This study aims to explore the effects of arachidonic acid lipoxygenase metabolism in vascular calcification. We used 5/6 nephrectomy and high-phosphorus feeding to establish a model of vascular calcification in mice. Six weeks after nephrectomy surgery, vascular calcium content was measured, and Alizarin Red S and Von Kossa staining were applied to detect calcium deposition in aortic arch. Control aortas and calcified aortas were collected for mass spectrometry detection of arachidonic acid metabolites, and active molecules in lipoxygenase pathway were analyzed. Real-time quantitative PCR was used to detect changes in the expression of lipoxygenase in calcified aortas. Lipoxygenase inhibitor was used to clarify the effect of lipoxygenase metabolic pathways on vascular calcification. The results showed that 6 weeks after nephrectomy surgery, the aortic calcium content of the surgery group was significantly higher than that of the sham group (P < 0.05). Alizarin Red S staining and Von Kossa staining showed obvious calcium deposition in aortic arch from surgery group, indicating formation of vascular calcification. Nine arachidonic acid lipoxygenase metabolites were quantitated using liquid chromatography/mass spectrometry (LC-MS) analysis. The content of multiple metabolites (12-HETE, 11-HETE, 15-HETE, etc.) was significantly increased in calcified aortas, and the most abundant and up-regulated metabolite was 12-HETE. Furthermore, we examined the mRNA levels of metabolic enzymes that produce 12-HETE in calcified blood vessels and found the expression of arachidonate lipoxygenase-15 (Alox15) was increased. Blocking Alox15/12-HETE by Alox15 specific inhibitor PD146176 significantly decreased the plasma 12-HETE content, promoted calcium deposition in aortic arch and increased vascular calcium content. These results suggest that the metabolism of arachidonic acid lipoxygenase is activated in calcified aorta, and the Alox15/12-HETE signaling pathway may play a protective role in vascular calcification.
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid ; Animals ; Arachidonate 12-Lipoxygenase ; Arachidonate 15-Lipoxygenase/metabolism* ; Arachidonic Acid ; Hydroxyeicosatetraenoic Acids ; Lipoxygenase/metabolism* ; Mice ; Signal Transduction ; Vascular Calcification

OBJECTIVE15-lipoxygenase (15-LO) is a prooxidant enzyme which is expressed in asthmatic lungs leading to formation of pro- and anti-inflammatory mediators. Gene expression profiling studies show the association between 15-LO and allergic asthma. This study was designed to observe the expression of 15-LO in lungs of asthmatic rats and examine the effects of dexamethasone on 15-lipoxygenase expression.

METHODSTwenty-seven male Sprague-Dawley (SD) rats were randomly divided into three groups: control, asthma and dexamethasone (DXM) intervention. An asthma model was prepared by sensitization and challenging with ovalbumin. The production of 15-LO in lung tissue homogenates was measured using ELISA.The expression of 15-LO mRNA in lungs was determined by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSThe levels of 15-LO mRNA and protein in the asthma group (0.51 ± 0.14 and 2080 ± 73 μg/mL, respectively) were lower than those in the control group (0.76 ± 0.15 and 2472 ± 106 μg/mL, respectively; P<0.01). DXM intervention increased significantly the levels of 15-LO mRNA and protein (1.02 ± 0.34 and 2562 ± 218 μg/mL) compared with the asthma group (P<0.01).

CONCLUSIONSThe production of 15-LO in lung tissues is reduced in asthmatic rats. DXM can increase the expression of 15-LO in lung tissues and thus might provide anti-inflammatory effects in asthmatic rats.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Arachidonate 15-Lipoxygenase ; analysis ; genetics ; Asthma ; drug therapy ; enzymology ; Dexamethasone ; pharmacology ; Lung ; enzymology ; pathology ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley

BACKGROUND AND OBJECTIVES: In oral cavity cancer (OCC) cells, the effects of interleukin-4 (IL-4) are various according to the cell specificity. However, if IL-4 induces apoptosis on OCC cells, the mediator of this apoptosis is uncertain. Therefore, we investigated whether apoptosis of OCC cells occurs by IL-4 and whether 15-lipoxygenase-1 (15-LO-1) induced by IL-4 is the possible mediator of this apoptosis. MATERIALS AND METHODS: SCC 1483 cells were used. Flow cytometry and poly ADP-ribose polymerase cleavage were used to examine apoptosis. Western blot analysis and reverse transcription-polymerase chain reaction were used to measure 15-LO-1 protein and mRNA. RESULTS: The inhibition of cell proliferation by more than 50% was noted from 10 ng/ml of IL-4. At this dose, apoptosis was observed and this apoptosis was inhibited by 2.2 microM caffeic acid. 15-LO-1 expression was observed from the 8 hour treatment of IL-4 and apoptosis increased after the 24 hour treatment of IL-4. In this apoptosis, caspase cascade, cyclooxygenase-2, and non-steroidal anti-inflammatory drugs-activated gene-1 (NAG-1) were not involved. CONCLUSION: IL-4 induced apoptosis in SCC 1483 OCC cells and 15-LO-1 induced by IL-4 may mediate this apoptotic pathway.
Adenosine Diphosphate Ribose ; Apoptosis* ; Arachidonate 15-Lipoxygenase ; Blotting, Western ; Cell Proliferation ; Cyclooxygenase 2 ; Flow Cytometry ; Interleukin-4* ; Mouth Neoplasms ; Mouth* ; RNA, Messenger ; Sensitivity and Specificity

Pulmonary arterial hypertension (PAH) is a rare disease with a complex aetiology characterized by elevated pulmonary artery resistance, which leads to progressive right ventricular failure and ultimately death. The aberrant metabolism of arachidonic acid in the pulmonary vasculature plays a central role in the pathogenesis of PAH. The levels of 15-lipoxygenase (15-LO) and 15-hydroxyeicosatetraenoic acid (15-HETE) are elevated in the pulmonary arterial endothelial cells (PAECs), pulmonary smooth muscle cells (PASMCs) and fibroblasts of PAH patients. Under hypoxia condition, 15-LO/15-HETE induces pulmonary artery contraction, promotes the proliferation of PAECs and PASMCs, inhibits apoptosis of PASMCs, promotes fibrosis of pulmonary vessels, and then leads to the occurrence of PAH. Here, we review the research progress on the relationship between 15-LO/15-HETE and hypoxic PAH, in order to clarify the significance of 15-LO/15-HETE in hypoxic PAH.
Arachidonate 15-Lipoxygenase ; Cell Proliferation ; Cells, Cultured ; Endothelial Cells ; Humans ; Hydroxyeicosatetraenoic Acids ; Hypoxia ; Myocytes, Smooth Muscle ; Pulmonary Arterial Hypertension ; Pulmonary Artery