Synthesis and characterization of fluorescent neutral lipid analogs containing N-(7-nitro-2,1,3-benzoxadiazol-4-yl)-aminohexanoic acid

McIntyre, Jonathan C.; Watson, D. K.; Sleight, Richard G.

doi:10.1016/0009-3084(93)90002-k

Cited by 6 publications

(3 citation statements)

References 20 publications

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“…Concentration dependent self-quenching of the NBD group is quite well-characterized (Nichols & Pagano, 1981;McIntyre et al, 1993;Prieto et al, 1994) and constitutes the basis of a large number of assays in membrane biology, such as the phase separation of lipids (Hoekstra, 1982;Wiener et al, 1985), phospholipid transfer between mixed micelles (Nichols, 1988), and phospholipase A 2 activity (Meyuhs et al, 1992). However, what we observe in this paper is not nonspecific quenching of fluorescence.…”

Section: Discussionmentioning

confidence: 99%

Membrane Organization at Low Cholesterol Concentrations: A Study Using 7-Nitrobenz-2-oxa-1,3-diazol-4-yl-Labeled Cholesterol

1996

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Cholesterol is most often found distributed nonrandomly in the plane of the bilayer, giving rise to cholesterol-rich and -poor domains. Many of these domains are thought to be crucial for the maintenance of membrane structure and function. However, such well-characterized domains generally occur in the membranes that contain relatively large amounts of cholesterol. Cholesterol organization in membranes containing very low amounts of cholesterol has not been investigated extensively. Recent evidence from differential-scanning calorimetric studies suggests that cholesterol may not form uniform monodisperse solutions, as assumed earlier, in the membranes even at very low concentrations. Fluorescent cholesterol analogues, when chosen carefully, offer a powerful approach for studying the distribution and organization of cholesterol in membranes at low concentrations. In this paper, we have studied the organization of cholesterol in membranes at very low concentrations (up to 5 mol % of the total lipid) using a fluorescent cholesterol analogue (NBD-cholesterol) which is labeled with the 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) group at the flexible acyl chain, without any alteration in the structural features necessary for proper membrane incorporation. Our results show that NBD-cholesterol exhibits local organization even at very low concentrations. This is consistent with the recently suggested model of cholesterol organization in membranes at low concentrations, involving the formation of transbilayer, tail-to-tail dimers The fluid mosaic model for biological membranes defines the membrane as an oriented, two-dimensional, viscous solution of proteins and lipids in instantaneous thermodynamic equilibrium (Singer & Nicolson, 1972). Although the basic tenets of this model have stood the test of time, one of its implicit assumptions, that the lipid molecules merely provide the milieu for the membrane proteins and that they are distributed uniformly throughout the bilayer, has been found to be not quite true. In fact, current evidence points out that both transverse and lateral regionalization, which can be described in terms of micro-and macrodomains, are common features of many biological membranes (Curtain et al., 1988;Edidin, 1992;Tocanne, 1992;Glaser, 1993;Jacobson et al., 1995).Cholesterol is one of the membrane components that is found, more often than not, distributed nonrandomly in structural and kinetic domains or pools in both biological and model membranes (Yeagle, 1985;Gennis, 1989;Schroeder et al., 1991;Liscum & Underwood, 1995). The reason for this lies in the rather unusual chemical structure of cholesterol that can by no means be considered a representative component of the "bulk" membrane (see Figure 1). The unusual shape of cholesterol also causes a phase separation in membranes at relatively high concentrations, resulting in the formation of cholesterol-rich and -poor domains that are structurally and compositionally so distinct that they can be observed histochemically and isolated physical...

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Section: Discussionmentioning

confidence: 99%

Membrane Organization at Low Cholesterol Concentrations: A Study Using 7-Nitrobenz-2-oxa-1,3-diazol-4-yl-Labeled Cholesterol

1996

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“…The C6-NBD-DAG standard was produced by phospholipase C treatment of C6-NBD-PC. The C6-NBD-TG standard was synthesized from C6-NBD-DAG (18). After chromatography with CHCl 3 :CH 3 OH: 30% NH 4 OH (65:35:5, v/v) to 10 cm, plates were air dried for 30 min and then chromatographed to the top of the plate in CHCl 3 :CH 3 OH: CH 3 COOH (100:2.5:5, v/v).…”

Section: Materials-silicamentioning

confidence: 99%

Acylglycerol Recycling from Triacylglycerol to Phospholipid, Not Lipase Activity, Is Defective in Neutral Lipid Storage Disease Fibroblasts

Igal

Coleman

1996

Journal of Biological Chemistry

119

116

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We have examined the developmental expression of acetylcholinesterase (AChE) during the process of neuronal differentiation from a pluripotent stem cell. P19 embryonic carcinoma cells form embryoid bodies, which, when cultured with retinoic acid, are induced to differentiate into neurons and glia. No AChE activity is present in the undifferentiated stem cells, and mRNA protection analyses do not detect AChE mRNA. Commitment to a neuronal differentiation pathway results in increased levels of AChE mRNA, production of a tetrameric form of the enzyme, and secretion of AChE into the culture medium. Concomitant with subsequent morphological differentiation into neurons, enzyme secretion diminishes and AChE becomes largely tethered to the neuronal cell membranes. The enzyme is attached to the cell surface as a globular tetramer. Its hydrodynamic properties are consistent with association through a noncatalytic hydrophobic subunit rather than anchorage by a glycophospholipid tail. No change in the rate of transcription of the Ache gene was detected during the course of differentiation, suggesting that the gene is actively transcribed at very early stages of development. Results suggest that stabilization of a labile mRNA governs the increase in AChE mRNA and gene product. The studies presented indicate that an early event in neuronal differentiation is the stabilization of the mRNA leading to expression of a secreted form of AChE. A subsequent step associated with neurite outgrowth results in a transition from secretion of the tetrameric enzyme to its localization on the cell membrane.

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“…The reaction of chloro-7-nitrobenzofurazan with 12-aminodecanoic acid proceeded in the presence of NaHCO 3 in ethanol to afford the corresponding NBF-conjugated decanoic acid 4 in 56% yield. 30 The coupling reaction of 3 29,31 with 4 gave the fluorescence-labeled hydrophobic tail function 5 in 55% yield. Removal of the TBS group in compound 5, followed by ester formation with bromoacetylbromide afforded the fluorescencelabeled lipid precursor 6 in 68% yield in two steps.…”

Section: Synthesismentioning

confidence: 99%

Design and synthesis of fluorescence-labeled closo-dodecaborate lipid: its liposome formation and in vivo imaging targeting of tumors for boronneutron capture therapy

et al. 2012

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The fluorescence-labeled closo-dodecaborane lipid (FL-SBL) was synthesized from (S)-(+)-1,2-isopropylideneglycerol as a chiral starting material. FL-SBL was readily accumulated into the PEGylated DSPC liposomes prepared from DSPC, CH, and DSPE-PEG-OMe by the post insertion protocol. The boron concentrations and the fluorescent intensities of the FL-SBL-labeled DSPC liposomes increased with the increase of the additive FL-SBL, and the maximum emission wavelength of the liposomes appeared at 531 nm. A preliminary in vivo imaging study of tumor-bearing mice revealed that the FL-SBL-labeled DSPC liposomes were delivered to the tumor tissue but not distributed to hypoxic regions.

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Synthesis and characterization of fluorescent neutral lipid analogs containing N-(7-nitro-2,1,3-benzoxadiazol-4-yl)-aminohexanoic acid

Cited by 6 publications

References 20 publications

Membrane Organization at Low Cholesterol Concentrations: A Study Using 7-Nitrobenz-2-oxa-1,3-diazol-4-yl-Labeled Cholesterol

Membrane Organization at Low Cholesterol Concentrations: A Study Using 7-Nitrobenz-2-oxa-1,3-diazol-4-yl-Labeled Cholesterol

Acylglycerol Recycling from Triacylglycerol to Phospholipid, Not Lipase Activity, Is Defective in Neutral Lipid Storage Disease Fibroblasts

Design and synthesis of fluorescence-labeled closo-dodecaborate lipid: its liposome formation and in vivo imaging targeting of tumors for boronneutron capture therapy

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