ELECTROPHORETIC EVIDENCE FOR ISOZYMES OF ARYLAMIDASE ("AMINOPEPTIDASE") IN THE RAT. I. RENAL, SERUM AND URINARY ENZYMES SPLITTING dl-ALANYL-2-NAPHTHYLAMIDE AND l-LEUCYL-2-NAPHTHYLAMIDE

1964 ◽  
Vol 12 (12) ◽  
pp. 869-874 ◽  
Author(s):  
BENITO MONIS
Keyword(s):  
Present Report ◽  
Molecular Forms ◽  
Supernatant Fraction ◽  
Blood Levels ◽  
Rat Tissues ◽  
Bilateral Nephrectomy ◽  
Rat Urine ◽  
Urinary Enzyme ◽  
Renal Ablation ◽  
Starting Point

Bilateral nephrectomy elicited no changes of serum blood levels of arylamidase assayed with two chromogenic substrates (l-leucyl-2-naphthylamide and dl-alanyl-2-naphthylamide) two days after the operation. Since rat urine contains similar enzymes it was postulated that the urinary enzyme is of renal origin and distinct from serum arylamidase. For this purpose, electrophoretic studies were undertaken. Starch block (for quantitative determinations) and starch gel for zymograms of arylamidase were used. It was shown that both procedures demonstrated the identity of urinary and renal arylamidase, which was distinct from the serum enzyme. The renal and urinary enzyme showed two distinct isozymes: one remaining at the origin and the other migrating somewhat less than the isozyme in serum. It is postulated that the isozyme remaining at the origin corresponds to a membrane-bound form, whereas the electrophoretically-mobile isozyme is found in the cellular supernatant fraction. The arylamidase from serum was represented by a single enzyme which migrated farthest towards the anode. Histochemical procedures for the demonstration of arylamidase activity in tissues at the light microscopic level permit the localization of enzyme(s) that can hydrolyze synthetic chromogenic naphthylamides containing l-leucyl and dl-alanyl groups (1). Rat kidneys have a high concentration of histochemically demonstrable arylamidase in the proximal convoluted tubules (2). Blood levels of arylamidase in the normal adult rat vary within a narrow range (3). The observation of arylamidase activity in rat urine raised several questions that led to the studies which are the basis of the present report. The relationship of serum and urinary arylamidase was the starting point of this investigation. It was speculated that if the urinary enzyme had its origin in the serum, bilateral nephrectomy should alter blood levels of the enzyme. If no such changes occurred, this would suggest that the urinary enzyme was released from the proximal convoluted tubule. In fact, total renal ablation led to no significant variation of serum arylamidase activity. It was therefore postulated that urinary arylamidase originated in the kidneys and that both are distinct from serum arylamidase. To test this hypothesis, zone electrophoretic studies were undertaken using two different supporting media. It will be shown that distinct molecular forms or isozymes of arylamidase can be separated from rat tissues and fluids by differential electrophoretic mobility. The electrophoretic identity of urinary and renal arylamidase, both of which are distinct from the serum arylamidase, is demonstrated in this study.

Lightness Perception Can Be Affected by Surface Curvature from Stereopsis

Perception ◽  
1994 ◽  
Vol 23 (8) ◽  
pp. 869-881 ◽  
Author(s):  
David Buckley ◽  
John P Frisby ◽  
Jonathan Freeman
Keyword(s):  
Present Report ◽  
Three Dimensional ◽  
Surface Shape ◽  
Sharp Change ◽  
Planar Surface ◽  
Lightness Perception ◽  
Starting Point ◽  
Integrated Interpretation ◽  
Shape Data ◽  
Surface Lightness

It is demonstrated that lightness perception can be affected by shape from stereopsis. The starting point was a report by Knill and Kersten that the perceived lightness of a monocularly viewed surface can be affected by outline-contour cues indicating that the surface is three-dimensional (3-D). In that study stimuli consisted of two equally sized abutting regions each having the same vertical linear-intensity ramp, so that the horizontal abutting boundary of the two patches created a sharp change in intensity. When this version of the Craik-O'Brien - Cornsweet stimulus has a rectangular outline, it exhibits the standard simultaneous contrast illusion: equivalent patches in the top and bottom regions appear to have different brightness despite having the same luminance. Knill and Kersten replicated this phenomenon with stimuli whose outline-contour cues were consistent with a flat (planar) surface. They found, however, that the illusion was greatly reduced in stimuli with outlines consistent with two abutting 3-D quarter cylinders, for which equivalent regions in the two halves appeared of similar lightness. Knill and Kersten interpreted this effect in terms of surface-lightness computations that took into account 3-D surface shape to achieve an integrated interpretation of the luminance and shape data. In the present report three experiments are described for which these earlier findings were taken as the starting point. In the first experiment the results were replicated by the use of a different methodology. In the second experiment it was shown that shape-from-stereo can produce similar effects on lightness perception to that caused by shape-from-contour. Real 3-D objects with curved surfaces, luminance profiles of the Knill and Kersten type, and carefully controlled outline-contour cues were used so that the objects appeared flat when viewed monocularly but curved in 3-D when seen binocularly. The third experiment was a control confirming that the stereo effect was not simply due to differences caused by monocular versus binocular viewing. It is concluded that the human visual system uses stereo cues, as well as outline-contour cues, in the interpretation of luminance data to recover surface lightness.

Trace Elements Burden in Geriatric Hemodialysis Patients: A Prospective Multicenter Collaborative Study

1997 ◽  
Vol 20 (10) ◽  
pp. 553-556 ◽  
Author(s):  
K.Y. Hung ◽  
C.Y. Ho ◽  
Y.M. Kuo ◽  
S.H. Lee ◽  
S.J. Hseih ◽  
...  
Keyword(s):  
Aging Process ◽  
Collaborative Study ◽  
Atomic Absorption Spectrophotometry ◽  
The Body ◽  
Blood Levels ◽  
Blood Concentrations ◽  
Normal Limits ◽  
Starting Point ◽  
Cellulose Membranes ◽  
The Impact

Introduction Hemodialysis therapy and the aging process bave been reported to interfere with the trace element (TE) status in the body. This multicenter collaborative study on blood levels of TE in geriatric hemodialysis (HD) patients was carried out with an aim to clarify the impact of the aging process and HD therapy on the TE status in such patients. Methods One hundred and fifty-one HD patients (65 male, 86 female), all aged over 60 (68.2±3.86 y), and 112 elderly controls (58 male, 54 female) with a mean age of 67.5±3.03 were enrolled. All patients underwent standard HD for at least 6 months. The artificial kidneys used were hollow-fiber dialyzers of cellulose membranes. Water used for HD was prepared by reverse osmosis. Blood was collected in the morning prior to dialysis and after fasting. Atomic absorption spectrophotometry was applied to measure blood levels of Pb, Cd and Hg as well as plasma levels of Cu, Zn and Al. Data are presented as mean ± SEM. Student's t-test and linear regression were applied for statistics. Results Our geriatric chronic HD patients showed a marked elevation of blood Al and decreased plasma Zn concentrations. The blood levels of Cu, Cd, Pb and Hg were within normal limits. The blood concentrations of Al, Cu and Pb were significantly higher in HD patients than in the normal elderly controls, whereas Cd and Hg were similar in both groups, and Zn were lower in HD patients. Plasma Zn value decreased as dialysis duration increased, however there was a reversely linear correlation between plasma Zn and age. Conclusions This data reported can be regarded as a starting point and may serve as an insight to further studies on TE imbalances in geriatric and chronic HD populations.

Variable Distribution of Three Molecular Forms of Peptide Histidine Isoleucinamide in Rat Tissues: Identification of the Large Molecular Form as Peptide Histidine Valine-(1–42)*

Endocrinology ◽  
1989 ◽  
Vol 125 (5) ◽  
pp. 2645-2655 ◽  
Author(s):  
ANNICK CAUVIN ◽  
ANDRÉ VANDERMEERS ◽  
MARIE-CLAIRE VANDERMEERS-PIRET ◽  
PATRICK ROBBERECHT ◽  
JEAN CHRISTOPHE
Keyword(s):  
Molecular Form ◽  
Molecular Forms ◽  
Rat Tissues ◽  
Variable Distribution

scholarly journals Proteins of the kidney microvillar membrane. Structural and immunochemical properties of rat endopeptidase-2 and its immunohistochemical localization in tissues of rat and mouse

1989 ◽  
Vol 264 (2) ◽  
pp. 335-346 ◽  
Author(s):  
K Barnes ◽  
J Ingram ◽  
A J Kenny
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Cross Reactivity ◽  
Immunohistochemical Localization ◽  
Mouse Kidney ◽  
Molecular Forms ◽  
Rat Tissues ◽  
Weak Staining ◽  
Mouse Tissues ◽  
Rabbit Polyclonal Antibody ◽  
Solubilized Form

The phosphoramidon-insensitive endopeptidase-2 in rat renal brush borders was investigated by immunochemical approaches with a rabbit polyclonal antibody raised to the purified enzyme released from the membrane by papain. An immunoaffinity column successfully purified the detergent-solubilized form of endopeptidase-2. This preparation had an apparent subunit Mr of 80,000, and did not show the two subunits, of Mr 80,000 and 74,000, consistently found in the papain-solubilized forms, indicating that the latter resulted from proteolysis by papain. SDS/polyacrylamide-gel electrophoresis of non-reduced samples of the enzyme revealed a band of Mr 220,000, confirming the presence of disulphide-bridged subunits. Treatment with endoglycosidases H and F generated smaller molecular forms, indicating that endopeptidase-2 contained about 30% asparagine-linked carbohydrate and that a few of these oligosaccharide chains were of the high-mannose type. Treatment with phosphatidylinositol-specific phospholipase indicated that the enzyme did not possess a glycolipid membrane anchor. A survey of rat tissues examined immunohistochemically and by immunoblotting revealed that only the kidney and intestinal tract expressed the antigen in significant amounts. Although some weak staining was seen in salivary glands and thyroid, other organs and tissues including brain and spinal cord were negative by both immunochemical techniques. In the kidney the antigen was confined to the lumen of the proximal tubule and was seen mainly in the population of juxtamedullary nephrons. In the gut, luminal staining was observed throughout its whole length, from duodenum to rectum. Excellent cross-reactivity of the antibody with Balb/c mouse tissues was observed. Immunohistochemistry of mouse kidney and gut revealed a distribution identical with that observed in the rat. Immunopurification of the detergent-solubilized mouse kidney antigen showed it to be a protein containing disulphide-linked subunits of Mr 90,000. It possessed endopeptidase-2-like activity, but was more efficient in hydrolysing azo-casein and less efficient in hydrolysing a model substrate than the rat enzyme. The close similarity between rat endopeptidase-2 and mouse meprin is further supported by these results.

scholarly journals cDNA sequence and heterologous expression of monomeric spinach pullulanase: multiple isomeric forms arise from the same polypeptide

1998 ◽  
Vol 331 (3) ◽  
pp. 937-945 ◽  
Author(s):  
Andreas RENZ ◽  
Stephanie SCHIKORA ◽  
Roland SCHMID ◽  
Jens KOSSMANN ◽  
Erwin BECK
Keyword(s):  
Amino Acids ◽  
Isoelectric Focusing ◽  
Tertiary Structure ◽  
Transit Peptide ◽  
Amino Acid Sequences ◽  
Molecular Forms ◽  
Calculated Molecular Mass ◽  
Post Translational Modifications ◽  
Starting Point ◽  
Spinach Leaf

The spinach pullulanase gene was cloned and sequenced using peptide sequences of the purified enzyme as a starting point and employing PCR techniques and cDNA library screening. Its open reading frame codes for a protein of 964 amino acids which represents a precursor of the pullulanase. The N-terminal transit peptide consists of 65 amino acids, and the mature protein, comprising 899 amino acids, has a calculated molecular mass of 99 kDa. Pullulanase is a member of the α-amylase family. In addition to a characteristic catalytic (β/α)8-barrel domain, it contains a domain, F, that is specific for branching and debranching enzymes. Pullulanase cDNA was expressed in Escherichia coli, and the purified protein was compared with the enzyme from spinach leaves. Identity of the two proteins was confirmed in terms of catalytic properties, N-terminal amino acid sequences and molecular masses. The pullulanase produced by E. coli showed the same microheterogeneity as the spinach leaf enzyme: it could be resolved into two substrate-induced forms by electrophoresis in amylopectin-containing polyacrylamide gels, and, in the absence of substrate, into several free forms (charge isomers) by isoelectric focusing or chromatofocusing. Rechromatofocusing of single free forms resulted in the originally observed pattern of molecular forms. However, heterogeneity of the protein disappeared on isoelectric focusing under completely denaturing conditions when only one protein band was observed. Post-translational modifications such as glycosylation and phosphorylation could be excluded as potential explanations for the protein heterogeneity. Therefore the microheterogeneity of spinach leaf pullulanase results from neither genetic variation nor post-translational modifications, but is a property of the single unmodified gene product. The different interconvertible forms of the pullulanase represent protein populations of different tertiary structure of the same polypeptide.

Tissue Distribution of Active Principles from Alpiniae Oxyphyllae Fructus Extract: An Experimental Study in Rats

2019 ◽  
Vol 15 (3) ◽  
pp. 286-293 ◽  
Author(s):  
Qi Wen ◽  
Hai-Long Li ◽  
Shi-Ying Mai ◽  
Yin-Feng Tan ◽  
Feng Chen
Keyword(s):  
Small Intestine ◽  
Oral Administration ◽  
Tissue Distribution ◽  
Large Intestine ◽  
Rat Kidney ◽  
Rat Tissues ◽  
Rat Stomach ◽  
Ether Anesthesia ◽  
Starting Point ◽  
Exposure Levels

Background: Alpiniae Oxyphyllae Fructus (Yizhi in Chinese) have been widely used as an herbal medicine for the treatment of diuresis, enuresis and diarrhea in China. Many studies have deciphered some potential underlying mechanisms for its anti-diarrheal effects. However, tissue distribution of Yizhi constituents is warranted because pharmacological receptors are frequently located in tissues. Moreover, it is also interesting to know about the potential correlation between behavior in drug distribution and the observed pharmacological response. The aim of this study is to investigate tissue distribution behaviors of Yizhi constituents after oral administration of Yizhi extract to rats, focusing on 10 active principles. Methods: Twenty four male Sprague Dawley rats were given orally the Yizhi extract and fourteen tissue samples were collected after being killed by bleeding from the abdominal aorta under ether anesthesia at different time-points. The resulting tissues were excised and homogenized. Based on our previous reports, liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) was used to quantify the target analytes, as well as phase II metabolites, in the various biosamples. Results: Almost all the targeted Yizhi active principles and some glucuronidated metabolites were qualitatively measured in rat stomach, small intestine, large intestine, as well as liver. Nootkatone, yakuchinone A and tectochrysin were observed in the rat brain. In other rat tissues, these analytes had lower exposure or could not be detected. Consistently, quantitative analysis revealed that the Yizhi active principles dominantly distributed into gastrointestinal tissues followed by liver, the overall exposure levels ranking as follows: stomach > small intestine > large intestine > liver. Tissue concentrationtime profiles of the test active principles in rat stomach, small intestine, and large intestine were bimodal with two concentration peaks occurring at 0.5 and 4h after oral administration, respectively. The exposure levels in rat kidney and bladder were quite low. Conclusion: The active principles of Yizhi were specially distributed into gastrointestinal tissues after oral administration of its ethanol extract to rats. The tissue distribution behaviors partly supported its anti-diarrheal effects from a pharmacokinetic opinion. This paper will be useful as the starting point for studying the pharmacological activities of this traditional herb.

scholarly journals CHANGES IN LIPID CONTENT OF SERUM AND OF LIVER FOLLOWING BILATERAL RENAL ABLATION OR URETERAL LIGATION

1943 ◽  
Vol 77 (5) ◽  
pp. 473-486 ◽  
Author(s):  
A. W. Winkler ◽  
S. H. Durlacher ◽  
H. E. Hoff ◽  
E. B. Man
Keyword(s):  
Fatty Acids ◽  
Lipid Content ◽  
Serum Lipids ◽  
Cholesterol Content ◽  
Progressive Increase ◽  
Bilateral Nephrectomy ◽  
Significant Elevation ◽  
Glucose Administration ◽  
Renal Ablation ◽  
Total Fatty Acids

1. In the dog and monkey bilateral nephrectomy or ureteral ligation results in a marked, progressive increase of total fatty acids, of free and esterified cholesterol, of phospholipid, and of free fat of serum. 2. No such changes follow unilateral nephrectomy, splenectomy, or fasting. 3. The increase after bilateral nephrectomy is not inhibited by glucose administration. 4. A marked increase of the phospholipid and a less significant elevation in cholesterol content of the liver accompanies this increase of serum lipids after bilateral nephrectomy.

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