The Caspase Pathway of Linoelaidic Acid (9t, 12t-C18:2)-Induced Apoptosis in Human Umbilical Vein Endothelial Cells
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ORIGINAL ARTICLE
The Caspase Pathway of Linoelaidic Acid (9t, 12t-C18:2)-Induced Apoptosis in Human Umbilical Vein Endothelial Cells Qiu Bin • Huan Rao • Jiang-Ning Hu • Rong Liu • Ya-Wei Fan • Jing Li • Ze-Yuan Deng Xianfeng Zhong • Fang-Ling Du
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Received: 1 February 2012 / Accepted: 25 September 2012 / Published online: 13 October 2012 Ó AOCS 2012
Abstract Trans fatty acids (TFA) are reported to contribute to inflammation and coronary heart disease. The study aim was to investigate the proapoptotic effects of two double bond TFA (TDTFA) on human umbilical vein endothelial cells (HUVEC). The HUVEC were grown in media supplied with linoelaidic acid (9t,12t-C18:2) at 50, 100, 200, 400 lmol/l for 24 or 48 h to examine the effects of TDTFA on the viability and apoptosis of these cells. Flow cytometry analysis and confocal scanning were used to measure apoptosis, cell binding of Annexin V and propidium iodide uptake. Colorimetric assay and RT-PCR were used to analyze enzyme activities and mRNA expression of caspase-3, -8 and -9 in HUVEC. Results showed that 9t,12t-C18:2 inhibited the viability of HUVEC in a dose-dependent and time-dependent manner. The percentages of 9t,12t-C18:2 induced apoptotic and necrotic cells significantly increased compared with that of the control. The activities and mRNA expression of caspase-8, -9 and -3 were significantly increased in 9t,12t-C18:2 treated cells compared to that of the control. Addition of
Q. Bin H. Rao J.-N. Hu R. Liu Y.-W. Fan (&) J. Li Z.-Y. Deng (&) X. Zhong State Key Laboratory of Food Science and Technology, Institute for Advanced Study, Nanchang University, Nanchang 330047, Jiangxi, China e-mail: [email protected] Z.-Y. Deng e-mail: [email protected]; [email protected] Q. Bin e-mail: [email protected] Q. Bin F.-L. Du Institute of Agro-Food Science and Technology, Shandong Academy of Agricultural Sciences, Jinan 250100, China
specific inhibitors of caspase-8 (z-IETD-fmk) and caspase9 (z-LEHD-fmk) to HUVEC was found to completely inhibit 9t,12t-C18:2-induced activation of caspase-3, and z-IETD-fmk inhibited the activation of caspase-9. Meanwhile, it was found that mRNA expression of Bid, Smac/ DIABLO and the release of mitochondrial cytochrome c were significantly elevated by 9t,12t-C18:2 treatment. These results suggest that 9t,12t-C18:2 may induce apoptosis of HUVEC through activating caspase-8, -9 and -3. Both the death receptor pathway and the mitochondrial pathway may be involved in the apoptosis induced by 9t,12t-C18:2. Keywords Linoelaidic acid Human umbilical vein endothelial cells Caspase Apoptosis
Abbreviations ERK HDL HUVEC LDL MAPK PI ROS Smac/DIABLO
TFA TDTFA HAPES CHAPS DTT
Extracellular signal-regulated kinases High-density lipoprotein Human umbilical vein endothelial cells Low-density lipoprotein Mitogen-activated protein kinase Propidium iodide Reactive oxygen species Second mitochondria-derived activator of caspase/direct IAP binding protein with low pI Trans fatty acids Two double bonds TFA 4-(2-Hydroxyethyl)-1p
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