/kg) or saline intraperitoneally. Blood and kidney samples were collected at 24 h and 72 h after cisplatin administration. The changes in expression of 4-hydroxy-2-nonenal protein (4-HNE, oxidative stress marker) and cyclooxygenase-2 (cox-2), activities of superoxide dismutases and caspase-3, and phosphorylation of extracellular signal-regulated kinase (ERK) were examined.Key findingsCisplatin increased the expression of 4-HNE and cox-2, and phosphorylation of ERK in IL-6^?/? mice than in WT mice. On the other hand, activity of superoxide dismutase, an anti-oxidative enzyme, was significantly decreased in the kidney obtained from IL-6^?/? mice after cisplatin administration.SignificanceOur findings suggest that IL-6 plays a protective role in the development of cisplatin-induced acute renal failure through upregulation of anti-oxidative stress factors.     

Maleate effects on kidney peptidases and proteinuria of male and female rats. Histochemical and biochemical studies

The effects of maleate on membrane-bound and lysosomal peptidases were studied histochemically in the kidney and biochemically in the kidney and the urine of male and female rats 6 h after the administration of two different doses of sodium maleate (150 and 300 mg/kg body weight). Additionally, the proteinuria of experimental animals was electrophoretically analysed to detect maleate-induced alterations in the urinary protein composition. The histochemistry of the brush-border peptidases (aminopeptidase A, gamma-glutamyltransferase) showed dose-dependent maleate effects in the late pars convoluta and the pars recta of the proximal tubule (blurring of the brush-border enzyme reaction pattern). The female animals were more severely affected by both maleate doses. After maleate treatment, enzyme-activity measurements in the kidney homogenate supernatant and urine indicated dose-dependent structural destruction of the proximal tubule, especially of brush-border membranes, and revealed an increase in enzyme excretion. Both the maleate-induced enzyme excretion and proteinuria were more pronouncedly increased in females than in males. Electrophoretic analysis of urinary proteins revealed alterations in the urinary-protein composition after maleate treatment, which favoured the excretion of proteins with a molecular weight higher than 20,000 daltons. Again, sex-related differences in the maleate effects were demonstrated. The results indicate that maleate causes alterations in the brush-border membranes and, especially at higher doses, results in cellular destruction selectively in the late proximal tubule of rat kidneys. Selectivity was also encountered in the maleate effects on urinary-protein composition, suggesting that the tubular alterations lead to an inhibition of the reabsorption of mainly high-molecular-weight proteins. Although the nature of the effects was independent of sex, it appears that females are less well protected against tubular damage caused by maleate.     

Morphometric study of the ultrastructure of nephrons and collecting tubules in the kidney of rats with renal and spontaneous hypertension.

The convoluted proximal and straight distal tubules and the medullary collecting ducts in kidneys of rats with ischaemic renal hypertension and with genetic spontaneous hypertension were studied by means of electron microscopic morphometry. The volume of mitochondria, the area of their cristae, of the outer surface and of membranes of the intercellular labyrinth, and other ultrastructural characterisitcs were calculated. No significant differences were found in proximal tubules between experimental and control animals, although in the distal tubules in both experiments the coefficient characterizing the level of morphologic organization of mitochondria, which takes into account their basic morphometric parameters, was reduced in hypertensive animals as compared with the intact ones. The volume of mitochondria and the area of their cristae in collecting ducts, and also the area of membranes of the intercellular labyrinth were increased. Our results suggest that in hypertension the reabsorption of substances from the proximal tubules is essentially normal, that it is reduced at the beginning of the distal tubules but is intensified in the collecting ducts.     

Dopamine D4 receptor expression in rat kidney: evidence for pre- and postjunctional localization.

Dopamine D4 receptors mediate inhibition of vasopressin-dependent sodium reabsorption by dopamine in collecting tubules. At present, the distribution of D4 receptors in other renal districts remains an open issue. The renal distribution of D4 receptor was assessed in normally innervated and denervated male Sprague-Dawley rats by quantitative immunohistochemistry using an anti-dopamine D4 receptor rabbit polyclonal antibody. D4 receptor protein immunoreactivity was observed perivascularly in the adventitia and the adventitia-media border. The density of perivascular dopamine D4 receptor was higher in afferent and efferent arterioles than in other segments of the renal vascular tree. Renal denervation abolished perivascular dopamine D4 receptor protein immunoreactivity. In renal tubules, the epithelium of collecting tubules showed the highest dopamine D4 receptor protein immunoreactivity, followed by the epithelium of proximal and distal tubules. No dopamine D4 receptor protein immunoreactivity was observed in the epithelium of the loop of Henle. Denervation did not change dopamine D4 receptor protein immunoreactivity in renal tubules. These results indicate that rat kidney expresses dopamine D4 receptors located both prejunctionally and nonprejunctionally in collecting, proximal, and distal tubules. This suggests that the dopamine D4 receptor may be involved in the control of neurotransmitter release and in renal hemodynamic and tubule function.     

All-trans retinoic acid ameliorates inflammatory response mediated by TLR4/NF-κB during initiation of diabetic nephropathy

Diabetic nephropathy (DN) is the leading cause of renal failure worldwide and its complications have become a public health problem. Inflammation, oxidative stress and fibrosis play central roles in the progression of DN that lead to renal failure. Potential deleterious effect of inflammation in early evolution of DN is not fully disclosed. Therefore, it is relevant to explore therapies that might modulate this process in order to reduce DN progression. We explored the beneficial effect of all-trans retinoic acid (ATRA) in early inflammation in glomeruli, proximal and distal tubules in streptozotocin (STZ)-induced diabetes. ATRA was administered (1 mg/kg daily by gavage) on days 3 to 21 after STZ administration. It was found that 21 days after STZ injection, diabetic rats exhibited proteinuria, increased natriuresis and loss of body weight. Besides, diabetes induced an increase in interleukins [IL-1β, IL-1α, IL-16, IL-13, IL-2; tumor necrosis factor alpha (TNF-α)] and transforming growth factor-beta 1 (TGF-β1), chemokines (CCL2, CCL20, CXCL5 and CXCL7), adhesion molecules (ICAM-1 and L-selectin) and growth factors (GM-CSF, VEGF, PDGF) in glomeruli and proximal tubules, whereas ATRA treatment remarkably ameliorated these alterations. To further explore the mechanisms through which ATRA decreased inflammatory response, the NF-κB/p65 signaling mediated by TLR4 was studied. We found that ATRA administration attenuates the TLR4/NF-κB inflammatory signaling and prevents NF-κB nuclear translocation in glomeruli and proximal tubules.