Fatty acids induce apoptosis in human smooth muscle cells depending on chain length, saturation, and duration of exposure.
OBJECTIVE
Plasma free fatty acid (FFA) concentrations are increased in states of insulin resistance. Therefore, this study evaluated apoptosis and underlying mechanisms induced by selected nutritional FFAs, a defined FFA-mix, and human plasma containing high FFA concentrations in human smooth muscle cells (HSMCs).


METHODS
HSMCs were incubated (24-72 h) with selected FFAs (100-300 micromol/l), an FFA-mix (palmitic-/stearic-/oleic-/linoleic-/alpha-linolenic acid=2.6/1/3.6/9/1; 300-900 micromol/l), or with high FFA-plasma (600 micromol/l) versus respective control cultures. Apoptosis, caspase activation, and protein expression were determined by DNA-fragmentation assays, flow cytometry, and Western blots, respectively.


RESULTS
Exposure (24h) of HSMCs to 300 micromol/l stearic-, oleic-, linoleic-, alpha-linolenic-, and arachidonic acid induced apoptosis, correlating (p<0.01) with the FFAs' chain length (r=0.602) and number of FFA double bonds (r=0.956). After 48 h, 100 micromol/l of all tested FFAs - including palmitic acid - were already sufficient to trigger HSMCs' cell death. FFA-exposure resulted in activation of caspases and apoptosis was completely abolished by co-incubation with caspase inhibitors and negatively correlated (p<0.01) with the base-excision repair protein XRCC1 (r=-0.765) and with c-myc's antagonist mad (r=-0.916), whereas positive correlations (p<0.01) were found for protein expression of the proto-oncogene c-myc (r=0.972) and the transcription factor E2F-1 (r=0.971). Exposure of HSMCs to the defined FFA-mix and to plasma samples from individuals with elevated plasma FFAs supported the results obtained by defined FFA stimulation.


CONCLUSIONS
Since smooth muscle cells surround the macrophage/foam cell/lipid-laden artheromatous core of atherosclerotic lesions with a protective fibrous cap, their FFA-induced HSMC apoptosis could contribute to progression of atherosclerosis by thinning of the fibrous cap and subsequent plaque destabilization.
Artwohl
Michaela
M
Department of Internal Medicine III, Clinical Division of Endocrinology and Metabolism, Medical University of Vienna, Währinger Gürtel 18-20, A-1090 Vienna, Austria. michaela.artwohl@meduniwien.ac.at
Lindenmair
Andrea
A
Roden
Michael
M
Waldhäusl
Werner-Klaus
WK
Freudenthaler
Angelika
A
Klosner
Gabriele
G
Ilhan
Aysegül
A
Luger
Anton
A
Baumgartner-Parzer
Sabina M
SM
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology*; D016718:Arachidonic Acid / Q000494:pharmacology; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D045304:Cytochromes c / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D050687:E2F1 Transcription Factor / Q000378:metabolism; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology*; D006801:Humans; D019787:Linoleic Acid / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D019301:Oleic Acid / Q000494:pharmacology; D019308:Palmitic Acid / Q000494:pharmacology; D000090063:Proto-Oncogene Mas; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D013229:Stearic Acids / Q000494:pharmacology; D014470:Umbilical Cord / Q000166:cytology; D000076105:X-ray Repair Cross Complementing Protein 1; D017962:alpha-Linolenic Acid / Q000494:pharmacology
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D005230:Fatty Acids, Nonesterified; C000715470:MAS1 protein, human; C079439:MXD1 protein, human; C489427:MYC protein, human; D000090063:Proto-Oncogene Mas; D016271:Proto-Oncogene Proteins c-myc; D012097:Repressor Proteins; D013229:Stearic Acids; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human; D017962:alpha-Linolenic Acid; D016718:Arachidonic Acid; D019301:Oleic Acid; D019308:Palmitic Acid; C031183:stearic acid; D045304:Cytochromes c; D019787:Linoleic Acid; D020169:Caspases