Inhibition of acyl-CoA: cholesterol acyltransferase in chinese hamster ovary (CHO) cells by short-chain ceramide and dihydroceramide

Neale D. Ridgway

doi:10.1016/0005-2760(95)00009-2

Inhibition of acyl-CoA: cholesterol acyltransferase in chinese hamster ovary (CHO) cells by short-chain ceramide and dihydroceramide

Neale D. Ridgway

Medicine

Medicine

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27 Citations (Scopus)

Résumé

The biological activity of ceramide, an intermediate in the synthesis and catabolism of sphingolipids, has been shown to be mimicked by short-chain N-acyl analogues. A potential role for ceramide in modulating cholesterol esterification was investigated using a series of short-chain ceramides and dihydroceramides. Acyl-CoA:cholesterol acyltransferase (ACAT) in CHO cells was inhibited rapidly (< 30 min) and in a dose-dependent fashion by two N-acyl analogues of naturally occurring d-erythro-ceramide, N-acetyl-sphingosine (d-erythro-C₂-ceramide) and N-hexanoyl-sphingosine (d-erythro-C₆-ceramide). At 10 μM d-erythro-C₂-ceramide, esterification of cholesterol was inhibited by 95% in CHO cells grown in delipidated serum, and 80-85% in cells grown in 25-hydroxycholesterol or human low-density lipoprotein (LDL). d-erythro-C₂-Ceramide did not inhibit [¹⁴C]oleate-labelling of triacylglycerol and phospholipid. Inhibition of cholesterol esterification in cells and isolated membranes required the d-erythro (2S,3R) configuration (the l-threo isomer of C₂-ceramide was not inhibitory) and an N-acyl group (sphingosine and sphinganine did not inhibit). dl-erythro-C₂-Dihydroceramide was also a potent ACAT inhibitor in isolated membranes (IC₅₀, 0.2 μM) and cells indicating lack of requirement for a 4-trans double bond. Consistent with results for C₂-ceramides, dl-threo-C₂-dihydroceramide was not inhibitory in cells or in vitro. Long-chain ceramide and N-palmitoyl-dihydroceramide did not inhibit ACAT in isolated membranes. Compared to d-erythro-C₂-ceramide, d-erythro-C₆- and C₄-ceramide were slightly weaker inhibitors of ACAT in isolated membranes. Thus, N-acyl chain length could influence inhibition, either by altering the effective concentration of ceramide in membranes or affinity for the ACAT enzyme. Short-chain ceramides and dihydroceramides are the first ACAT inhibitors described with structural similarity to a naturally occurring compound.

Langue d'origine	English
Pages (de-à)	39-46
Nombre de pages	8
Journal	Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
Volume	1256
Numéro de publication	1
DOI	https://doi.org/10.1016/0005-2760(95)00009-2
Statut de publication	Published - avr. 28 1995

Note bibliographique

Funding Information:
~ This work was supported by a grant and scholarship from the Medical Research Council of Canada, and an establishment grant from Izaak Walton Killiam Children's Hospital. Abbreviations: ACAT, acyl-CoA:cholesterol acyltransferase; BSA, bovine serum albumin; C8-APPD, L-threo-2-(N-octanoyl-amino)-l- phenyl-l,3-propanodiol; C2-ceramide, N-acetyl-sphingosine; C2-dihydroceramide, N-acetyl-sphinganine; C4-ceramide, N-butyl-sphingosine; C 6-ceramide, N-hexanoyl-sphingosine; L-MAPP, L-threo-2-(N-myristoyl- amino)-l-phenyl-l-propanol; D-MAPP, D-erythro-2-(N-myristoyl-amino)- 1-phenyl-l-propanol; TNF, tumor necrosis factor. * Corresponding author. Fax: + 1 (902) 494 1394.

ASJC Scopus Subject Areas

Biophysics
Biochemistry
Endocrinology

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10.1016/0005-2760(95)00009-2

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title = "Inhibition of acyl-CoA: cholesterol acyltransferase in chinese hamster ovary (CHO) cells by short-chain ceramide and dihydroceramide",

abstract = "The biological activity of ceramide, an intermediate in the synthesis and catabolism of sphingolipids, has been shown to be mimicked by short-chain N-acyl analogues. A potential role for ceramide in modulating cholesterol esterification was investigated using a series of short-chain ceramides and dihydroceramides. Acyl-CoA:cholesterol acyltransferase (ACAT) in CHO cells was inhibited rapidly (< 30 min) and in a dose-dependent fashion by two N-acyl analogues of naturally occurring d-erythro-ceramide, N-acetyl-sphingosine (d-erythro-C2-ceramide) and N-hexanoyl-sphingosine (d-erythro-C6-ceramide). At 10 μM d-erythro-C2-ceramide, esterification of cholesterol was inhibited by 95% in CHO cells grown in delipidated serum, and 80-85% in cells grown in 25-hydroxycholesterol or human low-density lipoprotein (LDL). d-erythro-C2-Ceramide did not inhibit [14C]oleate-labelling of triacylglycerol and phospholipid. Inhibition of cholesterol esterification in cells and isolated membranes required the d-erythro (2S,3R) configuration (the l-threo isomer of C2-ceramide was not inhibitory) and an N-acyl group (sphingosine and sphinganine did not inhibit). dl-erythro-C2-Dihydroceramide was also a potent ACAT inhibitor in isolated membranes (IC50, 0.2 μM) and cells indicating lack of requirement for a 4-trans double bond. Consistent with results for C2-ceramides, dl-threo-C2-dihydroceramide was not inhibitory in cells or in vitro. Long-chain ceramide and N-palmitoyl-dihydroceramide did not inhibit ACAT in isolated membranes. Compared to d-erythro-C2-ceramide, d-erythro-C6- and C4-ceramide were slightly weaker inhibitors of ACAT in isolated membranes. Thus, N-acyl chain length could influence inhibition, either by altering the effective concentration of ceramide in membranes or affinity for the ACAT enzyme. Short-chain ceramides and dihydroceramides are the first ACAT inhibitors described with structural similarity to a naturally occurring compound.",

author = "Ridgway, {Neale D.}",

note = "Funding Information: ~ This work was supported by a grant and scholarship from the Medical Research Council of Canada, and an establishment grant from Izaak Walton Killiam Children's Hospital. Abbreviations: ACAT, acyl-CoA:cholesterol acyltransferase; BSA, bovine serum albumin; C8-APPD, L-threo-2-(N-octanoyl-amino)-l- phenyl-l,3-propanodiol; C2-ceramide, N-acetyl-sphingosine; C2-dihydroceramide, N-acetyl-sphinganine; C4-ceramide, N-butyl-sphingosine; C 6-ceramide, N-hexanoyl-sphingosine; L-MAPP, L-threo-2-(N-myristoyl- amino)-l-phenyl-l-propanol; D-MAPP, D-erythro-2-(N-myristoyl-amino)- 1-phenyl-l-propanol; TNF, tumor necrosis factor. * Corresponding author. Fax: + 1 (902) 494 1394.",

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doi = "10.1016/0005-2760(95)00009-2",

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T1 - Inhibition of acyl-CoA

T2 - cholesterol acyltransferase in chinese hamster ovary (CHO) cells by short-chain ceramide and dihydroceramide

AU - Ridgway, Neale D.

N1 - Funding Information: ~ This work was supported by a grant and scholarship from the Medical Research Council of Canada, and an establishment grant from Izaak Walton Killiam Children's Hospital. Abbreviations: ACAT, acyl-CoA:cholesterol acyltransferase; BSA, bovine serum albumin; C8-APPD, L-threo-2-(N-octanoyl-amino)-l- phenyl-l,3-propanodiol; C2-ceramide, N-acetyl-sphingosine; C2-dihydroceramide, N-acetyl-sphinganine; C4-ceramide, N-butyl-sphingosine; C 6-ceramide, N-hexanoyl-sphingosine; L-MAPP, L-threo-2-(N-myristoyl- amino)-l-phenyl-l-propanol; D-MAPP, D-erythro-2-(N-myristoyl-amino)- 1-phenyl-l-propanol; TNF, tumor necrosis factor. * Corresponding author. Fax: + 1 (902) 494 1394.

PY - 1995/4/28

Y1 - 1995/4/28

N2 - The biological activity of ceramide, an intermediate in the synthesis and catabolism of sphingolipids, has been shown to be mimicked by short-chain N-acyl analogues. A potential role for ceramide in modulating cholesterol esterification was investigated using a series of short-chain ceramides and dihydroceramides. Acyl-CoA:cholesterol acyltransferase (ACAT) in CHO cells was inhibited rapidly (< 30 min) and in a dose-dependent fashion by two N-acyl analogues of naturally occurring d-erythro-ceramide, N-acetyl-sphingosine (d-erythro-C2-ceramide) and N-hexanoyl-sphingosine (d-erythro-C6-ceramide). At 10 μM d-erythro-C2-ceramide, esterification of cholesterol was inhibited by 95% in CHO cells grown in delipidated serum, and 80-85% in cells grown in 25-hydroxycholesterol or human low-density lipoprotein (LDL). d-erythro-C2-Ceramide did not inhibit [14C]oleate-labelling of triacylglycerol and phospholipid. Inhibition of cholesterol esterification in cells and isolated membranes required the d-erythro (2S,3R) configuration (the l-threo isomer of C2-ceramide was not inhibitory) and an N-acyl group (sphingosine and sphinganine did not inhibit). dl-erythro-C2-Dihydroceramide was also a potent ACAT inhibitor in isolated membranes (IC50, 0.2 μM) and cells indicating lack of requirement for a 4-trans double bond. Consistent with results for C2-ceramides, dl-threo-C2-dihydroceramide was not inhibitory in cells or in vitro. Long-chain ceramide and N-palmitoyl-dihydroceramide did not inhibit ACAT in isolated membranes. Compared to d-erythro-C2-ceramide, d-erythro-C6- and C4-ceramide were slightly weaker inhibitors of ACAT in isolated membranes. Thus, N-acyl chain length could influence inhibition, either by altering the effective concentration of ceramide in membranes or affinity for the ACAT enzyme. Short-chain ceramides and dihydroceramides are the first ACAT inhibitors described with structural similarity to a naturally occurring compound.

AB - The biological activity of ceramide, an intermediate in the synthesis and catabolism of sphingolipids, has been shown to be mimicked by short-chain N-acyl analogues. A potential role for ceramide in modulating cholesterol esterification was investigated using a series of short-chain ceramides and dihydroceramides. Acyl-CoA:cholesterol acyltransferase (ACAT) in CHO cells was inhibited rapidly (< 30 min) and in a dose-dependent fashion by two N-acyl analogues of naturally occurring d-erythro-ceramide, N-acetyl-sphingosine (d-erythro-C2-ceramide) and N-hexanoyl-sphingosine (d-erythro-C6-ceramide). At 10 μM d-erythro-C2-ceramide, esterification of cholesterol was inhibited by 95% in CHO cells grown in delipidated serum, and 80-85% in cells grown in 25-hydroxycholesterol or human low-density lipoprotein (LDL). d-erythro-C2-Ceramide did not inhibit [14C]oleate-labelling of triacylglycerol and phospholipid. Inhibition of cholesterol esterification in cells and isolated membranes required the d-erythro (2S,3R) configuration (the l-threo isomer of C2-ceramide was not inhibitory) and an N-acyl group (sphingosine and sphinganine did not inhibit). dl-erythro-C2-Dihydroceramide was also a potent ACAT inhibitor in isolated membranes (IC50, 0.2 μM) and cells indicating lack of requirement for a 4-trans double bond. Consistent with results for C2-ceramides, dl-threo-C2-dihydroceramide was not inhibitory in cells or in vitro. Long-chain ceramide and N-palmitoyl-dihydroceramide did not inhibit ACAT in isolated membranes. Compared to d-erythro-C2-ceramide, d-erythro-C6- and C4-ceramide were slightly weaker inhibitors of ACAT in isolated membranes. Thus, N-acyl chain length could influence inhibition, either by altering the effective concentration of ceramide in membranes or affinity for the ACAT enzyme. Short-chain ceramides and dihydroceramides are the first ACAT inhibitors described with structural similarity to a naturally occurring compound.

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Inhibition of acyl-CoA: cholesterol acyltransferase in chinese hamster ovary (CHO) cells by short-chain ceramide and dihydroceramide

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