首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
Cell cultures are increasingly used in the evaluation of chemically-induced nephrotoxicity. The utili of renal cell culture systems in toxicology would be improved, however, if better characterized and more specific markers of toxicity were available. High resolution proton nuclear magnetic resonance (1H NMR) spectroscopy is well suited to the study of toxicological events and has identified many novel markers of nephrotoxicity in vivo. In this study, 1H NMR spectroscopy has been used to characterize the biochemical composition of two renal cell lines of different nephronal origin, LLC-PK1 (pig proximal tubule) and Madin-Darby canine kidney (MDCK, distal tubule). The early biochemical responses of these cell lines to the model proximal tubular toxin S-(1,2dichlorovinyl)i-L-cysteine (DCVC) and the renal medullary toxin 2-chloroethanamine (CEA) have also been investigated. For each line, 500 MHz 1H NMR spectra of protein-free acetone extracts of cells and culture medium gave characteristic and reproducible profiles of low MW constituents, including amino and organic acids, glucose and soluble membrane precursors, such as choline and myo-inositol. Treatment-related changes in several low MW compounds not routinely measured in toxicological studies were revealed by NMR specboscopy before marked cytotoxicity was observed by phase contrast microscopy. For example, LLC-PK1 cells treated with 60 μM DCVC showed a marked decrease in intracellular choline levels within 3 h which suggests an effect on the balance of choline synthesis and utilization. Wrthin 9 h of treatment with DCVC there were decreases in intracellular acetate and alanine concentrations which may be indicative of a decrease in fatty acid oxidation and biglyceride metabolism accompanied by an increase in gluconeogenesis. In MDCK cells, 1 h post treatment with 5 mM CEA, intracellular glycine was decreased. This study indicates the potential power and applicability of 1H NMR spectroscopy for evaluating the biochemical and metabolic effects of toxins in cell culture systems and provides a novel approach to identifying new markers of tissue damage.  相似文献   

2.
Suspensions of proximal tubules were obtained by collagenase digestion of rat renal cortex followed by centrifugation on a percoll gradient. NAD content in tubules incubated at 37 degrees C was decreased by 40-60% compared with tubules incubated at 4 degrees C. This change occurred within 30 min and was maintained for up to 2 hr. Inhibitors of NAD hydrolysing enzymes prevented the depletion of cellular NAD at 37 degrees C. Acute changes in proximal tubule NAD content at 37 degrees C were not accompanied by changes in phosphate uptake by brush border membrane vesicles subsequently prepared from the same tubules. In contrast, incubation of tubules with parathyroid hormone (10(-6) M) produced the expected inhibition (20%) of brush border membrane transport of phosphate. One implication of these findings is that acute changes in total NAD content of proximal tubules at 37 degrees C may not influence the phosphate transport system in the renal brush border membrane. Other interpretations are discussed.  相似文献   

3.
In the intact kidney, renal proximal tubule cells accumulate p-aminohippurate (PAH) via a basolateral, probenecid- and sodium-sensitive transport system. Primary cultures of rabbit proximal tubule cells retain sodium-glucose co-transport in culture, but little is known about PAH transport in this system. Purified proximal tubule cells from a rabbit were grown in culture and assessed for PAH and alpha-methyl-D-glucoside uptake capacities as well as proximal tubule marker enzyme activities. Control PAH uptake on collagen-coated filters (20 +/- 3 pmol/mg protein.min; n = 8) was not significantly different from uptake in the presence of 1 mM probenecid (19 +/- 4 pmol/mg protein.min; n = 8). Uptake from the basal side of the cell was 3.9 +/- 0.7 times greater than that from the apical side. In multi-well plate studies, the uptake was significantly reduced by removing sodium from the medium and stimulated by coating the wells with collagen. Glutarate (10 mM) had no effect on the uptake of PAH. Other differentiated proximal tubule characteristics were retained in culture, including the ability to form domes and to transport glucose by a phlorizin-sensitive system. Phlorizin-sensitive 1 mM alpha-methyl-D-glucoside uptake was 134 +/- 42 pmol/mg protein.min (n = 7; P less than 0.02). The proximal tubule marker enzymes alkaline phosphatase and gamma-glutamyltranspeptidase, increased in activity in the cultures after confluence. It was concluded that whereas some differentiated properties were retained during primary culture of rabbit proximal tubule cells, the PAH transport system was selectively lost or modified from that present in the intact kidney.  相似文献   

4.
Acute renal failure was induced in male rats by the subcutaneous injectioon of 4 mg HgC12 per kg body weight. Enzyme activities of the proximal tubule were studied histochemically at six time intervals from 15 min to 24 h. The enzyme studied were alkaline phosphatase, 5'-nucleotidase, acid phosphatase, alpha-glycerophosphate dehydrogenase (NAD-independent), malic dehydrogenase, succinic dehydrogenase, latic dehydrogenase, glucose-6-phosphate dehydrogenase and glucose-6-phosphatase. Decreases in activity were observed for alkaline phosphatase and 5'-nucleotidase after 15 min. Acid phosphatase was decreased after 30 min. These three enzymes returned to control levels after 3 h, but malic dehydrogenase and alpha-glycerophosphate dehydrogenase were decreased at this time interval. Succinic dehydrogenase was first decreased after 6 h. The earliest morphological changes detectable by light microscopy were observed in pars recta tubules in the medullary rays after 6 h, a time when all enzymes studied showed widespread decreased activity throughout the proximal tubule. After 24 h, the pars convoluta appeared morphologically normal but the pars recta was necrotic and exhibited calcification, whereas enzyme activity was decreased (absent in some cases) in both pars convoluta and pars recta. These results support the hypothesis that Hg++, when given in a sublethal dose, is associated with early histochemical changes in the brush border of the proximal tubule, which may be related to early changes in sodium reabsorption and to the subsequent development of acute renal failure. The observation that changes in plasma membrane-associated enzymes occur early and prior to alterations in enzymes of mitochondria and the endoplasmic reticulum suggests that Hg++ interacts initially with the plasma membrane.  相似文献   

5.
Protection of tissues from oxygen toxicity is one of the major prerequisites to aerobic life. Since a wide variety of xenobiotics with prooxidant activity is excreted by the kidney, renal tubule cells should be protected from hazardous oxygen species. Because intravenously injected Cu/Zn-type superoxide dismutase (SOD) is rapidly excreted in the urine in its intact form, effective dismutation of superoxide radicals cannot be achieved in vivo by intravenously administered SOD. To scavenge superoxide radicals and inhibit their toxic effects in and around renal tubule cells, a hexamethylene-diamine (AH)-conjugated SOD (AH-SOD) was synthesized. When injected intravenously into the rat, (125)I-labeled AH-SOD disappeared from the circulation with a half-life of 3 min and accumulated in the kidney. After 30 min of administration, more than 80% of the radioactivity derived from AH-SOD was found to localize in the kidney without being excreted in the urine. Immunohistochemical examination revealed that, 60 min after administration, the major part of AH-SOD localized in renal proximal tubule cells. Kinetic analysis using right-side-out-oriented renal brush border vesicles revealed that AH-SOD bound to their membrane surface by some mechanism which was inhibited by AH but not by heparin and albumin. These results indicated that AH-SOD rapidly underwent renal glomerular filtration, bound to apical plasma membranes of proximal tubule cells, and localized in these cells for a fairly long time without being excreted in the urine. Thus, AH-SOD might permit studies on the role of superoxide radicals in and around renal proximal tubule cells.  相似文献   

6.
Hydrolysis of NAD by a nucleotide pyrophosphatase of renal membrane fractions has been reported previously. The aim of the present study was to localize this enzyme in the rat kidney. Nucleotide pyrophosphatase was assayed in glomeruli, in three parts of the proximal tubule and in four parts of the distal tubule dissected form freeze-dried sections. Nucleotide pyrophosphatase activity, expressed in mumol X min-1 X mg protein-1, ranged between 9.8 and 32.3 in the proximal tubular segments and between 1.1 and 2.7 in the distal tubular segments. It was 3.4 in the glomeruli. The enrichment of the activity during the purification of brush border vesicles was measured. A ten-fold higher specific activity was found in the brush border vesicles as compared to the renal cortical homogenates. Thus, most of the renal nucleotide pyrophosphatase appears to be localized in the luminal membrane of the proximal tubule. A permeabilization of the membrane did not increase the activity of brush border vesicles. This indicates that all catalytic sites are accessible at the outer surface of the membrane.  相似文献   

7.
Summary The reabsorption of ovalbumin double labelled with fluorescein isothiocyanate (FITC) in the kidneys of normal and castrated male and female rats was investigated using fluorometry and fluorescence microscopy. The animals received an intravenous injection of either 2 or 7 mg fluorescein-thiocarbamyl (FTC)-ovalbumin per kilogram bodyweight (bw) and were killed 4 or 8 min post-injection. Animals injected with unlabelled ovalbumin (7.0 mg/kg bw) served as controls. Fluorescence microscopy revealed that FTC-ovalbumin was reabsorbed exclusively in the renal proximal tubule, the highest level of reabsorption being observed in its first part. Four and eight minutes after the mjection, FTC-ovalbumin was only observed in apical reabsorption vacuoles, with lysosomes exhibiting no specific fluoreseence. Fluorometric determinations for the renal homogenate supernatant showed that the renal reabsorption of FTC-ovalbumin was up to 24% higher in normal females than in normal males. Castration resulted in a significant increase in renal reabsorption in male rats (up to 38%; significant), whereas a minor decrease was observed in castrated females. The renal uptake differences in normal and castrated animals are discussed in the light of the sex-hormone-dependent catabolism of lysosomal proteins in the renal proximal tubule of rats.Supported by the Deutsche Forschungsgemeinschaft (SFB 105)  相似文献   

8.
Summary The renal tubules of the paired pronephros in early larvae (ammocoetes) of two lamprey species, Lampetra fluviatilis and Petromyzon marinus, were studied by use of light-, scanning- and transmission electron microscopy. They consist of (1) a variable number of pronephric tubules (3 to 6), and (2) an excretory duct. By fine-structural criteria, the renal tubules can be divided into 6 segments. Each pronephric tubule is divided into (1) the nephrostome and (2) the proximal tubule, the excretory duct consisting of (3) a common proximal tubule followed by (4) a short intermediate segment, and then by a pronephric duct composed of (5) a cranial and (6) a caudal section. The epithelium of the nephrostome displays bundles of cilia. The cells of the proximal tubule possess a brush border, many endocytotic organelles and a system of canaliculi (tubular invaginations of the basolateral plasmalemma). The same characteristics are encountered in the epithelium of the common proximal tubule; however, the number of these specific organelles decreases along the course of this segment in a posterior direction. In the intermediate segment, the epithelium appears structurally nonspecialized. The cells of the cranial pronephric duct lack a brush border; they have an extensive system of canaliculi and numerous mitochondria. The caudal pronephric duct is lined by an epithelium composed of light and dark cells; the latter are filled with mitochondria and the former contain mucus granules beneath the luminal plasmalemma. The tubular segments found in the pronephros are the same in structure and sequence as in the lamprey opisthonephroi. However, only the nephrostomes and proximal tubules occur serially in the pronephros, while the common proximal tubule, the intermediate segment and the cranial pronephric duct form portions of a single excretory duct.This paper is dedicated to the memory of Professor W. Bargmann, long-time editor of Cell and Tissue Research, the author of a splendid review on the structure of the vertebrate kidney and a master of German scientific writing.  相似文献   

9.
L Sievers  P Kugler 《Histochemistry》1986,86(2):215-220
The reabsorption of ovalbumin double labelled with fluorescein isothiocyanate (FITC) in the kidneys of normal and castrated male and female rats was investigated using fluorometry and fluorescence microscopy. The animals received an intravenous injection of either 2 or 7 mg fluorescein-thiocarbamyl (FTC)-ovalbumin per kilogram bodyweight (bw) and were killed 4 or 8 min post-injection. Animals injected with unlabelled ovalbumin (7.0 mg/kg bw) served as controls. Fluorescence microscopy revealed that FTC-ovalbumin was reabsorbed exclusively in the renal proximal tubule, the highest level of reabsorption being observed in its first part. Four and eight minutes after the injection, FTC-ovalbumin was only observed in apical reabsorption vacuoles, with lysosomes exhibiting no specific fluorescence. Fluorometric determinations for the renal homogenate supernatant showed that the renal reabsorption of FTC-ovalbumin was up to 24% higher in normal females than in normal males. Castration resulted in a significant increase in renal reabsorption in male rats (up to 38%; significant), whereas a minor decrease was observed in castrated females. The renal uptake differences in normal and castrated animals are discussed in the light of the sex-hormone-dependent catabolism of lysosomal proteins in the renal proximal tubule of rats.  相似文献   

10.
The relationship between the covalent binding, uptake, and toxicity produced by pentachlorobutadienyl-L-cysteine (PCBC) was examined in rabbit renal proximal tubules (RPT), renal basolateral membrane vesicles, and isolated renal cortical mitochondria. Renal proximal tubules rapidly metabolized PCBC to a reactive intermediate that bound to tubular protein. Approximately 70–90% of PCBC found in the cell at any given time was bound to protein. PCBC initially uncoupled oxidative phosphorylation, followed by a 45% reduction of state 3 respiration and a 90% decrease in cellular adenosine triphosphate (ATP) levels. These events preceded cell death. Isolated mitochondria also metabolized PCBC to a reactive intermediate that bound to mitochondrial protein and initiated mitochondrial toxicity. These results show that. PCBC-induced mitochondrial dysfunction occurred as a result of mitochondrial bioactivation and that the mitochondrion is the critical subcellular target in PCBC toxicity. Aminooxyacetic acid (AOAA), an inhibitor of cysteine conjugate β-lyase, reduced the covalent binding of PCBC-equivalents to tubular protein by approximately 90% and decreased but did not prevent the toxic effects produced by PCBC on RPT respiration and cellular ATP levels. AOAA delayed but had no effect on the overall extent of cell death produced by PCBC. The protective effect of AOAA was independent of any effects on PCBC uptake. These results show that AOAA decreased but did not prevent the metabolism of PCBC by cysteine conjugate β-lyase. The partial inhibition of PCBC metabolism, and hence, PCBC-induced cell death by AOAA, may be related to limited concentrations of AOAA within the tubule cell or mitochondria.  相似文献   

11.
The relationship between the covalent binding, uptake, and toxicity produced by pentachlorobutadienyl-L-cysteine (PCBC) was examined in rabbit renal proximal tubules (RPT), renal basolateral membrane vesicles, and isolated renal cortical mitochondria. Renal proximal tubules rapidly metabolized PCBC to a reactive intermediate that bound to tubular protein. Approximately 70-90% of PCBC found in the cell at any given time was bound to protein. PCBC initially uncoupled oxidative phosphorylation, followed by a 45% reduction of state 3 respiration and a 90% decrease in cellular adenosine triphosphate (ATP) levels. These events preceded cell death. Isolated mitochondria also metabolized PCBC to a reactive intermediate that bound to mitochondrial protein and initiated mitochondrial toxicity. These results show that PCBC-induced mitochondrial dysfunction occurred as a result of mitochondrial bioactivation and that the mitochondrion is the critical subcellular target in PCBC toxicity. Aminooxyacetic acid (AOAA), an inhibitor of cysteine conjugate beta-lyase, reduced the covalent binding of PCBC-equivalents to tubular protein by approximately 90% and decreased but did not prevent the toxic effects produced by PCBC on RPT respiration and cellular ATP levels. AOAA delayed but had no effect on the overall extent of cell death produced by PCBC. The protective effect of AOAA was independent of any effects on PCBC uptake. These results show that AOAA decreased but did not prevent the metabolism of PCBC by cysteine conjugate beta-lyase. The partial inhibition of PCBC metabolism, and hence, PCBC-induced cell death by AOAA, may be related to limited concentrations of AOAA within the tubule cell or mitochondria.  相似文献   

12.
Primary cultures of renal rabbit proximal tubule cells were initiated from a pure suspension of proximal tubule fragments. Proximal tubule cells were grown in a hormone-supplemented, serum-free medium containing low concentrations of antibiotics. Confluent monolayers exhibited multicellular dome formation, indicating the presence of transepithelial solute and water transport. Ultrastructural examination revealed a monolayer of polarized epithelial cells with tight junctions and sparse membraneous microvilli facing the culture medium. Time course biochemical characterization was performed using a palette of 12 enzymes, representative of important metabolic functions or pathways. Brush-border-associated enzymes (gamma-glutamyl transpeptidase and alanine aminopeptidase) were moderately reduced throughout the culture whereas alkaline phosphatase was markedly decreased at confluency. Mitochondrial and lysosomal marker enzymes were well preserved over the culture period. Glutathione-S-transferase activity remained stable during the 16-day culture period investigated. Glycolysis enzyme activities (lactate dehydrogenase and hexokinase) were enhanced, as a function of culture age. Na(+)-K(+)-ATPase activity rise was concomitant with the increase of glycolysis marker enzymes. In contrast, the gluconeogenesis marker enzyme, glucose-6-phosphatase, fell dramatically to reach a low level equivalent to 4% of the activity measured in isolated proximal tubules. Primary cultures exhibited several differentiated functions of the proximal tubule cell: (a) PTH alone was able to induce a significant stimulation of adenylate cyclase activity, unlike isoproterenol, thyrocalcitonin, and arginine vasopressin, and (b) sodium-dependent alpha-methylglucoside (AMG) transport was detected. This AMG uptake was selectively inhibited by phlorizin (5 X 10(-3) M), which is a competitive inhibitor of glucose uptake at the apical membrane. Complete characterization made it possible to investigate hitherto unexplored aspects of in vitro cultured proximal tubule cells. This primary culture model could provide a useful and reliable tool to investigate in vitro renal proximal tubule function, under normal conditions or after a drug-induced toxicity.  相似文献   

13.
M Shima  Y Seino  S Torikai  M Imai 《Life sciences》1988,43(4):357-363
Using isolated glomeruli and nephron segments obtained from collagenase treated rabbit kidneys, we examined the in vitro degradation of alpha-human atrial natriuretic polypeptide (alpha-hANP). The ANP-degrading activity was measured by the amount of immunoreactive ANP remaining after incubation of about 50 fmoles alpha-hANP with each tissue preparation for 7.5 min. The sequence of degrading activity among isolated nephron segments was as follows: proximal straight tubule greater than proximal convoluted tubule greater than cortical collecting tubule greater than distal convoluted tubule greater than cortical thick ascending limb. A single glomerulus exhibited the degrading activity which was comparable to approximately 50% of the activity of 1 mm proximal convoluted tubule. Phosphoramidon, an inhibitor of endopeptidase, prevented the degradation of ANP in proximal convoluted tubule and glomerulus by 68% and 89%, respectively, but not in cortical thick ascending limb and cortical collecting tubule. From these results, we conclude that the degradation of ANP by endopeptidase occurs mainly in the proximal tubule and glomerulus.  相似文献   

14.
The organic anion p-aminohippurate (PAH) is specifically secreted by the renal proximal tubule. The possibility was examined that the probenecid sensitive PAH transport system (which is involved in this secretory process in renal proximal tubule cells in vivo) is retained in primary cultures of rabbit kidney proximal tubule cells. Significant 3H-PAH uptake into primary cultures of proximal tubule cells was observed. After 10 min, 150 pmole PAH/mg protein had accumulated intracellularly. Given an intracellular fluid volume of 10 microliter/mg protein, the intracellular PAH concentration was estimated to be 15 microM. The initial rate of PAH uptake (when 50 microM PAH was in the uptake buffer) was inhibited 50% by 2 mM probenecid. Intact monolayers also exhibited Na+-dependent alpha methyl-D-glucoside uptake (an apical marker). Basolateral membranes were purified from primary rabbit kidney proximal tubule cell cultures. Probenecid sensitive PAH uptake into the membrane vesicles derived from the primary cultures was observed. The rate of PAH uptake was equivalent to that obtained with vesicles obtained from the rabbit renal cortex. No significant Na+-dependent D-glucose uptake into the vesicles was observed, indicating that primarily basolateral membrane vesicles had indeed been obtained.  相似文献   

15.
Proximal and distal tubule suspensions were prepared from kidneys of Sprague-Dawley rats by an isolation procedure on a PercollR gradient. The marker enzymes alkaline phosphatase (brush border) and hexokinase (cytoplasmic) as well as p-aminohippurate transport capacity, gluconeogenic activity and electron microscopy were used to characterize the two kidney tubule suspensions. The results of this study indicate that cytochrome P-450 is localized to the proximal tubular cells and that the O-deethylation of 7- ethoxycoumarin was higher in the proximal than distal fraction. Both proximal and distal tubules showed glucuronidation and deacetylation capacities and a relatively equal distribution of non-protein sulfhydryls. These studies demonstrate metabolic heterogeneity of the nephron, the proximal tubule being the main site of renal xenobiotic metabolism. Understanding of metabolic heterogeneity of proximal and distal kidney tubules should provide important information regarding cell specific mechanisms of nephrotoxicity.  相似文献   

16.
The relationship between the covalent binding, uptake, and toxicity produced by S-(1,2-dichlorovinyl)-L-cysteine (DCVC) and S-(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC) was investigated in suspensions of rabbit renal proximal tubules (RPT). The DCVC and TFEC at concentrations of 25 μM produced a time-dependent (1–6 hours) loss of RPT viability. The TFEC was bio-transformed rapidly by β-lyase to a reactive metabolite which bound covalently to tubular protein. Approximately 63% of the TFEC-equivalents inside the cell were bound to protein. Covalent binding of TFEC-equivalents was associated with a 30% decrease in tubular basal and state 3 respiration, a sevenfold increase in lipid peroxidation, and, ultimately, cell death. The DCVC was biotransformed rapidly to a reactive metabolite which bound covalently to tubular protein. Approximately 90% of the DCVC-equivalents inside the cell were bound covalently to tubular protein. Following exposure to 25 μM DCVC, the binding of DCVC-equivalents was associated with a 17-fold increase in lipid peroxidation but, in contrast to TFEC, had no effect on tubular respiration. However, exposure of RPT to 100 μM DCVC resulted in a ninefold increase in the binding of DCVC- equivalents and a 30% decrease in tubular state 3 respiration. The β-lyase inhibitor aminooxyacetic acid (AOAA) blocked the covalent binding, mitochondrial dysfunction, lipid peroxidation, and cell death produced by TFEC. The AOAA decreased the covalent binding and the lipid peroxidation produced by DCVC by approximately 60–70% but had no effect on cell death. These results suggest that mitochondria! bioactivation of TFEC by β-lyase is critical for TFEC-induced mitochondrial dysfunction and the resulting cell death. These results also suggest that cytosolic bioactivation and binding, but not mitochondrial bioactivation and dysfunction, are important in the toxicity produced by DCVC to rabbit RPT. The lack of protection against DCVC toxicity by AOAA may be related to incomplete inhibition of DCVC metabolism or bioactivation of DCVC by pathways other than β-lyase.  相似文献   

17.
2 bromoethanamine hydrobromide (BEA) has been widely considered to be a target selective nephrotoxin that causes necrosis of the medulla in 24-48 h, but recent reports suggest that early cortical injury is also associated with this lesion. In order to assess the cortical effects of BEA (100 mg kg-1 bw single ip injection), several urinary markers of renal injury were evaluated over a 7 day period in male Wistar Albino rats. Hexachlorobutadiene (HCBD 150 mg kg-1 bw in peanut oil ip), a renal toxin which targets selectively for the proximal tubule, was used as a comparison. After BEA treatment, urinary levels of alanine aminopeptidase, gamma-glutamyl-transpeptidase, alkaline phosphatase and glucose increased transiently. Each of the proximal tubule marker enzymes peaked earlier following HCBD treatment and elevation of alanine aminopeptidase and gamma glutamyl transpeptidase was sustained for longer periods than for BEA. Following BEA treatment, lactate dehydrogenase rose prominently on day 1 followed by a return to control values on day 2 and a further rise on day 3 and remained high until the end of the study. BEA also increased the urinary excretion of total protein and albumin. After HCBD treatment, lactate dehydrogenase showed a transient elevation and glucose levels were slightly increased. Based on the present observations the changes induced by BEA administration on urinary markers of renal injury are different from those observed following HCBD treatment. These findings suggest that BEA toxicity also involves other parts of the kidney besides the papilla.  相似文献   

18.
Summary Hydrolysis of NAD by a nucleotide pyrophosphatase of renal membrane fractions has been reported previously. The aim of the present study was to localize this enzyme in the rat kidney. Nucleotide pyrophosphatase was assayed in glomeruli, in three parts of the proximal tubule and in four parts of the distal tubule dissected form freezedried sections. Nucleotide pyrophosphatase activity, expressed in mol·min–1·mg protein–1, ranged between 9.8 and 32.3 in the proximal tubular segments and between 1.1 and 2.7 in the distal tubular segments. It was 3.4 in the glomeruli. The enrichement of the activity during the purification of brush border vesicles was measured. A tenfold higher specific activity was found in the brush border vesicles as compared to the renal cortical homogenates. Thus, most of the renal nucleotide pyrophosphatase appears to be localized in the luminal membrane of the proximal tubule. A permeabilization of the membrane did not increase the activity of brush border vesicles. This indicates that all catalytic sites are accessible at the outer surface of the membrane.Supported by the Swiss National Foundation, grant nr. 3.813.084  相似文献   

19.
The alpha macroglobulins of rat serum.   总被引:5,自引:0,他引:5       下载免费PDF全文
Cortex of rat kidney was homogenized and fractions enriched in plasma membrane, endoplasmic reticulum or brush border were prepared by several techniques of differential centrifugation. The identity and homogeneity of the membrane fragments were investigated by assaying marker enzymes and by transmission and scanning electron microscopy. Kallikrein was present in both plasma-membrane- and endoplasmic-reticulum-enriched fractions isolated by two fractionation procedures. Kallikrein was highly concentrated in a plasma-membrane fraction but was absent from the brush-border membrane of proximal tubular cells. Cells of transplanted renal tumours of the rat, originating from the proximal tubule, had no kallikrein activity. Kininase activity, angiotensin I-converting enzyme (kininase II) and angiotensinase were found in a plasma-membrane-enriched fraction and especially in the fraction containing isolated brush border. It is suggested that after renal kallikrein is synthesized on endoplasmic reticulum, it is subsequently reoriented to a surface membrane for activation and release. Renal kallikrein may enter the tubular filtrate distal to the proximal tubules. The brush-border membrane of proximal tubule is the major site of inactivation of kinins and angiotensin II..  相似文献   

20.
Summary Morphological examination of kidney biopsies from patients with glomerulonephritis and hematuria has revealed the presence of erythrocytes within epithelial cells of the proximal tubule. This observation suggested that the proximal tubule might be capable of phagocytizing morphologically intact erythrocytes. To examine this possibility small quantities of heparinized autologous blood were injected into surface convolutions of proximal tubules of the rat kidney using standard micropuncture techniques. At time intervals ranging from 10 min to 120 h after injection, the kidneys were preserved for light and transmission electron microscopy by drip-fixation with a half-strength Karnovsky's glutaraldehyde-formaldehyde fixative.During the initial 6 h there was a flattening of the brush border and accumulation of electron-dense material representing hemoglobin in apical vacuoles and in lysosome-like structures. From 6 to 15 h after micropuncture, there was progressive loss of the brush border and the simultaneous formation of pseudopodia-like evaginations that extended from the apical plasma membrane and surrounded the individual erythrocytes. By 18 and 24 h, erythrocytes were observed in the proximal tubule cells. At later time intervals, edema, lymphocytic infiltration, and fibrosis were observed in the interstitium. In addition, crystalline structures were present in the lumen and the cells of both proximal and distal tubules. These findings suggest that in addition to their well-established ability to pinocytize hemoglobin and other proteins, the cells of the proximal tubule are capable of phagocytizing morphologically intact autologous erythrocytes. It is possible that phagocytosis by the proximal tubule cells may play a role in the disposal of erythrocytes from the tubular fluid in hematuric conditions.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号