Changes in cell phospholipid metabolism in vitro in the presence of HEMA and its degradation products

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Objectives: Diacylglycerol-kinase (DAG-kinase) is an enzyme that phosphorylates diacylglycerol (DAG) to phosphatidic acid (PA), which serves as a precursor to phosphoglycerides involved in cell signaling or as cell membrane structural components. DAG-kinase can be inhibited by diacylethylene glycols (DAEG). We hypothesize that 2-hydroxyethyl methacrylate (HEMA) may alter phosphorylation of DAG to PA following intracellular formation of DAEG. Methods: Cultured rabbit kidney (RK13) epithelial cells were treated with HEMA, EG, or known inhibitors of DAG-kinase for 24 h, then exposed to [32P]O-4 in the presence of a synthetic diacylglycerol for 2 h. Other cultures were radiolabeled with [3H]-oleic acid for 24 h, then exposed to HEMA for an additional 24 h. The cells were harvested and the lipids extracted. Radioactive lipids were separated by thin layer chromatography, located by autoradiography, and quantitated as cpm/ug protein. Cell cultures treated with HEMA were homogenized and the DAG-kinase activity was assayed and expressed as cpm/ug protein. Data were analyzed by one-way ANOVA and Newman-Keuls Multiple Comparison Test. Results: Cultures exposed to HEMA or known DAG-kinase inhibitors exhibited reduced incorporation of radioactivity in the PA fraction compared to control cultures. Direct assays of DAG-kinase activity from cells exposed to HEMA demonstrated decreased enzyme activity. Evaluation of cell phospholipid synthesis showed altered formation of phosphatidylethanolamine and phosphatidylcholine. Significance: Results suggest that HEMA impairs formation of PA, possibly by acylation of EG released by hydrolysis of the HEMA and resultant production of the inhibitor DAEG. The decreased availability of PA may alter PA-dependent cell structural lipid pathways and lipid-dependent signaling pathways, altering cell growth.

Original languageEnglish (US)
Pages (from-to)297-302
Number of pages6
JournalDental Materials
Volume16
Issue number4
DOIs
StatePublished - Jan 1 2000

Fingerprint

Phospholipids
Diacylglycerol Kinase
Metabolism
Phosphatidic Acids
Degradation
Lipids
Acids
Glycols
Diglycerides
Cell signaling
Proteins
Thin layer chromatography
Acylation
Phosphorylation
Oleic acid
Radioactivity
Enzyme activity
Cell growth
Cell membranes
Glycerophospholipids

Keywords

  • 2-Hydroxyethyl methacrylate
  • Cell culture
  • Dental resin
  • Diacylglycerol-kinase
  • Phosphatidic acid
  • Phospholipid

ASJC Scopus subject areas

  • Materials Science(all)
  • Dentistry(all)
  • Mechanics of Materials

Cite this

Changes in cell phospholipid metabolism in vitro in the presence of HEMA and its degradation products. / Schuster, G. S.; Caughman, Gretchen B; Rueggeberg, Frederick.

In: Dental Materials, Vol. 16, No. 4, 01.01.2000, p. 297-302.

Research output: Contribution to journalArticle

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abstract = "Objectives: Diacylglycerol-kinase (DAG-kinase) is an enzyme that phosphorylates diacylglycerol (DAG) to phosphatidic acid (PA), which serves as a precursor to phosphoglycerides involved in cell signaling or as cell membrane structural components. DAG-kinase can be inhibited by diacylethylene glycols (DAEG). We hypothesize that 2-hydroxyethyl methacrylate (HEMA) may alter phosphorylation of DAG to PA following intracellular formation of DAEG. Methods: Cultured rabbit kidney (RK13) epithelial cells were treated with HEMA, EG, or known inhibitors of DAG-kinase for 24 h, then exposed to [32P]O-4 in the presence of a synthetic diacylglycerol for 2 h. Other cultures were radiolabeled with [3H]-oleic acid for 24 h, then exposed to HEMA for an additional 24 h. The cells were harvested and the lipids extracted. Radioactive lipids were separated by thin layer chromatography, located by autoradiography, and quantitated as cpm/ug protein. Cell cultures treated with HEMA were homogenized and the DAG-kinase activity was assayed and expressed as cpm/ug protein. Data were analyzed by one-way ANOVA and Newman-Keuls Multiple Comparison Test. Results: Cultures exposed to HEMA or known DAG-kinase inhibitors exhibited reduced incorporation of radioactivity in the PA fraction compared to control cultures. Direct assays of DAG-kinase activity from cells exposed to HEMA demonstrated decreased enzyme activity. Evaluation of cell phospholipid synthesis showed altered formation of phosphatidylethanolamine and phosphatidylcholine. Significance: Results suggest that HEMA impairs formation of PA, possibly by acylation of EG released by hydrolysis of the HEMA and resultant production of the inhibitor DAEG. The decreased availability of PA may alter PA-dependent cell structural lipid pathways and lipid-dependent signaling pathways, altering cell growth.",
keywords = "2-Hydroxyethyl methacrylate, Cell culture, Dental resin, Diacylglycerol-kinase, Phosphatidic acid, Phospholipid",
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AU - Caughman, Gretchen B

AU - Rueggeberg, Frederick

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Y1 - 2000/1/1

N2 - Objectives: Diacylglycerol-kinase (DAG-kinase) is an enzyme that phosphorylates diacylglycerol (DAG) to phosphatidic acid (PA), which serves as a precursor to phosphoglycerides involved in cell signaling or as cell membrane structural components. DAG-kinase can be inhibited by diacylethylene glycols (DAEG). We hypothesize that 2-hydroxyethyl methacrylate (HEMA) may alter phosphorylation of DAG to PA following intracellular formation of DAEG. Methods: Cultured rabbit kidney (RK13) epithelial cells were treated with HEMA, EG, or known inhibitors of DAG-kinase for 24 h, then exposed to [32P]O-4 in the presence of a synthetic diacylglycerol for 2 h. Other cultures were radiolabeled with [3H]-oleic acid for 24 h, then exposed to HEMA for an additional 24 h. The cells were harvested and the lipids extracted. Radioactive lipids were separated by thin layer chromatography, located by autoradiography, and quantitated as cpm/ug protein. Cell cultures treated with HEMA were homogenized and the DAG-kinase activity was assayed and expressed as cpm/ug protein. Data were analyzed by one-way ANOVA and Newman-Keuls Multiple Comparison Test. Results: Cultures exposed to HEMA or known DAG-kinase inhibitors exhibited reduced incorporation of radioactivity in the PA fraction compared to control cultures. Direct assays of DAG-kinase activity from cells exposed to HEMA demonstrated decreased enzyme activity. Evaluation of cell phospholipid synthesis showed altered formation of phosphatidylethanolamine and phosphatidylcholine. Significance: Results suggest that HEMA impairs formation of PA, possibly by acylation of EG released by hydrolysis of the HEMA and resultant production of the inhibitor DAEG. The decreased availability of PA may alter PA-dependent cell structural lipid pathways and lipid-dependent signaling pathways, altering cell growth.

AB - Objectives: Diacylglycerol-kinase (DAG-kinase) is an enzyme that phosphorylates diacylglycerol (DAG) to phosphatidic acid (PA), which serves as a precursor to phosphoglycerides involved in cell signaling or as cell membrane structural components. DAG-kinase can be inhibited by diacylethylene glycols (DAEG). We hypothesize that 2-hydroxyethyl methacrylate (HEMA) may alter phosphorylation of DAG to PA following intracellular formation of DAEG. Methods: Cultured rabbit kidney (RK13) epithelial cells were treated with HEMA, EG, or known inhibitors of DAG-kinase for 24 h, then exposed to [32P]O-4 in the presence of a synthetic diacylglycerol for 2 h. Other cultures were radiolabeled with [3H]-oleic acid for 24 h, then exposed to HEMA for an additional 24 h. The cells were harvested and the lipids extracted. Radioactive lipids were separated by thin layer chromatography, located by autoradiography, and quantitated as cpm/ug protein. Cell cultures treated with HEMA were homogenized and the DAG-kinase activity was assayed and expressed as cpm/ug protein. Data were analyzed by one-way ANOVA and Newman-Keuls Multiple Comparison Test. Results: Cultures exposed to HEMA or known DAG-kinase inhibitors exhibited reduced incorporation of radioactivity in the PA fraction compared to control cultures. Direct assays of DAG-kinase activity from cells exposed to HEMA demonstrated decreased enzyme activity. Evaluation of cell phospholipid synthesis showed altered formation of phosphatidylethanolamine and phosphatidylcholine. Significance: Results suggest that HEMA impairs formation of PA, possibly by acylation of EG released by hydrolysis of the HEMA and resultant production of the inhibitor DAEG. The decreased availability of PA may alter PA-dependent cell structural lipid pathways and lipid-dependent signaling pathways, altering cell growth.

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KW - Cell culture

KW - Dental resin

KW - Diacylglycerol-kinase

KW - Phosphatidic acid

KW - Phospholipid

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