Conjugated Linoleic Acid Reduces Phorbol Ester-Induced Prostaglandin F2 Production by Bovine Endometrial Cells (original) (raw)
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Journal of Dairy Science, 2006
Recent studies have implicated n-3 polyunsaturated fatty acids in the reduction of eicosanoid production in the bovine uterus. The objective of this study was to determine whether the effect of eicosapentaenoic acid (EPA; C 20:5 , n-3) on PGF 2α production by bovine endometrial (BEND) cells is influenced by the quantity of linoleic acid (C 18:2 , n-6) in the incubation medium. Confluent BEND cells were incubated in the absence (control) or presence of 100 M of EPA for 24 h. After incubation, cells were rinsed and then stimulated with phorbol 12,13-dibutyrate (PDBu; 100 ng/mL) for 6 h. Additional sets of culture dishes were treated with a combination of EPA and increasing n-6/n-3 fatty acid ratios for 24 h and then challenged with PDBu for 6 h. The PDBu stimulated PGF 2α secretion and upregulated steadystate concentrations of prostaglandin endoperoxide synthase-2 and peroxisome proliferator-activated receptor delta mRNA within 6 h. Preincubation of BEND cells with EPA for 24 h decreased PGF 2α response to phorbol ester, but had no detectable effects on prostaglandin endoperoxide synthase-2 or peroxisome proliferator-activated receptor delta mRNA abundance in PDBu-stimulated BEND cells. The inhibitory effect of EPA on PGF 2α production was reverted in BEND cells treated with an increasing n-6-ton -3 fatty acid ratio. Findings indicate that the net inhibition of endometrial PGF 2α bioynthesis by n-3 fatty acids may vary depending on the ratio of n-6 to n-3 fatty acids in the uterus.
American Journal of Veterinary Research, 2013
Objective-To determine the effects of 2 conjugated linoleic acid (CLA) isomers (cis-9, trans-11 and trans-10, cis-12) on synthesis of prostaglandin (PG) E 2 and F 2α and expression of prostaglandin H synthase-2 (PGHS-2) of adult and fetal bovine endometrial epithelial cells in vitro. Sample-Primary cultures of endometrial epithelial cells obtained from 4 adult cows and 4 fetal bovine carcasses. Procedures-Cells were exposed to 0, 50, 100, or 200µM cis-9, trans-11 or trans-10, cis-12 CLA isomers for 24 hours. Culture media collected before and after 6 hours of stimulation of cells with phorbol 12-myristate 13-acetate were assayed to detect PGE 2 and PGF 2α via ELISA. After stimulation, cells were collected for western blot analysis to quantify PGHS-2. Results-Concentrations of PGF 2α and PGE 2 were significantly lower in culture media of adult and fetal endometrial epithelial cells exposed to any concentration of either CLA than they were in media of cells not exposed to CLAs. The trans-10, cis-12 CLA isomer seemed to decrease PG production more markedly than did the cis-9, trans-11 CLA isomer. Most concentrations of both CLAs significantly reduced culture media PGE 2 :PGF 2α concentration ratios of cells. Exposure of cells to CLAs did not affect expression of PGHS-2 protein.
Journal of Equine Veterinary Science, 2009
Increased secretion of prostaglandin F 2 ␣ (PGF 2 ␣) within the uterus because of uterine inflammation can cause luteolysis and result in early embryonic loss. Supplementation with polyunsaturated fatty acids (PUFAs) has been shown to influence PG production in many species, although the effects on the mare remain unknown. The present study aimed to determine fatty acid uptake in equine endometrial explants and evaluate their influence on PG secretion and expression of enzymes involved in PG synthesis in vitro. Equine endometrial explants were treated with 100 M arachidonic acid, eicosapentaenoic acid, or docosahexaenoic acid and then challenged with oxytocin (250 nM) or lipopolysaccharide (LPS; 1 g/mL). Production of PGF 2 ␣ and PG E 2 (PGE 2 ) was measured, and mRNA expression of enzymes involved in PG synthesis was determined with quantitative real-time PCR. Media concentrations of PGF 2 ␣ and PGE 2 were higher (P Ͻ 0.0001) from endometrial explants challenged with oxytocin or LPS compared with controls despite which fatty acid was added. Only DHA lowered (P Ͻ 0.0001) media concentrations of PGF 2 ␣ and PGE 2 from explants. Endometrial explants stimulated with oxytocin had increased expression of PG-endoperoxide synthase 1 (PTGS1; P Ͻ 0.02), PG-endoperoxide synthase 2 (PTGS2; P Ͻ 0.001), PG F 2 ␣ synthase (PGFS; P Ͻ 0.01), PG E 2 synthase (PGES; P Ͻ 0.01), and phospholipase A 2 (PLA 2 ; P Ͻ 0.005) compared with controls and regardless of fatty acid treatment; whereas stimulation with LPS increased expression of PTGS2 (P Ͻ 0.004), PGFS (P Ͻ 0.03), PGES (P Ͻ 0.01), and PLA 2 (P Ͻ 0.01) compared with controls and regardless of fatty acid treatment. Treatment with PUFAs, specifically DHA, can influence PG secretion in vitro through mechanisms other than enzyme expression.
Prostaglandins & Other Lipid Mediators, 1998
A diet rich in n-3 polyunsaturated fatty acid (PUFA) may reduce the intrauterine production of prostaglandins and prolong pregnancy. We tested this hypothesis by assessing the influence of various PUFAs on the spontaneous production of PGE 2 and PGF 2␣ from decidual cell cultures. In addition, we assessed prostaglandin and cytokine production stimulated by lipopolysaccharides (LPS) in order to mimic parturition where infection is involved.
International Journal of Cancer, 2007
Conjugated linoleic acid (CLA) is a naturally occurring fatty acid, which has been shown to exert beneficial effects against breast carcinogenesis. It has been reported that CLA could modulate cellular proliferation and differentiation through the activation of peroxisome proliferator-activated receptors (PPARs). Among different PPAR isotypes, PPARc is involved in growth inhibition of transformed cells. Ligands of PPARc are considered as potential anticancer drugs, so CLA was tested for its ability to induce PPARc expression in MCF-7 breast cancer cells. The effects of CLA and of a specific synthetic PPARc antagonist were evaluated on cell growth as well as on parameters responsible for cell growth regulation. We demonstrated here that CLA stimulated the expression of PPARc to levels up to control and caused PPARc translocation into the nucleus. Furthermore, the overexpression of PPARc positively correlates with the inhibition of cell proliferation and with the modulation of ERK signaling induced by CLA; in all cases the administration of the antagonist reverted CLA effects. The PPAR-signaling pathway is connected with the b-catenin/Ecadherin pathway, thus we evaluated CLA effects on the expression and cellular distribution of these proteins, which are involved in cell adhesion and responsible for invasive behavior. The treatment with CLA determined the up-regulation and the redistribution of b-catenin and E-cadherin and the antagonist reverted only the effect on b-catenin. These studies indicate that CLA regulates PPARc expression by selectively acting as an agonist and may influence cell-cell adhesion and invasiveness of MCF-7 cells.
Theriogenology, 2019
Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor family of ligand-dependent transcription factors. PPARs are important regulators of glucose and fatty acid metabolism, apoptosis, angiogenesis, cell proliferation and differentiation, and immune response. Their possible role in the female reproductive tract was demonstrated. In the present study, cultured luminal epithelial (LE) and stromal (ST) cells of the porcine endometrium were used to examine (1) the effect of conceptus exposed medium (CEM) on mRNA and protein expression and DNA binding activity of PPARA, PPARD, and PPARG isoforms, and (2) the effect of PPARA, PPARD, and PPARG agonists on the expression of selected genes, apoptosis, and cell proliferation. The addition of CEM stimulated PPARA expression and DNA binding activity of this isoform in LE and ST cells (P < 0.05). Increased expression of PPARD mRNA in the presence of CEM was detected in ST cells (P < 0.05), while the concentration of PPARG transcripts decreased in response to CEM in both cell types (P < 0.05). LE and ST cells of the pig endometrium possess PPARA, PPARD, and PPARG proteins, with clear nuclear staining visible predominately in ST cells. In LE cells, activation of PPARG with 15-deoxy-D12,14-prostaglandin(PG)J2 down-regulated the expression of genes encoding amino acid transporter 1 (SLC38A1), leukemia inhibitory factor (LIF) and enzymes involved in PG synthesis (P < 0.05). In ST cells, activation of PPARD isoform with both agonists used (L-165,041 and cPGI2) and PPARG isoform with 15-deoxy-D12,14-PGJ2 increased vascular endothelial growth factor A (VEGFA) mRNA expression (P < 0.05). Moreover, GW9578 (PPARA agonist) and 15-deoxy-D12,14-PGJ2 stimulated glucose transporter 1 (SLC2A1) gene expression in ST cells. 15-deoxy-D12,14-PGJ2 was also effective in up-regulation of the ratio of BAX/BCL2 mRNA expression and active caspase-3 concentration in ST cells (P < 0.05). Finally, GW9578 stimulated LE and ST cell proliferation, while rosiglitazone (PPARG agonist) increased the number of viable ST but not LE cells. In conclusion, this study demonstrated that conceptus products differentially modulate PPARs expression and activity in the porcine endometrium. Activation of PPARs may in turn affect nutrient transport, PG synthesis, angiogenesis, apoptosis, or cell proliferation in this tissue. Therefore, PPAR isoforms seem to play an important role in development and function of the porcine uterus.
Conjugated linoleic acid is a potent naturally occurring ligand and activator of PPARα
Journal of Lipid Research, 1999
We have previously shown that a mixture of dietary conjugated derivatives of linoleic acid (conjugated linoleic acid, CLA) induces peroxisome proliferator-responsive enzymes and modulates hepatic lipid metabolism in vivo. The present studies demonstrate that CLA is a high affinity ligand and activator of peroxisome proliferator-activated receptor ␣ (PPAR ␣) and induces accumulation of PPARresponsive mRNAs in a rat hepatoma cell line. Using a scintillation proximity assay (SPA), CLA isomers were shown to be ligands for human PPAR ␣ with a rank order of potency of (9Z,11E) Ͼ (10E,12Z) Ͼ (9E,11E) Ͼ furan-CLA (IC 50 values from 140 n M to 400 n M). Levels of acyl-CoA oxidase (ACO), liver fatty acid-binding protein (L-FABP), and cytochrome P450IVA1 (CYP4A1) mRNA were induced by CLA in FaO hepatoma cells. Even though linoleate and CLA were incorporated into lipids of hepatoma cells to the same extent, linoleate had little or no effect on ACO, CYP4A1, or L-FABP mRNA. In agreement with its binding potency, (9Z,11E)-CLA was the most efficacious PPAR ␣ activator in the mouse PPAR ␣-GAL4(UAS) 5-CAT reporter system. These data indicate that CLA is a ligand and activator of PPAR ␣ and its effects on lipid metabolism may be attributed to transcriptional events associated with this nuclear receptor. Also, (9Z,11E)-CLA is one of the most avid fatty acids yet described as a PPAR ␣ ligand.-Moya-Camarena, S.
Journal of Endocrinology, 2004
We investigated the effect of n-6 polyunsaturated fatty acids (PUFAs) on prostaglandin (PG) production by the uterus. A mixed population of endometrial cells (epthelium and stroma) from late-gestation ewes were cultured in defined medium containing linoleic acid (LA, 18:2, n-6), -linolenic acid (GLA, 18:3, or arachidonic acid (AA, 20:4, in concentrations of 0 (control), 20 or 100 µM. After 45 h in test medium with or without added PUFAs, cells were challenged with control medium (CM), oxytocin (OT, 250 nM), lipopolysaccharide (LPS, 0·1 µg/ml) or dexamethasone (DEX, 5 µM) for 22 h in the continued presence of the same concentration of PUFA and the medium was collected for measurement of PGF 2 and PGE 2 . Supplementation with LA inhibited the production of PGF 2 but did not alter PGE 2 , whereas GLA and AA increased production of both PGs. All PUFA supplements thus increased the ratio of PGE 2 to PGF 2 (E:F ratio) two-to threefold. In control cells, OT and LPS challenges stimulated the production of PGF 2 and PGE 2 . In all challenge groups, the concentrations of PGF 2 in response to PUFAs followed the same pattern -LA<control<GLA<AA -but there were significant alterations in responsiveness as a result of PUFA treatment. In the cells supplemented with 100 µM AA, there was no further increase in PGF 2 output in the presence of OT or LPS and when 100 µM GLA was present neither LPS nor OT stimulated PGE 2 significantly. When LPS was given to AA-supplemented cells, the E:F ratio was increased. DEX did not change PGE 2 production in control or LA-treated cells, but the cells produced significantly less PGF 2 , so the E:F ratio was increased. In contrast, in GLA-and AA-treated cells, DEX reduced the production of both PGF 2 and PGE 2 , so the E:F ratio was unaltered. In summary, the study showed altered production of PGs in the presence of different PUFAs according to their position in the n-6 metabolic pathway. The type of PUFA present affected responsiveness to OT, LPS and DEX and also changed the ratio of PGE 2 to PGF 2 produced. The possible implications of this work are discussed in relation to the effect of diet on term and pre-term labour, which both require upregulation of the endometrial PG synthetic pathway.
Journal of Dairy Science, 2009
Studying long-chain fatty acid (LCFA) effects on gene network expression in bovine cells could provide useful information for future practical applications. An optimized in vitro system that does not require tissue collection or cell isolation could fill a niche in the study of PPARα activity in ruminants. Specific aims were to optimize culture conditions in Madin-Darby bovine kidney (MDBK) cells to achieve maximal mRNA expression of known peroxisome proliferator-activated receptor-α (PPARα) target genes using palmitate (16:0) as a representative LCFA. Variables included length of incubation time, use of albumin-bound (4:1 molar proportion) 16:0 (A16:0), or addition of insulin. A first time-course experiment tested culturing cells in Dulbecco's modified Eagle's medium with 150 μM PPAR ligand Wy-14643 (WY) and A16:0. A second experiment tested the effects of albumin and insulin using 150 μM of 16:0 without albumin or insulin (−Alb/−Ins), 16:0 without albumin plus 5 mg/L of bovine insulin (−Alb/+Ins), A16:0 without insulin (+Alb/−Ins), or a control. A third experiment was a preliminary metabolic characterization of cells and assessed intracellular lipid droplet formation after treatment with 150 μM of 16:0 or an ethanol control. For all experiments, cells were harvested at 0, 6, 12, 18, and 24 h posttreatment. In experiments 1 and 2, mRNA expression was assessed by quantitative PCR of selected PPARα target genes as well as PPARα coactivators (ACOX1, CPT1A, ACADVL, ACSL1, PPARA, PPARGC1A, LPIN1). In experiment 1, there was a linear increase in mRNA expression of CPT1A (~500%) and ACSL1 (50 to 200%) by 6 h of incubation with both WY and A16:0. The LPIN1 mRNA increased by >100% by 6 h only with A16:0. Further, there was a linear increase in expression of PPARA (~100%) with A16:0 through 24 h of incubation. In experiment 2, insulin increased, and coupling LCFA with albumin tended to delay the response in expression of CPT1A and ACSL1 to 16:0. Data indicated a toxic effect of 150 μM free 16:0 as assessed by cell counts after 12 h of incubation. In experiment 3, MDBK cells appeared to use glucose and AA as energy sources and were able to secrete triglycerides. In addition, MDBK cells cultured with 150 μM of 16:0 had a substantial uptake of LCFA and synthesized intracellular lipid droplets. Overall, results indicated that a 6-h incubation with free LCFA and addition of insulin was suitable to detect marked effects on mRNA expression of PPARα target genes in MDBK cells.
Journal of dairy science, 2009
Studying long-chain fatty acid (LCFA) effects on gene network expression in bovine cells could provide useful information for future practical applications. An optimized in vitro system that does not require tissue collection or cell isolation could fill a niche in the study of PPARalpha activity in ruminants. Specific aims were to optimize culture conditions in Madin-Darby bovine kidney (MDBK) cells to achieve maximal mRNA expression of known peroxisome proliferator-activated receptor-alpha (PPARalpha) target genes using palmitate (16:0) as a representative LCFA. Variables included length of incubation time, use of albumin-bound (4:1 molar proportion) 16:0 (A16:0), or addition of insulin. A first time-course experiment tested culturing cells in Dulbecco's modified Eagle's medium with 150 microM PPAR ligand Wy-14643 (WY) and A16:0. A second experiment tested the effects of albumin and insulin using 150 microM of 16:0 without albumin or insulin (-Alb/-Ins), 16:0 without albumin plus 5 mg/L of bovine insulin (-Alb/+Ins), A16:0 without insulin (+Alb/-Ins), or a control. A third experiment was a preliminary metabolic characterization of cells and assessed intracellular lipid droplet formation after treatment with 150 microM of 16:0 or an ethanol control. For all experiments, cells were harvested at 0, 6, 12, 18, and 24 h posttreatment. In experiments 1 and 2, mRNA expression was assessed by quantitative PCR of selected PPARalpha target genes as well as PPARalpha coactivators (ACOX1, CPT1A, ACADVL, ACSL1, PPARA, PPARGC1A, LPIN1). In experiment 1, there was a linear increase in mRNA expression of CPT1A (approximately 500%) and ACSL1 (50 to 200%) by 6 h of incubation with both WY and A16:0. The LPIN1 mRNA increased by >100% by 6 h only with A16:0. Further, there was a linear increase in expression of PPARA (approximately 100%) with A16:0 through 24 h of incubation. In experiment 2, insulin increased, and coupling LCFA with albumin tended to delay the response in expression of CPT1A and ACSL1 to 16:0. Data indicated a toxic effect of 150 microM free 16:0 as assessed by cell counts after 12 h of incubation. In experiment 3, MDBK cells appeared to use glucose and AA as energy sources and were able to secrete triglycerides. In addition, MDBK cells cultured with 150 microM of 16:0 had a substantial uptake of LCFA and synthesized intracellular lipid droplets. Overall, results indicated that a 6-h incubation with free LCFA and addition of insulin was suitable to detect marked effects on mRNA expression of PPARalpha target genes in MDBK cells.