1.Effects of PPAR-alpha activation on oleic acid-induced steatosis and expression of heme oxygenase-1 in HepG2 cells.
Jing-jing ZHAO ; Long-feng ZHAO ; Hui YANG ; Li ZHANG
Chinese Journal of Hepatology 2013;21(3):218-221
OBJECTIVETo investigate the effects of peroxisome proliferator activated receptor-alpha (PPAR-a) activation on oleic acid (OA)-induced steatosis and hepatic expression of heme oxygenase-1 (HO-1) using an in vitro cell model system.
METHODSA steatosis human hepatocyte in vitro model system was established by treating HepG2 cells with 0.2 mmol/L of oleic acid for 24 hours. The steatosis cells were then divided into four groups for an additional 24 hours of treatment with 0.2 mmol/L of oleic acid alone (model control group) or with 5, 10 or 50 pnol/L of fenofibrate (FF, a selective PPAR-a agonist; experimental groups). Untreated HepG2 cells served as non-steatosis controls. Effect of PPAR-a activation on fat accumulation was detected by Oil Red O staining and on intracellular triglyceride (TG) levels by enzymatic assay. mRNA and protein expression of PPAR-alpha and HO-1 were quantified by real-time PCR and immunocytochemistry, respectively. One-way ANOVA and the LSD t-test were used for between-group comparisons, and correlation analysis was performed with the Pearson's correlation coefficient.
RESULTSThe steatosis model control cells showed significantly increased TG deposition (379.98 +/- 23.19 mg/g protein, vs. non-steatosis controls F = 148.56, P< 0.01), significantly decreased mRNA and protein expression of PPAR-alpha (0.42 +/- 0.38,F= 177.64,P< 0.01 and 0.47 +/- 0.14, F= 120.76,P< 0.01) and HO-1 (0.36 +/- 0.66, F= 74.77,P< 0.01 and 0.26 +/- 0.10,F= 119.90,P<0.01). FF (5, 10 and 50 micromol/L) inhibited the steatosis induced by OA in a concentration-dependent manner (294.00 +/- 19.80, 250.33 +/- 9.96, and 196.99 +/- 9.14, F = 148.56, P <0.01) and increased the mRNA and protein expression of PPAR-alpha (0.55 +/- 0.65, 0.85 +/- 0.61, and 1.31 +/- 0.36,F= 177.64,P< 0.01; 0.82 + 0.11, 1.31 +/- 0.16, and 1.75 +/- 0.13, F= 120.76,P <0.01) and HO-1 (0.62 +/- 0.05, 0.84 +/- 0.07, and 1.30 +/- 0.11,F= 74.77,P <0.01; 0.44 +/- 0.08, 0.81 +/- 0.08, 1.20 +/- 0.10,F= 119.90,P< 0.01).
CONCLUSIONActivation of PPAR-a prevents OA-induced steatosis in HepG2 cells, and HO-1 may function as a downstream effector of this mechanism.
Fatty Liver ; chemically induced ; Heme Oxygenase-1 ; metabolism ; Hep G2 Cells ; Humans ; Oleic Acid ; pharmacology ; PPAR alpha ; metabolism ; Triglycerides ; metabolism
2.Lipid Accumulation and IL-6 Production in L02 Hepatocytes Induced by Sodium Oleate: Dose and Time Dependence.
Chen CHEN ; Jun Sheng HUO ; Qin ZHUO ; Yan LI ; Yan MA ; Jing Bo WANG ; Xu Dong JIA
Biomedical and Environmental Sciences 2021;34(12):998-1004
To explore interleukin-6 (IL-6) production and characterize lipid accumulation in L02 hepatocytes induced by sodium oleate. L02 hepatocytes were incubated with 0, 37.5, 75, 150, 300, 600, or 1,200 μmol/L sodium oleate for 24 h, and the supernatant was collected to detect the concentration of IL-6. L02 hepatocytes were incubated with 300, 150, 75, or 0 μmol/L sodium oleate for 0-24 h. The supernatant was collected for detection of IL-6 and free fatty acids. L02 hepatocytes treated with 300 μmol/L sodium oleate for 0-24 h were stained with Oil Red O. With extended sodium oleate incubation time, IL-6 levels increased, and free fatty acids decreased. After 24 h incubation, IL-6 levels increased as sodium oleate increased from 37.5 to 300 μmol/L (
Dose-Response Relationship, Drug
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Hepatocytes/metabolism*
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Humans
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Interleukin-6/metabolism*
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Lipid Metabolism
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Oleic Acid/administration & dosage*
;
Time Factors
3.Effects of dilution rate and carbon-to-nitrogen ratio on lipid accumulation by Rhodosporidium toruloides under chemostat conditions.
Hongwei SHEN ; Guojie JIN ; Cuimin HU ; Zhiwei GONG ; Fengwu BAI ; Zongbao K ZHAO
Chinese Journal of Biotechnology 2012;28(1):56-64
The objective of this work is to investigate how dilution rate and carbon-to-nitrogen (C/N) ratio affects lipid accumulation by Rhodosporidium toruloides AS 2.138 9 in continuous culture. Under steady-state conditions, the increase in dilution rate led to the decrease in lipid content and lipid yield. The highest lipid yield and lipid content at D = 0.02 h(-1) were 0.18 g lipid/g sugar and 57.1%, respectively, while the highest lipid productivity and biomass productivity were obtained at D = 0.14 h(-1). The increase in C/N ratio led to the increase in lipid content. The highest lipid content of 38% was obtained at C/N = 237. The highest lipid yield of 0.12 g lipid/g sugar was obtained at C/N = 92. However, the highest lipid productivity of 0.12 g/(L x h) was obtained at C/N = 32. No significant changes were observed in terms of fatty acid composition of the lipid produced under different C/N ratios, and these three fatty acids, palmitic acid, stearic acid and oleic acid, took over 85% in all samples.
Basidiomycota
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growth & development
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metabolism
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Batch Cell Culture Techniques
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Carbon
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metabolism
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Culture Media
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Fatty Acids
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metabolism
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Glucose
;
metabolism
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Lipids
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biosynthesis
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Nitrogen
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metabolism
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Oleic Acid
;
biosynthesis
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Palmitic Acid
;
metabolism
4.Effect of oleic acid on the proliferation and secretion of pro-inflammatory mediators of human normal fibroblasts and scar fibroblasts.
Qiong JIANG ; Bo-yu WU ; Xiao-dong CHEN
Chinese Journal of Burns 2012;28(6):444-450
OBJECTIVETo investigate the effect of oleic acid on the proliferation and secretion of pro-inflammatory mediators of human normal fibroblasts and scar fibroblasts.
METHODSHuman normal fibroblasts and scar fibroblasts were cultured in vitro and respectively divided into seven groups according to the random number table, with 8 samples in each group. Cells in blank control (BC) group were routinely cultured without addition of other agents. Cells in ethanol-control (EC) group were cultured with medium with the addition of 2% absolute ethanol. Cells in oleic acid groups were cultured with the addition of oleic acid in concentration of 0.25, 0.50, 1.00, 2.00, or 4.00 mmol/L in 2% absolute ethanol. The growth of cells in each group was observed with trypan blue staining on post culture day (PCD) 1-5. On PCD 2, structure of cells in BC, EC, and 1.00 mmol/L oleic acid groups was observed under inverted phase contrast microscope and transmission electron microscope; cell cycle of BC, EC, and 1.00 mmol/L oleic acid groups was measured by flow cytometer; cell proliferation activity in each group was measured by MTT assay; the level of NO in supernatant was assayed by Griess assay; the levels of TNF-α, IL-1β, IL-6, and IL-8 in supernatants in each group were determined by enzyme-linked immunosorbent assay. Data were processed with multifactor and repeated measurement design analysis of variance.
RESULTS(1) There was no significant difference in each index of normal fibroblasts and scar fibroblasts between BC group and EC group. (2) The numbers of normal fibroblasts and scar fibroblasts in 2.00 and 4.00 mmol/L oleic acid groups were significantly lower than those in corresponding BC and EC groups on PCD 2-5 (with F values respectively 13.773 and 11.344, P values all below 0.01). (3) On PCD 2, the numbers of normal fibroblasts and scar fibroblasts in 1.00 mmol/L oleic acid groups decreased, and the cells were aggregating, rounding, and easy to drop off. Cellular membrane disruption, vacuolar degeneration of mitochondrion, pyknosis, and lipid droplets were observed. (4) The percentages of G0/G1 and G2/M phases of normal fibroblasts in 1.00 mmol/L oleic acid group [(93.56 ± 9.98)%, (2.01 ± 0.75)%] were significantly higher than those in BC group [(84.23 ± 10.96)%, (0.37 ± 0.16)%, with F values respectively 3.026, 34.751, P < 0.05 or P < 0.01], while the percentage of normal fibroblasts in S phase [(4.42 ± 0.87)%] was markedly lower than that in BC group [(16.06 ± 1.74)%, F = 136.120, P < 0.01]. The percentages of scar fibroblasts of G0/G1 and G2/M phases in 1.00 mmol/L oleic acid group [(93.86 ± 13.90)%, (1.89 ± 0.66)%] were significantly higher than those in BC group [(83.88 ± 10.42)%, (0.41 ± 0.17)%, with F values respectively 3.529, 32.710, P < 0.05 or P < 0.01], and the percentage of scar fibroblasts in S phase [(3.87 ± 0.63)%] was markedly lower than that in BC group [(15.89 ± 2.02)%, F = 116.508, P < 0.01]. (5) The proliferation rates of normal fibroblasts and scar fibroblasts in 0.50-4.00 mmol/L oleic acid groups were significantly lower than those in corresponding BC and EC groups (with F values respectively 215.945 and 194.555, P < 0.05 or P < 0.01). (6) The content of NO in supernatant of normal fibroblasts in all oleic acid groups was obviously higher than that in BC and EC groups (F = 30.240, P < 0.05 or P < 0.01). The contents of NO in supernatants of scar fibroblasts in 1.00-4.00 mmol/L oleic acid groups were significantly higher than that in BC and EC groups (F = 12.495, P < 0.01). The contents of TNF-α and IL-6 in supernatants of normal fibroblasts and scar fibroblasts in 2.00 and 4.00 mmol/L oleic acid groups were obviously higher than those in corresponding BC and EC groups (with F(TNF-α) values respectively 6.911, 3.818, F(IL-6) values respectively 16.939, 11.600,P < 0.05 or P < 0.01). The contents of IL-1β in supernatants of normal fibroblasts and scar fibroblasts in groups of every concentration of oleic acid were significantly higher than those in corresponding BC and EC groups (with F values respectively 25.117, 9.137, P values all below 0.01). The contents of IL-8 in supernatants of normal fibroblasts in 1.00-4.00 mmol/L oleic acid groups were markedly higher than those in BC and EC groups (F = 2.717, P < 0.05 or P < 0.01). The contents of IL-8 in supernatants of scar fibroblasts in 2.00 and 4.00 mmol/L oleic acid groups were significantly higher than those in BC and EC groups (F = 3.338, P < 0.05). There was no statistically significant difference in above indexes between normal fibroblasts and scar fibroblasts in the same concentration of oleic acid group (with F values from 0.120 to 3.766, P values all above 0.05).
CONCLUSIONSAlthough oleic acid in high concentration inhibits the proliferation of scar fibroblasts, it also inhibits the proliferation of normal fibroblasts. Oleic acid in high concentration can cause excessive and continued inflammatory reaction by promoting the secretion of pro-inflammatory mediators of normal fibroblasts and scar fibroblasts, and they are detrimental to wound healing.
Cell Proliferation ; drug effects ; Cells, Cultured ; Cicatrix ; metabolism ; Fibroblasts ; cytology ; drug effects ; secretion ; Humans ; Inflammation Mediators ; metabolism ; Oleic Acid ; pharmacology
5.Lipid synthesis by an acidic acid tolerant Rhodotorula glutinis.
Zhangnan LIN ; Hongjuan LIU ; Jian'an ZHANG ; Gehua WANG
Chinese Journal of Biotechnology 2016;32(3):339-346
Acetic acid, as a main by-product generated in the pretreatment process of lignocellulose hydrolysis, significantly affects cell growth and lipid synthesis of oleaginous microorganisms. Therefore, we studied the tolerance of Rhodotorula glutinis to acetic acid and its lipid synthesis from substrate containing acetic acid. In the mixed sugar medium containing 6 g/L glucose and 44 g/L xylose, and supplemented with acetic acid, the cell growth was not:inhibited when the acetic acid concentration was below 10 g/L. Compared with the control, the biomass, lipid concentration and lipid content of R. glutinis increased 21.5%, 171% and 122% respectively when acetic acid concentration was 10 g/L. Furthermore, R. glutinis could accumulate lipid with acetate as the sole carbon source. Lipid concentration and lipid yield reached 3.20 g/L and 13% respectively with the initial acetic acid concentration of 25 g/L. The lipid composition was analyzed by gas chromatograph. The main composition of lipid produced with acetic acid was palmitic acid, stearic acid, oleic acid, linoleic acid and linolenic acid, including 40.9% saturated fatty acids and 59.1% unsaturated fatty acids. The lipid composition was similar to that of plant oil, indicating that lipid from oleaginous yeast R. glutinis had potential as the feedstock of biodiesel production. These results demonstrated that a certain concentration of acetic acid need not to be removed in the detoxification process when using lignocelluloses hydrolysate to produce microbial lipid by R. glutinis.
Acetic Acid
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Biofuels
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Biomass
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Culture Media
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Fatty Acids
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Hydrolysis
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Industrial Microbiology
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Lignin
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chemistry
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Linoleic Acid
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Lipids
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biosynthesis
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Oleic Acid
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Rhodotorula
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metabolism
6.Effect of peroxisome proliferator-activated receptors activators on plasminogen activator inhibitor-1 expression in HepG-2 cells.
Yan-Li HE ; Xin ZHOU ; Ping YE ; Hong FANG ; Yong-Xue LIU ; Cheng-Hua LUO ; Qiong WANG
Chinese Journal of Applied Physiology 2003;19(3):298-301
AIMTo investigate the effect of different peroxisome proliferator-activated receptors (PPARs) activators on plasminogen activator inhibitor-1 in HepG-2 cell line and explore the effect of PPARs on PAL-1 gene expression.
METHODSStearic acid, oleic acid, linoleic acid, fenofibrate, pioglitazone were used in the treatment of HepG-2 cell culture. The level of PAI-1 and PPARs mRNA was measured by reverse transcription-polymerase chain reaction (RT-PCR) and the level of PAI-1 activity and PPARs protein was determined by colorimetric assay and western blotting respectively.
RESULTSThe mRNA and activity of PAI-1 significantly increased in the groups of oleic acid and linoleic acid compared with the control, but decreased in the group of fenofibrate. There were no significant changes in both groups of stearic acid and pioglitazone. The alterations in the level of PPARs mRNA and protein were not detected in all the treated groups compared with the control.
CONCLUSIONPeroxisome proliferator-activated receptors activators play important roles in the PAI-1 gene expression and regulation. It is likely mediated by the activation of PPARalpha, but there might be other mechanisms.
Fenofibrate ; pharmacology ; Hep G2 Cells ; Humans ; Linoleic Acid ; pharmacology ; Oleic Acid ; pharmacology ; Peroxisome Proliferator-Activated Receptors ; agonists ; metabolism ; Plasminogen Activator Inhibitor 1 ; genetics ; metabolism ; RNA, Messenger ; genetics
7.Changes of carbon monoxide, nitric oxide levels and heme oxygenase system in acute respiratory distress syndrome induced by oleic acid.
He-Liang LIU ; Jin-Yuan ZHAO ; Li CHEN
Chinese Journal of Preventive Medicine 2004;38(4):240-243
OBJECTIVETo investigate possible role of carbon monoxide (CO) and heme oxygenase (HO) in the pathogenesis of acute respiratory distress syndrome (ARDS) induced by oleic acid (OA) and to compared with that induced by nitric oxide (NO).
METHODSARDS model was established in rats by oleic acid injection and concentrations of CO and NO in pulmonary arterial, carotid jugular blood and bronchoalveolar lavage fluid (BALF) were measured sequentially. Immunohistochemical method was used to determine the expression of HO in the lung.
RESULTSPulmonary arterial pressure in ARDS rats elevated 10 min after OA injection [(13.80 +/- 1.87) mm Hg to (19.51 +/- 5.02) mm Hg]. At 0.5 h after OA injection, concentration of CO in pulmonary artery began to increase and was markedly higher at 2 h than that in control rats [(0.135 +/- 0.010) g/L versus (0.116 +/- 0.005) g/L] (P < 0.01), also higher than that in carotid artery [(0.117 +/- 0.013) g/L] and in jugular vein [(0.107 +/- 0.018) g/L] in the same group, and maintained at a relatively high level thereafter. Concentration of CO in BALF also increased at 0.5 - 24 h and diminished at 72 h, as compared with that in controls. Concentration of NO in blood of pulmonary and systemic circulation all elevated markedly at 0.5 h and 2 h after OA injection, and then declined to normal at 12 h. Concentration of NO in BALF was significantly higher than that in controls. Arterial blood gas analysis showed that PaO2 markedly decreased in ARDS rats, especially at 2 h after OA injection. HO-2 could be expressed in the lung tissues of normal rats with immunohistochemical method, the strongest in epithelial cells of the bronchi, and HO-1 could only be expressed in pulmonary blood vessel walls, bronchial epithelial cells, alveolar epithelial cells and inflammatory cells of ARDS rats, lasting for 72 h after OA injection, consistent with that of CO level.
CONCLUSIONARDS rats showed a lastecl increase of CO level in pulmonary blood circulation, suggesting CO/HO system might play a more important role in modulation of blood vessel tension than NO might do in pathogenesis of ARDS.
Animals ; Carbon Monoxide ; metabolism ; Heme Oxygenase (Decyclizing) ; metabolism ; Male ; Nitric Oxide ; metabolism ; Oleic Acid ; Rats ; Rats, Sprague-Dawley ; Respiratory Distress Syndrome, Adult ; metabolism
8.Changes in liquid clearance of alveolar epithelium after oleic acid-induced acute lung injury in rats.
Jun TAO ; Tian-de YANG ; Xiang-rui CHEN ; He HUANG
Chinese Journal of Traumatology 2004;7(2):122-126
OBJECTIVEImpaired active fluid transport of alveolar epithelium may involve in the pathogenesis and resolution of alveolar edema. The objective of this study was to explore the changes in alveolar epithelial liquid clearance during lung edema following acute lung injury induced by oleic acid.
METHODSForty-eight Wistar rats were randomly divided into six groups, i.e., injured, amiloride, ouabain, amiloride plus ouabain and terbutaline groups. Twenty-four hours after the induction of acute lung injury by intravenous oleic acid (0.25 ml/kg), 5% albumin solution with 1.5 microCi (125)I-labeled albumin (5 ml/kg) was delivered into both lungs via trachea. Alveolar liquid clearance (ALC), extravascular lung water (EVLW) content and arterial blood gases were measured one hour thereafter.
RESULTSAt 24 h after the infusion of oleic acid, the rats developed pulmonary edema and severe hypoxemia, with EVLW increased by 47.9% and ALC decreased by 49.2%. Addition of either 2x10(-3) M amiloride or 5x10(-4) M ouabain to the instillation further reduced ALC and increased EVLW. ALC increased by approximately 63.7% and EVLW decreased by 46.9% with improved hypoxemia in the Terbutaline (10(-4) M) group, compared those in injured rats. A significant negative correlation was found between the increment of EVLW and the reduction of ALC.
CONCLUSIONSActive fluid transport of alveolar epithelium might play a role in the pathogenesis of lung edema in acute lung injury.
Adrenergic beta-Agonists ; pharmacology ; Animals ; Epithelium ; metabolism ; Oleic Acid ; adverse effects ; Pulmonary Alveoli ; metabolism ; Random Allocation ; Rats ; Rats, Wistar ; Respiratory Distress Syndrome, Adult ; chemically induced ; metabolism ; Terbutaline ; pharmacology
9.Effect of ulinastain on the expression of hemeoxygenase-1 in oleic acid-induced acute lung injury in rats.
Li-Yan JIN ; Jun-Mei XU ; Zhi-Biao HE ; Wen-Yan RUAN ; Xiang-Ping CHAI
Journal of Central South University(Medical Sciences) 2007;32(4):675-678
OBJECTIVE:
To explore the effect of ulinastain on the expression of hemeoxy genase-1 (HO-1) in oil acid-induced acute lung injury in rats.
METHODS:
The animal model of acute lung injury was established by oil acid. Thirty SD rats were randomly divided into 3 groups: the blank control group (A), the acute lung injury group (B) and the acute lung injury group (C) followed by injecting 100 mL/kg ulinastatin. Each group consisted of 10 rats. Group A were given 0.2 mL/kg natural solution through the trial vein; Group B and C were given 0.2 mL/kg oil-acid through trial vein, while group C were injected 100mL/kg ulinastatin by the peritoneal cavity after injecting oil acid. After 4 hours, the rates of respiration were counted and blood samples were cramped out through the heart puncture for blood gas analysis. The expressions of hemeoxygenase-1 and the pathologic construction changes were determined by HE staining in the lower right lung of rats in the 3 groups.
RESULTS:
The respiration dysfunction caused by oil acid could be prominently improved by ulinastain. There was only a little expression of hemeoxygenase-1 in the lung of Group A, but the expression increased in Group B and significatively increased in Group C.
CONCLUSION
Ulinastatin may protect the rats from acute lung injury through increasing the expression of HO-1.
Acute Lung Injury
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chemically induced
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metabolism
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Animals
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Glycoproteins
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pharmacology
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Heme Oxygenase (Decyclizing)
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metabolism
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Lung
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drug effects
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metabolism
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Male
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Oleic Acid
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adverse effects
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Rats
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Rats, Sprague-Dawley
10.Identification and functional analysis of soybean stearoyl-ACP Δ⁹ desaturase (GmSAD) gene family.
Mimi DENG ; Baoling LIU ; Zhilong WANG ; Jin'ai XUE ; Hongmei ZHANG ; Runzhi LI
Chinese Journal of Biotechnology 2020;36(4):716-731
Stearoyl-ACP Δ⁹ desaturase (SAD) catalyzes the synthesis of monounsaturated oleic acid or palmitoleic acid in plastids. SAD is the key enzyme to control the ratio of saturated fatty acids to unsaturated fatty acids in plant cells. In order to analyze the regulation mechanism of soybean oleic acid synthesis, soybean (Glycine max) GmSAD family members were genome-wide identified, and their conserved functional domains and physicochemical properties were also analyzed by bioinformatics tools. The spatiotemporal expression profile of each member of GmSADs was detected by qRT-PCR. The expression vectors of GmSAD5 were constructed. The enzyme activity and biological function of GmSAD5 were examined by Agrobacterium-mediated transient expression in Nicotiana tabacum leaves and genetic transformation of oleic acid-deficient yeast (Saccharomyces cerevisiae) mutant BY4389. Results show that the soybean genome contains five GmSAD family members, all encoding an enzyme protein with diiron center and two conservative histidine enrichment motifs (EENRHG and DEKRHE) specific to SAD enzymes. The active enzyme protein was predicted as a homodimer. Phylogenetic analysis indicated that five GmSADs were divided into two subgroups, which were closely related to AtSSI2 and AtSAD6, respectively. The expression profiles of GmSAD members were significantly different in soybean roots, stems, leaves, flowers, and seeds at different developmental stages. Among them, GmSAD5 expressed highly in the middle and late stages of developmental seeds, which coincided with the oil accumulation period. Transient expression of GmSAD5 in tobacco leaves increased the oleic acid and total oil content in leaf tissue by 5.56% and 2.73%, respectively, while stearic acid content was reduced by 2.46%. Functional complementation assay in defective yeast strain BY4389 demonstrated that overexpression of GmSAD5 was able to restore the synthesis of monounsaturated oleic acid, resulting in high oil accumulation. Taken together, soybean GmSAD5 has strong selectivity to stearic acid substrates and can efficiently catalyze the biosynthesis of monounsaturated oleic acid. It lays the foundation for the study of soybean seed oleic acid and total oil accumulation mechanism, providing an excellent target for genetic improvement of oil quality in soybean.
Fatty Acid Desaturases
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genetics
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metabolism
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Gene Expression Profiling
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Oleic Acid
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biosynthesis
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Phylogeny
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Plant Proteins
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genetics
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Seeds
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chemistry
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Soybeans
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classification
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enzymology
;
genetics