Separation of lipid classes by thin-layer chromatography

Skipski, Vladimir P.; Smolowe, Anson F.; Sullivan, R.Clark; Barclay, Marion

doi:10.1016/0005-2760(65)90047-0

Cited by 299 publications

(90 citation statements)

References 18 publications

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“…The different lipid fractions were separated on pre-coated PLC plates according to Skipski et ul. [12,13]. For transmethylation of the fatty acids the method of Pohl t>t al.…”

Section: Methodsmentioning

confidence: 99%

Active Ca Transport of Sarcoplasmic Reticulum during Experimental Uremia. Changes in Kinetics and Lipid Composition

Heimberg¹,

Matthews²,

Ritz³

et al. 1976

Eur J Biochem

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The Ca-transport system of sarcoplasmic vesicles of rabbits is altered by experimental uremia. 1. The influx rate constant of the experimental membranes decrease with a resulting decrease of the calcium influx rate.2. The experimental membranes transport a smaller amount of Ca2+ per mol of ATP split than the controls, i.e. their transport ratio is decreased.3. The calcium permeability of the experimental membranes increases with a resulting decreased concentrating ability.4. The phosphatide content but not the cholesterol content of the experimental membranes dccreases with a consequent increase of the cholesterol/phosphatide ratio.5. The fatty acid pattern of total phosphatides of the experimental membranes changes. A relative decrease of palmitic acid and oleic acid occurs and a relative increase of stearic, arachidonic and higher unsaturated fatty acids.6. The altered lipid composition of the membranes does not change the temperature dependence of the kinetics, In recent years it was shown that function and membrane composition of the fragment sarcoplasmic reticulum are affected by different models of disease showed kinetic changes of the sarcoplasmic reticulum during experimental uremia. In the following study these changes are analyzed in detail. Alterations of the lipid-protein interactions of the membrane were found which might be relevant for the understanding of disturbances of active transport processes during uremia.The changed lipid-protein interactions, correlated with kinetic changes, represent a new model of these interactions within intact membranes of fragmented sarcoplasmic reticulum which until now was not available in vifro and thus might allow some more insight into the lipid-protein interactions of the membrane. MATERIALS AND METHODSRabbits (2 -2.5 kg) of either sex were made uremic by 516 nephrectomy (removal of the right kidney, ligation of pole arteries of the left kidney under nembutal anaesthesia in one session. Control animals were sham operated. After the operation the rabbits were kept on pair-feeding conditions and sacrified after 7 days. During this time the serum urea level rose from 41.2 k 5.8 mg% to above 80 m g x .The vesicles were prepared from the psoas muscle and the white muscles of the hind legs according to Nagai et al. [9]. Neither a higher degree of protein nor of mitochondria1 impurities of the pathological preparations could be detected by gel-electrophoresis (Fig. I), electron microscopy (courtesy of D r Agostini) or by means of sodium azide, 5 mM of which showed no depressive action on the ATPase activity in both preparations.The ATPase activity of the vesicles was recorded by a pH stat instrument (Radiometer, Copenhagen) which was standardized by parallel P, measurements according to Rockstein and Herron [lo]. The Catransport activity was measured by the millipore filtration technique. For special conditions of the different kinetic measurements see legends.The vesicle lipids were extracted with a 20-fold amount of chloroform/methanol (2/1) and washed according t...

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“…The different lipid fractions were separated on pre-coated PLC plates according to Skipski et ul. [12,13]. For transmethylation of the fatty acids the method of Pohl t>t al.…”

Section: Methodsmentioning

confidence: 99%

Active Ca Transport of Sarcoplasmic Reticulum during Experimental Uremia. Changes in Kinetics and Lipid Composition

Heimberg¹,

Matthews²,

Ritz³

et al. 1976

Eur J Biochem

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“…The lipids were resolved on silica gel H plates developed in chloroform/methanol/acetic acid/ water (25:15:4:2). The plates were developed in the same solvent system three times (50). Spots were identified by comparison to the migration patterns of known standards following staining with Dragendorf, ninhydrin, and sulfuric acid charring.…”

Section: Analysis Of Lipid Compositionmentioning

confidence: 99%

Chondrocytes Utilize a Cholesterol-Dependent Lipid Translocator To Externalize Phosphatidylserine

et al. 2006

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During endochondral ossification, growth plate chondrocytes release plasma membrane (PM) derived matrix vesicles (MV), which are the site of initial hydroxyapatite crystal formation. MV constituents which facilitate the mineralization process include the integral membrane ectoenzymes alkaline phosphatase (ALPase) and nucleotide pyrophosphatase phosphodiesterase (NPP1/PC-1), along with a phosphatidylserine-(PS-) rich membrane surface that binds annexins and calcium, resulting in enhanced calcium entry into MV. In this study, we determined that chick growth plate MV were highly enriched in membrane raft microdomains containing high levels of cholesterol, glycophosphatidylinositol-(GPI-) anchored ALPase, and phosphatidylserine (PS) localized to the external leaflet of the bilayer. To determine how such membrane microdomains arise during chondrocyte maturation, we explored the role of PM cholesterol-dependent lipid assemblies in regulating the activities of lipid translocators involved in the externalization of PS. We first isolated and determined the composition of detergent-resistant membranes (DRMs) from chondrocyte PM. DRMs isolated from chondrocyte PM were enhanced in ganglioside 1 (GM1) and cholesterol as well as GPI-anchored ALPase. Furthermore, these membrane domains were enriched in PS (localized to the external leaflet of the bilayer) and had significantly higher ALPase activity than non-cholesterol-enriched domains. To understand the role of cholesterol-dependent lipid assemblies in the externalization of PS, we measured the activities of two lipid transporters involved in PS externalization, aminophospholipid translocase (APLT) and phospholipid scramblase (PLSCR1), during maturation of a murine chondrocytic cell line, N1511. In this report, we provide the first evidence that maturing chondrocytes express PLSCR1 and have scramblase activity. We propose that redistribution of PS is dependent on an increase in phospholipid scramblase activity and a decrease in APLT activity. Lastly, we show that translocator activity is most likely to be modulated by membrane cholesterol levels through a membrane raft microdomain. † This work was supported by USPHS Grant DE13576. * Address correspondence to this author. . battagli@biochem.dental.upenn.edu. HHS Public Access Author manuscriptBiochemistry. Author manuscript; available in PMC 2016 January 29. Author Manuscript Author ManuscriptAuthor Manuscript Author ManuscriptVertebrate skeletal growth occurs largely through a highly regulated process known as endochondral ossification. During this process mesenchymal cells condense and differentiate into chondrocytes to form a cartilage template. Chondrocytes proliferate and then undergo a complex maturation process associated with an increase in cell size (hypertrophy). This hypertrophic stage is characterized by upregulation of the expression of genes encoding ALPase, 1 type X collagen, and matrix metalloproteinase-13, followed by mineralization of the extracellular matrix. The zones of proliferating, prehypertro...

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“…CPIB (sodium salt) was obtained from Nissan Kagaku (Tokyo, Japan). [1-14C] (Folch et al, 1957) and applied to thin layer chromatography (Skipski et al, 1965). The radioactivities in cholesterol and triglyceride fraction were counted.…”

Section: Chemicalsmentioning

confidence: 99%

Modification of intracellular glucose metabolism in human skin fibroblast preincubated with p‐chlorophenoxyisobutyrate.

Nishide

Shirai

Saitō

et al. 1987

Brit J Clinical Pharma

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1 The effect of p-chlorophenoxyisobutyrate (CPIB) on glucose metabolism human skin fibroblasts was examined.2 CPIB increased the incorporation of 2-deoxy-D-[U-14C]glucose into skin fibroblasts.3 CPIB decreased [14C]C02 production from D-[U-14C]glucose but did not affect pyruvate dehydrogenase activity. 4 CPIB reduced fatty acid oxidation activity and cholesterol synthesis but increased triglyceride synthesis. 5 These effects of CPIB were observed both in the presence and in the absence of insulin. 6 One possible mechanism of CPIB on reducing plasma glucose may be due to the increase of glucose incorporation into the cells and triglyceride synthesis in the cells.

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Separation of lipid classes by thin-layer chromatography

Cited by 299 publications

References 18 publications

Active Ca Transport of Sarcoplasmic Reticulum during Experimental Uremia. Changes in Kinetics and Lipid Composition

Active Ca Transport of Sarcoplasmic Reticulum during Experimental Uremia. Changes in Kinetics and Lipid Composition

Chondrocytes Utilize a Cholesterol-Dependent Lipid Translocator To Externalize Phosphatidylserine

Modification of intracellular glucose metabolism in human skin fibroblast preincubated with p‐chlorophenoxyisobutyrate.

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