Effects of Heparin on the Lipoproteins in Hyperlipemia. An Electrophoretic Study of the Serum Alpha and Beta Lipoproteins After Their Separation by Fractionation of the Plasma Proteins or Ultracentrifugal Flotation 1

Herbst, F.; Lever, Walter F.; Lyons, Mary E.; Hurley, Nancy A.

doi:10.1172/jci103106

Cited by 33 publications

(11 citation statements)

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“…Paper electrophoresis was performed essentially according to the method of Grassmann and Hannig (8,9). After being heated at 110°for 10 minutes, the paper ing 2 per cent agar, for 17 hours at 180 V and 40 mA.…”

Section: Methodsmentioning

confidence: 99%

“…Paper electrophoresis was performed essentially according to the method of Grassmann and Hannig (8,9). After being heated at 110°for 10 minutes, the paper strips were stained with Amidoschwarz 1OB.2 The intensity of the color was read in an "Elphor" densitometer.2 The relative percentage of the separated proteins was determined planimetrically from the curve obtained by plotting the optical density as a function of distance on the paper strip.…”

Section: Methodsmentioning

confidence: 99%

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Characterization of the Proteins of Human Synovial Fluid in Certain Disease States 1

Schmid¹,

MacNair²

1956

J. Clin. Invest.

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In search of the similarities and differences between blood plasma and synovial fluid, on one hand, and normal and pathological joint fluid on the other, it appeared important to investigate systematically the protein components of joint fluid. Investigations of the synovial fluid proteins by electrophoresis have been carried out by several authors (1-4). The identification of these proteins has been initiated (1,5). Fractionation with ammonium sulfate to obtain the albuminglobulin ratio (6) and demonstration of definite properties of certain proteins by immunochemical methods and enzymatic reactions have been reported (1,5).This publication presents a study on the characterization of the proteins in pathological synovial fluids. MATERIALS AND METHODSThe collection of synovial fluid and plasma as well as the procedure for the fractionation of their proteins have been described in a previous publication (7). All fluids were drawn from knee joints of patients with rheumatoid or traumatic arthritis or neuroarthropathy, and were obtained through the courtesy of the clinical staff of the Arthritis Unit. Most of the fluids deposited a precipitate within 20 hours although acid-citrate-dextrose (ACD) solution had been added. Prior to fractionation the joint fluids were treated with hyaluronidase (7).The electrophoretic and ultracentrifugal analyses were carried out in a Perkin-Elmer electrophoresis apparatus and a Spinco Model E ultracentrifuge, respectively. Paper electrophoresis was performed essentially according to the method of Grassmann and Hannig (8,9). After being heated at 110°for 10 minutes, the paper ing 2 per cent agar, for 17 hours at 180 V and 40 mA. The arrow "Electroph." indicates the direction of the electrophoretic movement of the proteins, and the arrow "Endosmosis" indicates the direction of the electroendosmosis. After the electrophoresis was completed, the troughs between the agar sections were filled with horse serum against human serum. Therefore, the precipitin zones formed symmetrical patterns. The positions of albumin and 7-globulin were found by applying these proteins as references on adjoining sections. The precipitin zones of synovial fluid (a) appeared less intense than those of the serum because of its lower protein concentration. The line corresponding to albumin, however, was very distinct due to the relatively high albumin concentration in this joint fluid.Whatman No. 1 paper (24 cm. wide) soaked in the borate buffer were used as electric connection between the ends of the agar plate and the adjoining borate buffer reservoirs. The buffer chambers, in turn, were again connected to holes filled with glass wool with a second set of buffer chambers in which the electrodes were placed. The electrodes were made by winding 120 cm. of stainless steel wire (0.05 cm.) around a glass tube with diameter of 1.5 cm. To secure an even level of the buffer in all 4 compartments, the two inner reservoirs were connected with each other by a thin rubber tubing. After half an hour of temperature equilibra...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Characterization of the Proteins of Human Synovial Fluid in Certain Disease States 1

Schmid¹,

MacNair²

1956

J. Clin. Invest.

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“…It also seems highly probable that lipoproteins in pathological as well as some normal sera will not always appear in the groups indicated in the three dimensional diagram shown above. The yellow lipoprotein described by Hunter in certain normal sera (23) and the lipoproteins appearing post heparin (13,24) may be examples.…”

Section: Discussionmentioning

confidence: 94%

The Α2 LIPOPROTEINS OF HUMAN SERUM. CORRELATION OF ULTRACENTRIFUGAL AND ELECTROPHORETIC PROPERTIES

Kunkel¹,

Trautman²

1956

J. Clin. Invest.

149

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Two main groups of lipoproteins have been characterized in the serum of normal adults.These have been termed the a1 and the 8,/ lipoproteins on the basis of their electrophoretic mobility (1-5). The broad spectrum of low density lipoproteins that have been observed particularly in patients with arteriosclerosis by means of the analytical ultracentrifuge (6) have not been classified in terms of electrophoretic mobility because of the difficulties in characterization by classical electrophoretic procedures (7-9). Nevertheless, these lipoproteins have usually been considered as /3 lipoproteins.Electrophoresis on filter paper, obviating solution density problems and permitting lipid analyses of the more stable whole serum has revealed the two main "alpha" and "beta" lipoprotein components. Another fraction observed at the site of origin is considered to represent adsorbed lipoproteins (4, 5) particularly the chylomicrons (10).This fraction is not observed by zone electrophoresis in a starch supporting medium, but an additional lipoprotein peak in the a2 region has been found in the sera of fasting normal (4, 11) and arteriosclerotic (12) individuals. Relatively little information is available regarding this component except that it was shown to rise after meals, particularly fatty meals, as indicated by phospholipid determinations (11).The present report represents an attempt to define the a2 lipoproteins further in terms of both zone electrophoretic and density ultracentrifugalproperties. This fraction was examined in the sera of a limited group of fasting normal and arteriosclerotic individuals and was found to consist primarily of lipoproteins less dense than d2"1.018. METHODSPreparative ultracentrifugation of the human sera was carried out in accordance with the basic principles of differential density centrifugation of the lipoproteins as described by Lindgren, Elliott, and Gofman (13) and de Lalla and Gofman (14) with very minor modifications. The Spinco swinging bucket rotor SW39L was used in the Spinco Model E ultracentrifuge instead of the angle preparative rotors. Various low molecular weight solutes were used to effect density increments for isolation of low density fractions and initially layered solutions were routinely employed.The purposes of this study were met by a gross fractionation with no density gradient of solvent but rather one due to the distribution of the serum albumin after the centrifugation to effect sufficient compartmentization to quantitatively maintain the separation of the serum into top and bottom fractions. Very slow acceleration and deceleration were employed, approximately 1000 rpm per min. In the Model LH preparative machine the insertion of a drive current meter enabled acceleration and braking at a given constant current similar to the operation of the Model E. This required constant manual slow movement of the speed dial. In general the Model L machine proved less satisfactory for accurate work with the swinging bucket rotor because of disturbance just prior to stopping.In ...

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“…The total content of cholesterol and its esters does not change significantly, although the concentration of unesterified cholesterol rises in the lipoproteins of higher density [Lindgren et al, 1955]. If heparin is injected when the blood does not contain lipid particles of high triglyceride content there are no changes in the ultracentrifugal pattern [Lever, Herbst and Lyons, 1955]. These changes following injection of heparin can most easily be interpreted in terms of a lipolytic reaction analogous to that which occurs in vitro.…”

Section: The Properties Of Clearing Factormentioning

confidence: 99%

“…This does not seem likely, however. Herbst, Lever, Lyons and Hurley [1955] isolated, by chemical fractionation and by ultracentrifugal analysis, the a-and ,B-lipoproteins from lipaemic human plasma after heparin injection. Both lipoproteins showed an increased electrophoretic mobility, but there was no abnormally high concentration of lipid in the a-lipoprotein fraction.…”

Section: The Properties Of Clearing Factormentioning

confidence: 99%

Untitled

1957

Exp Physiol

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THE possibility that heparin or related chemical substances might be concerned in the transport of fat in the body was suggested by a chance observation made by Hahn in 1943. He noticed that when dogs with a gross alimentary lipwemia were transfused with blood containing heparin as the anticoagulant, samples of plasma withdrawn one to five minutes after the transfusion were completely clear. Hahn found that this clearing of alimentary lipaemia also occurred when animals were given intravenous injections of heparin; it did not occur when heparin was mixed with lipsemic plasma in vitro. Later, Anderson and Fawcett [1950] found that a clearing effect could be produced in vitro when plasma from an animal previously injected with heparin (heparinized plasma) was mixed with lipaemic plasma. These two phenomena, described as the heparin clearing reaction in vivo and in vitro have been confirmed in many animal species including birds and reptiles. The purpose of this review is to describe the investigation of these phenomena and to discuss their possible significance in fat transport. The reaction in vitro has received the most attention and will be discussed first. The events which occur in the lipaemic animal following the injection of heparin can then be considered in relation to the findings in vitro. THE HEPARIN CLEARING REACTION IN VITRO The Mechanism of ClearingThe turbidity of the plasma in alimentary lipawmia is due to the presence of large numbers of visible lipid particles of varying size. They include the chylomicra and a variety of low density lipoprotein complexes. These particles are composed primarily of triglyceride but also contain varying amounts of protein, phosphatide and cholesterol [Laurell, 1954;Robinson, 1955;Lindgren, Nichols and Freeman, 1955;Havel, Eder and Bragdon, 1955]. The experiments of Anderson and Fawcett [1950] showed that, following heparin injection, plasma contained a component, the so-called clearing factor, which had the property of changing these lipid complexes so that they no longer interfered with the transmission of light. This clearing property of heparinized plasma can be demonstrated in vitro in a number of ways: by mixing heparinized plasma with lipaemic plasma [Anderson and Fawcett, 1950], or by adding fatty chyle or isolated turbid lipoproteins to heparinized plasma [French, Robinson and Florey, 1953; Brown, Boyle 151

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Effects of Heparin on the Lipoproteins in Hyperlipemia. An Electrophoretic Study of the Serum Alpha and Beta Lipoproteins After Their Separation by Fractionation of the Plasma Proteins or Ultracentrifugal Flotation 1

Cited by 33 publications

References 18 publications

Characterization of the Proteins of Human Synovial Fluid in Certain Disease States 1

Characterization of the Proteins of Human Synovial Fluid in Certain Disease States 1

The Α2 LIPOPROTEINS OF HUMAN SERUM. CORRELATION OF ULTRACENTRIFUGAL AND ELECTROPHORETIC PROPERTIES

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