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Proteomics Insights into the Gene Network of cis9, trans11-Conjugated Linoleic Acid Biosynthesis in Bovine Mammary Gland Epithelial Cells.

ABSTRACT: The objective of the study was to elucidate the stearoyl-coenzyme A desaturase (SCD1)-dependent gene network of c9, t11-CLA biosynthesis in MAC-T cells from an energy metabolism perspective. The cells were divided into the CAY group (firstly incubated with CAY10566, a chemical inhibitor of SCD1, then incubated with trans-11-octadecenoic acid, (TVA)), the TVA group (only TVA), and the control group (without CAY, TVA). The c9, t11-CLA, and TVA contents were determined by gas chromatography. The mRNA levels of SCD1 and candidate genes were analyzed via real-time PCR. Tandem mass tag (TMT)-based quantitative proteomics, bioinformatic analysis, parallel reaction monitoring (PRM), and small RNA interference were used to explore genes involved in the SCD1-dependent c9, t11-CLA biosynthesis. The results showed that the SCD1 deficiency led by CAY10566 blocked the biosynthesis of c9, t11-CLA. In total, 60 SCD1-related proteins mainly involved in energy metabolism pathways were primarily screened by TMT-based quantitative proteomics analysis. Moreover, 17 proteins were validated using PRM analysis. Then, 11 genes were verified to have negative relationships with SCD1 after the small RNA interference analysis. Based on the above results, we concluded that genes involved in energy metabolism pathways have an impact on the SCD1-dependent molecular mechanism of c9, t11-CLA biosynthesis.

SUBMITTER: Peng L

PROVIDER: S-EPMC9264836 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundDue to high neutral lipids accumulation in the cytoplasm, in vitro-produced embryos from Bos primigenius indicus and their crosses are more sensitive to chilling and cryopreservation than those from Bos primigenius taurus. The objective of the present study was to evaluate the effects of trans-10, cis-12 conjugated linoleic acid (CLA) on the development and cryotolerance of crossbred Bos primigenius taurus x Bos primigenius indicus embryos produced in vitro, and cultured in the presence of fetal calf serum. Bovine zygotes (n = 1,692) were randomly assigned to one of the following treatment groups: 1) Control, zygotes cultured in Charles Rosenkrans 2 amino acid (CR2aa) medium (n = 815) or 2) CLA, zygotes cultured in CR2aa medium supplemented with 100 μmol/L of trans-10, cis-12 CLA (n = 877). Embryo development (cleavage and blastocyst rates evaluated at days 3 and 8 of culture, respectively), lipid content at morula stage (day 5) and blastocyst cryotolerance (re-expansion and hatching rates, evaluated 24 and 72 h post-thawing, respectively) were compared between groups. Additionally, selected mRNA transcripts were measured by Real-Time PCR in blastocyst stage.ResultsThe CLA treatment had no effect on cleavage and blastocyst rates, or on mRNA levels for genes related to cellular stress and apoptosis. On the other hand, abundance of mRNA for the 1-acylglycerol-3-phosphate 0-acyltransferase-encoding gene (AGPAT), which is involved in triglycerides synthesis, and consequently neutral lipid content, were reduced by CLA treatment. A significant increase was observed in the re-expansion rate of embryos cultured with trans-10, cis-12 CLA when compared to control (56.3 vs. 34.4%, respectively, P = 0.002). However, this difference was not observed in the hatching rate (16.5 vs. 14.0%, respectively, P = 0.62).ConclusionsThe supplementation with trans-10, cis-12 CLA isomer in culture medium reduced the lipid content of in vitro produced bovine embryos by reducing the gene expression of 1-acylglycerol 3-phosphate 0-acyltransferase (AGPAT) enzyme. However, a possible improvement in embryo cryotolerance in response to CLA, as suggested by increased blastocyst re-expansion rate, was not confirmed by hatching rates.

Project description:BackgroundConjugated linoleic acid (cis-9, trans-11 CLA) and trans-11 vaccenic acid (VA) are found naturally in ruminant-derived foods. CLA has been shown to have numerous potential health related effects and has been extensively investigated. More recently, we have shown that VA has lipid-lowering properties associated with reduced hepatic lipidogenesis and chylomicron secretion in the JCR:LA-cp rat. The aim of this study was to evaluate potential additional hypolipidemic effects of purified forms of CLA and VA in an animal model of the metabolic syndrome (the JCR:LA-cp rat).MethodsTwenty four obese JCR:LA-cp rats were randomized and assigned to one of three nutritionally adequate iso-caloric diets containing 1% w/w cholesterol and 15% w/w fat for 16 wk: 1) control diet (CD), 2) 1.0% w/w cis-9, trans-11 CLA (CLA), 3) 1.0% w/w VA and 1% w/w cis-9, trans-11 CLA (VA+CLA). Lean rats were fed the CD to represent normolipidemic conditions.ResultsFasting plasma triglyceride (TG), total cholesterol and LDL-cholesterol concentrations were reduced in obese rats fed either the CLA diet or the VA+CLA diet as compared to the obese control group (p < 0.05, p < 0.001; p < 0.001, p < 0.01; p < 0.01, p < 0.001, respectively). The VA+CLA diet reduced plasma TG and LDL-cholesterol to the level of the normolipidemic lean rats and further decreased nonesterified fatty acids compared to the CLA diet alone. Interestingly, rats fed the VA+CLA diet had a higher food intake but lower body weight than the CLA fed group (P < 0.05). Liver weight and TG content were lower in rats fed either CLA (p < 0.05) or VA+CLA diets (p < 0.001) compared to obese control, consistent with a decreased relative protein abundance of hepatic acetyl-CoA carboxylase in both treatment groups (P < 0.01). The activity of citrate synthase was increased in liver and adipose tissue of rats fed, CLA and VA+CLA diets (p < 0.001) compared to obese control, suggesting increased mitochondrial fatty acid oxidative capacity.ConclusionWe demonstrate that the hypolipidemic effects of chronic cis-9, trans-11 CLA supplementation on circulating dyslipidemia and hepatic steatosis are enhanced by the addition of VA in the JCR:LA-cp rat.

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Proteomics Insights into the Gene Network of cis9, trans11-Conjugated Linoleic Acid Biosynthesis in Bovine Mammary Gland Epithelial Cells.

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