CONCENTRATION OF ACID PHOSPHATASE, RIBONUCLEASE, DESOXYRIBONUCLEASE, SS-Glucuronidase, AND CATHEPSIN IN "DROPLETS" ISOLATED FROM THE KIDNEY CELLS OF NORMAL RATS

Straus, Werner

doi:10.1083/jcb.2.5.513

Cited by 150 publications

(39 citation statements)

References 13 publications

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“…However, a few vacuoles in glomerular epithelial cells, and many vacuoles in proximal convoluted tubular cells, contained reaction product. This is somewhat suprising since lysosomes in tubular cells contain hydrolytic enzymes capable of inactivating peroxidase (25). It is possible that the observed persistence was due to the large dose of the tracer given (26).…”

Section: Resultsmentioning

confidence: 99%

Altered Functional Properties of the Renal Glomerulus in Autologous Immune Complex Nephritis

Schneeberger

Leber

Karnovsky

et al. 1974

The Journal of Experimental Medicine

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The present study was undertaken in order to investigate the nature of the glomerular damage in a form of experimental immune complex nephritis. The model of autologous immune complex nephritis (AIC) 1 (Heymann's nephritis) (1) was chosen for several reasons. First, the model bears a remarkable resemblance, in terms of ultrastructural and immunofluorescence features, to an important renal disease in man, membranous glomerulonephritis. Furthermore, the model is highly reproducible. In addition, the basic pathogenetic mechanism has been thoroughly documented (2-4). Briefly, the disease is produced in rats by immunization with homologous kidney preparations, which results in the formation of autoantibodies against a renal antigen, normally found in the brush border of proximal tubules. This antigen is presumed to enter the circulation in small amounts and combine with autoantibody to form complexes which deposit in glomeruli. 2 The complexes are detected as granular deposits of immunoglobulin and complement along the epithelial side of the glomerular basement membrane. The disease, like its human counterpart, is manifested by proteinuria, which is often heavy.Our approach was to use the enzymatic tracers horseradish peroxidase (HRP) (40,000 daltons) and catalase (240,000 daltons), and the electronopaque tracer ferritin (ca. 500,000 daltons) as probe molecules with which to assess the altered functional properties of the glomerular capillary wall. These ultrastructural tracers can be directly visualized, and their location within the glomerular capillary wall can be followed at sequential intervals after intravenous injection. Such protein tracers have been extensively used to identify * This study was supported by U.S. Public Health Service grants no. AM 16392 and AM 13132.1Abbreviations used in this paper: AIC, autologous immune complex; CFA, complete Freund's adjuvant; CL, capillary lumen; DAB, 3,3' diaminobenzidine-tetra-HC1; GBM, glomerniar basement membrane; HRP, horseradish peroxidase; US, urinary space.It is of interest that a related pathogenetic mechanism may operate in man. It has recently been reported that a tubular-derived antigen may be present in the glomerular immune deposits in some cases of human membranous glomerulonephritis (5).

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Section: Resultsmentioning

confidence: 99%

Altered Functional Properties of the Renal Glomerulus in Autologous Immune Complex Nephritis

Schneeberger

Leber

Karnovsky

et al. 1974

The Journal of Experimental Medicine

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“…Accordingly, in identifying these droplets as lysosomes we must rely in part on evidence obtained by others on cell fractions. Straus (31)(32)(33), for instance, found AcPase, /3-glucuronidase, ribonuclease, deoxyribonuclease, and cathepsin in partially purified fractions of "small-, .... intermediate-sized-," and "large droplets" isolated from normal rat kidney, and from the kidney of egg white-treated rats. He also followed changes in enzyme amounts and distribution patterns following egg white administration which apparently leads to the transformation of small droplets into FIGURE 18 Periphery of a large absorption droplet (ad), ~ hours after hemoglobin administration.…”

Section: Fmcre 17 Periphery Of An Absorption Droplet (Ad)mentioning

confidence: 99%

“…So far the evidence in case has been either indirect, i.e. derived from the study of cell fractions (31)(32)(33)37) or incomplete, i.e. based on the demonstration in situ of either an exogenous protein (38)(39)(40) or a lysosomal enzyme (8).…”

Section: F Miller and G E 1)alade Lytlc Activities In Protein Absomentioning

confidence: 99%

Lytic Activities in Renal Protein Absorption Droplets

Miller

Palade

1964

The Journal of Cell Biology

422

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The digestive cycle following reabsorption of hemoglobin by cells of the proximal convoluted tubules in mouse kidney and the uptake of ferritin by glomerular mesangial cells in the kidney of normal and nephrotic rats wereinvestigated by electron microscopical histochemical procedures.Mouse kidneys, sampled at closely spaced time points between 1 to 48 hours after intraperitoneal injection of hemoglobin, and rat (normal and nephrotic) kidneys, sampled at 30 minutes, 2 hours, and 48 hours after intravenous injection of ferritin, were fixed in glutaraldehyde, cut at 50/z on a freezing microtome, incubated for acid phosphatase and thiolacetate-esterase, and postfixed in OsO4. Satisfactory preservation of fine structure permitted the localization of the enzymatic reaction products on cell structures involved in uptake and digestion of exogenous proteins. The latter were identified either by their density (hemoglobin) or their molecular structure (ferritin).It was found that lysosomal enzymic activities and incorporated exogenous proteins occur together in the same membrane-bounded structures. In the cells of the proximal convolution, lytic activities become demonstrable within I hour after hemoglobin injection, appear first in apical vacuoles filled with hemoglobin, and persist in fully formed protein absorption droplets. At the end of the lytic cycle (~48 hours post injection), the cells have an increased population of polymorphic bodies which exhibit lytic activities. In smaller numbers, identical bodies occur in controls. It is concluded that they represent remnants of previous digestive events.The means by which the resorptive vacuoles acquire hydrolytic activities remain unknown. Fusion of newly formed vacuoles with residual bodies was not seen, and hemoglobin incorporation into such bodies was only occasionally encountered. Acid phosphatase activity was found sometimes in the Golgi complex, but enzyme transport from the complex to the resorbing vacuoles could not be established. Autolydc vacuoles containing mitochondria or mitochondrial remnants were frequently found during the early stages of hemoglobin resorption, but no definite conclusions about the mechanism involved in the segregation of endogenous material were obtained.In nephrotic rats ferritin was segregated in membrane-bounded bodies mainly in the mesangial cells and to a lesser extent in epithelial and endothelial cells. Most of these sites were marked by the reaction products of acid phosphatase and organophosphorus-resistant esterase and therefore identified as lysosomes connected with the digestion of incorporated exogenous proteins. 519on

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“…Granules of this type ("droplets"), ranging in size from 0.1 to 5 ~ diameter, have been isolated from the kidney cells of rats and were found to contain high concentrations of acid phosphatase, acid ribonuclease and deoxyribonuclease, cathepsin, and fl-glucuronidase (2,3). High concentrations of the same enzymes have been discovered in fractions isolated from rat liver by de Duve, Pressman, Gianetto, Wattiaux, and Appelmans (4) who called the granules in which these enzymes are concentrated "lysosomes.…”

Section: Identification Of Phagosomesmentioning

confidence: 99%

“…In other animals and age groups (19), the smaller size of the droplets may make their microscopic observation more difficult than in old rats. Since droplets have now been isolated from the kidneys of normal rats and have been characterized by their enzymatic properties (2,3), their differentiation from mitochondria should now be possible. The simple procedure of using 70 per cent alcohol and subsequent staining with basic fuchsin (section IV (a)) may also facilitate their differentiation.…”

Section: Identification Of Phagosomesmentioning

confidence: 99%

Rapid Cytochemical Identification of Phagosomes in Various Tissues of the Rat and their Differentiation from Mitochondria by the Peroxidase Method

Straus

1959

The Journal of Cell Biology

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101

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PLATES 86 TO 90(Received for publication, September 15, 1958) ABSTRACT 1. Granules characterized by their ability to segregate foreign proteins (phagosomes) were identified in the cells of many rat organs after intravenous administration of horseradish peroxidase, by using the conventional test with henzidine for the histochemical detection of peroxidase. The largest numbers of phagosomes were identified in kidney and liver. Considerable numbers were observed cytochemically in pancreas, prostate, epididymis, thymus, spleen, bone marrow, small intestine, heart, pituitary, and mouse mammary carcinoma.2. The variation in size of the phagosomes ranging from the limit of microscopic visibility up to 5 # diameter, previously described for kidney, was also observed to occur in many of the other organs. The average size of the phagosomes in different organs was also different, the phagosomes of the liver, for example being on the average smaller than those of the kidney, pancreas, and prostate.3. In squash preparations of kidney and liver, the phagosomes appeared often in curved rows following the course of the cell membranes of epithelial cells. In several other organs, they appeared aggregated in cells located in the vicinity of blood or ]ymphatic vessels or capillaries.4. After injection of peroxidase directly into the brain of a rabbit, a striking concentration of peroxidase was observed in phagosomes of endothelial cells of capillaries and vessels, surrounding the site of injection. It was suggested that this localization may offer an explanation for the so called blood-brain barrier.5. The cytochemical peroxidase method was applied to smears of isolated fractions of kidney and liver. Only the isolated phagosomes, but not the isolated nuclei, mitochondria, and microsomes, reacted with benzidine after administration of peroxidase. The contamination of conventionally prepared nuclear, mitochondrial, and microsomal fractions of kidney and liver with phagosomes of different sizes was observed. By correlating the cytochemical peroxidase test of smears of isolated fractions with the colorimctric determination of peroxidase, acid phosphatase, and cytochrome oxidase in the same fractions, the differentiation of the phagosomes from mitochondria and other cell granules was facilitated.6. The marked difference in the osmotic properties of phagosomes and mitochondria, detectable after treatment with 70 per cent alcohol, and the difference in their affinities towards basic fuchsin, made it possible to differentiate the phagosomes from the mitochondria. It was found by this simple procedure that kidney cells of normal rats contain a large number of phagosomes ranging in size from 0.5 to 3 #, whereas liver cells of normal rats contain relatively few phagosomes of this size but many smaller ones (0.2 to 0.5 # diameter). These increased in size after treatment of the rats with horseradish peroxidase.

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CONCENTRATION OF ACID PHOSPHATASE, RIBONUCLEASE, DESOXYRIBONUCLEASE, SS-Glucuronidase, AND CATHEPSIN IN "DROPLETS" ISOLATED FROM THE KIDNEY CELLS OF NORMAL RATS

Cited by 150 publications

References 13 publications

Altered Functional Properties of the Renal Glomerulus in Autologous Immune Complex Nephritis

Altered Functional Properties of the Renal Glomerulus in Autologous Immune Complex Nephritis

Lytic Activities in Renal Protein Absorption Droplets

Rapid Cytochemical Identification of Phagosomes in Various Tissues of the Rat and their Differentiation from Mitochondria by the Peroxidase Method

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