Cis-12 Cis-12 Conjugated Linoleic Acid Research Articles

Rumen microbiota-host transcriptome interaction mediates the protective effects of trans-10, cis-12 CLA on facilitating weaning transition of lambs.

Developing alternatives to antibiotics for prevention of gastrointestinal dysbiosis in early-weaning farmed animals is urgently needed. This study was to explore the potential effects of trans-10, cis-12 conjugated linoleic acid (CLA) on maintaining ruminal homeostasis of young ruminants during the weaning transition period. Thirty neonatal lambs were selected (6 lambs per group) and euthanized for rumen microbial and epithelial analysis. The lambs were weaned at 28d and experienced the following 5 treatments: euthanized on d 28 as the pre-weaning control (CON0), fed starter feed for 5 (CON5) or 21 (CON21) d, fed starter feed with 1% of CLA supplemented for 5 (CLA5) or 21 (CLA21) d. Results showed that the average daily weight gain and dry matter intake were significantly higher in CLA5 than CON5 group. As compared with the CON5 and CON21 group, the relative abundances of volatile fatty acid (VFA) producing bacteria including Bacteroides, Treponema, Parabacteroides and Anaerovibrio, as well as the concentrations of acetate, butyrate and total VFA were significantly increased in CLA5 and CLA21 group, respectively. Integrating microbial profiling and epithelial transcriptome results showed that 7 downregulated inflammatory signaling-related host genes IL2RA, CXCL9, CD4, CCR4, LTB, SPP1, and BCL2A1 with CLA supplementation were significantly negatively correlated with both VFA concentration and VFA producing bacteria, while 3 (GPX2, SLC27A2 and ALDH3A1) and 2 (GSTM3 and GSTA1) upregulated metabolism-related genes, significantly positively correlated with either VFA concentration or VFA producing bacteria, respectively. To confirm the effects of CLA on epithelial signal transduction, invitro experiment was further conducted by treating rumen epithelial cells without or with IL-17A+TNF-α for 12h after pretreatment of 100μM CLA or not (6 replicates per treatment). The results demonstrated the anti-inflammatory effect of CLA via suppressing the protein expression of NF-кB p-p65/p65 with the activation of peroxisome proliferator-activated receptor gamma (PPARγ). In conclusion, CLA supplementation enhanced the ruminal microbiota-driven transcriptional regulation in healthy rumen epithelial development via rumen VFA production, and CLA may therefore serve as an alternative way to alleviate early-weaning stress and improve physiological and metabolic conditions of young ruminants.

Stearic acid does not overcome conjugated linoleic acid trans-10, cis-12-induced milk fat depression in lactating ewes.

The objective of this study was to test the hypothesis that stearic acid (SA) supplementation increases milk fat content and overcomes the antilipogenic effects of trans-10, cis-12 conjugated linoleic acid (CLA) in lactating ewes. Twenty-eight Lacaune ewes (36 (sd 2) days in lactation; 70·5 (sd) 9·6 kg of body weight), producing 1·8 (sd 0·4) kg of milk/d, were used in a completely randomised design (seven ewes/treatment) for 21 d. The treatments were: (1) Control; (2) CLA (6·4 g/d of trans-10, cis-12 CLA); (3) SA (28 g/d of SA) and (4) SA in association with trans-10, cis-12 CLA (CLASA; 6·4 g/d of trans-10, cis-12 CLA plus 28 g/d of SA). All data were analysed using a mixed model that included the fixed effect of treatment and the random effect of ewe. SA did not alter milk fat content and yield relative to Control (91·9 v. 91·2 (sd 4·1) g/d). CLASA was not able to overcome the reduction in fat content and fat yield induced by CLA (75 v. 82 (sd 0·14) g/d). SA increased the relative abundance of CD36, fatty acid-binding protein 4 (FABP4) and PPAR-γ mRNA by 140, 112 and 68 % compared with CLASA. SA also reduced the relative abundance of acetyl-CoA carboxylase α promoter II and stearoyl-CoA desaturase (SCD) when compared with Control (45 and 39 %). Compared with CLA, CLASA treatment had no effect on the mRNA abundance of fatty acid synthase, lipoprotein lipase, CD36, SCD, FABP4, acylglycerolphosphate acyltransferase 6, sterol regulatory element-binding protein 1 and PPAR-γ. In conclusion, SA supplementation did not increase milk fat synthesis and did not overcome the CLA-induced milk fat depression when associated with trans-10, cis-12 CLA.

High rumen degradable starch decreased goat milk fat via trans-10, cis-12 conjugated linoleic acid-mediated downregulation of lipogenesis genes, particularly, INSIG1

BackgroundStarch is an important substance that supplies energy to ruminants. To provide sufficient energy for high-yielding dairy ruminants, they are typically fed starch-enriched diets. However, starch-enriched diets have been proven to increase the risk of milk fat depression (MFD) in dairy cows. The starch present in ruminant diets could be divided into rumen-degradable starch (RDS) and rumen escaped starch (RES) according to their different degradation sites (rumen or intestine). Goats and cows have different sensitivities to MFD. Data regarding the potential roles of RDS in milk fat synthesis in the mammary tissue of dairy goats and in regulating the occurrence of MFD are limited.ResultsEighteen Guanzhong dairy goats (day in milk = 185 ± 12 d) with similar parity, weight, and milk yield were selected and randomly assigned to one of three groups (n = 6), which were fed an LRDS diet (Low RDS = 20.52%), MRDS diet (Medium RDS = 22.15%), or HRDS diet (High RDS = 24.88%) for 5 weeks. Compared with that of the LRDS group, the milk fat contents in the MRDS and HRDS groups significantly decreased. The yields of short-, medium- and long-chain fatty acids decreased in the HRDS group. Furthermore, increased RDS significantly decreased ruminal B. fibrisolvens and Pseudobutyrivibrio abundances and increased the trans-10, cis-12 conjugated linoleic acid (CLA) and trans-10 C18:1 contents in the rumen fluid.A multiomics study revealed that the HRDS diet affected mammary lipid metabolism down-regulation of ACSS2, MVD, AGPS, SCD5, FADS2, CERCAM, SC5D, HSD17B7, HSD17B12, ATM, TP53RK, GDF1 and LOC102177400. Remarkably, the significant decrease of INSIG1, whose expression was depressed by trans-10, cis-12 CLA, could reduce the activity of SREBP and, consequently, downregulate the downstream gene expression of SREBF1.ConclusionsHRDS-induced goat MFD resulted from the downregulation of genes involved in lipogenesis, particularly, INSIG1. Specifically, even though the total starch content and the concentrate-to-fiber ratio were the same as those of the high-RDS diet, the low and medium RDS diets did not cause MFD in lactating goats.

Dietary Fat Does Not Overcome trans-10, cis-12 Conjugated Linoleic Acid Inhibition of Milk Fat Synthesis in Lactating mice.

Trans-10, cis-12 conjugated linoleic acid (CLA) is a potent inhibitor of milk fat synthesis in the cow and similarly reduces milk fat in rodents. The objective of this study was to determine whether dietary fat can overcome CLA inhibition of milk fat concentration in lactating mice. Wild type C57Bl/6J mice (n = 31) were fed semipurified diets containing either low fat (LF; 4% fat) or high fat (HF; 23.6% fat) starting 4-6 days postpartum. Dietary fat was increased by inclusion of high oleic sunflower oil. After 2 days on the experimental diets, lactating dams were orally dosed with either water (control) or trans-10, cis-12 CLA (20 mg/day) for 5 days. CLA treatment decreased pup growth similarly in both HF and LF diets. Milk fat percent was increased over 16% by the HF diet and decreased over 12% by CLA, but there was no interaction of dietary fat and CLA. Both CLA and the HF diet reduced the proportion of short- and medium-chain fatty acids that originate from de novo synthesis, and there was no interaction of diet and CLA. CLA had no effect on the percent of preformed fatty acids, but the HF diet increased their abundance. Dietary fat and CLA both modified mammary expression of lipogenic enzymes and regulators, but no interactions were observed. In conclusion, CLA reduced milk fat concentration and litter growth, but these effects were not overcome by increased dietary fat from high oleic sunflower oil. CLA inhibition of milk fat in the mammary gland is not substrate dependent, and the mechanism is independent from dietary supply of oleic acid.

Relationships between trans-10 shift indicators and milk fat traits in dairy ewes: Insights into milk fat depression

The causative relationship between ruminal biohydrogenation alterations known as the trans-10 shift and marine lipid-induced milk fat depression (MFD) has been questioned in dairy ewes, but the large intra- and inter-experiment differences in the levels of trans-10 shift indicators precludes firm conclusions. This meta-analysis was conducted to quantify the relationship between trans-10 shift milk indicators [i.e., trans-10,cis-12 conjugated linoleic acid (CLA), trans-10 18:1 or trans-10:trans-11 18:1 ratio] and lactation performance, mainly milk fat traits, in Assaf dairy ewes. A database comprising of 17 experiments including 50 dietary treatments was compiled. The diets were grouped into 2 dietary conditions: (1) non-MFD condition, which included 14 diets supplemented with plant oils and tannins that modified the milk fatty acid profile without lowering milk fat concentration and yield, and 12 respective control diets, and (2) MFD condition, which included 15 diets supplemented with marine lipids that induced the low fat-milk syndrome, and 9 respective control diets. The overall levels of milk trans-10 shift indicators did not differ between dietary conditions, but significant increments were found in the supplemented diets relative to their respective controls. A similar linear and negative (slope: −7.28 ± 2.816) relationship between milk fat concentration and trans-10,cis-12 CLA was found in both dietary conditions. However, the quadratic negative response of milk fat content to trans-10 18:1 concentration was greater in MFD than in non-MFD conditions. In the two dietary conditions, the relationship between milk trans-10,cis-12 CLA or trans-10 18:1 and de novo fatty acid concentration in milk was negative, whereas the relationship with preformed fatty acid proportion was positive. The magnitude of changes in both milk fatty acid groups in response to changes in these trans-10 shift indicators were greater in non-MFD compared with MFD conditions. Milk fat yield and trans-10 shift indicators were only negatively related in ewes that were fed marine lipid supplements, because reductions in de novo fatty acid yield with increased shift indicator levels were not counteracted by concomitant increments in preformed fatty acid secretion. The best-fit (lower root mean squared error, higher R2 and no prediction biases) equations of milk fat or major milk fatty acid groups, as concentration or yield, were based on milk trans-10 18:1 concentration. In conclusion, different responses between the two conditions to similar shift indicator levels support that the trans-10 shift would not be a major factor explaining the diet-induced low-fat milk syndrome in dairy ewes.

Effect of delipidant agents during in vitro culture on the development, lipid content, gene expression and cryotolerance of bovine embryos.

In vitro produced embryos are still sensitive to the freezing process which can be explained, in part, by the high-lipid accumulation that characterizes these embryos. Therefore, we aimed to evaluate the effect of delipidating agents, L-carnitine and the trans-10 cis-12 conjugated linoleic acid (CLA) isomer, on blastocyst development, lipid content, gene expression and cryotolerance when added to embryo culture media. Embryos were cultured in four different media: T1: control (n=616), synthetic oviduct fluid (SOF) media with 5% foetal bovine serum (FBS); T2: L-carnitine (n=648), SOF medium with 5% FBS and 0.6mg/ml of L-carnitine; T3: CLA (n=627), SOF medium with 5% FBS and 100μM trans-10 cis-12 CLA; and T4: L-carnitine+CLA: (n=597), SOF medium with 5% FBS plus 0.6mg/ml L-carnitine and 100μM trans-10 cis-12 CLA. Supplementation of culture medium with either or both delipidating agents reduced (p<.05) blastocyst rate on D7 (T1=49±3.5; T2=39±3.0; T3=42±3.9 and T4=39±3.9), but did not affected gene expression (p>.05). Although embryos cultured in the presence of L-carnitine contained fewer (p<.05) lipid droplets than the control embryos, they showed a lower re-expansion rate 24hr post-thaw than those (p<.05). In conclusion, although L-carnitine reduced the amount of lipids in cultured embryos, the use of L-carnitine and CLA during in vitro culture was not able to improve the embryo production and the response to cryopreservation.

Effect of trans-10, cis-12 conjugated linoleic acid (CLA) on expression of lipogenic genes is related to amount of trans-10, cis-12 CLA per unit of metabolic weight in lactating dairy ewes

This study evaluated the effect of trans-10, cis-12 CLA on the expression of genes involved in decreasing milk fat synthesis in two groups of lactating ewes with different metabolic weights. Twelve Lacaune ewes were alloted, according to metabolic weight (MW), into a Light Metabolic Weight (LMW) and Heavy Metabolic Weigth (HMW) groups, both of which were orally fed 30 g/d of rumen-unprotected CLA supplement containing 29.9% trans-10, cis-12. Milk samples were collected on days 0, 3 and 10 and mammary and adipose tissue biopsies were taken on days 3 and 10. RNA extraction, cDNA synthesis and RT-qPCR were performed and data were analyzed using the MIXED procedure of SAS as repeated measure in a completely randomized design. The trans-10, cis-12 CLA reduced milk fat synthesis between days 3 and 10 (P = 0.001), and the HMW group had a greater expression (P < 0.05) of the genes acetyl-CoA carboxylase alpha - promoter III (ACACA-α PIII), fatty acid synthase (FASN), stearoil CoA desaturase 1 (SCD1) and thyroid hormone responsive spot14 (THRSP) in the mammary gland and of acetyl-CoA carboxylase alpha - promoter I (ACACA-α PI) and leptin (P < 0.05) in adipose tissue. Our findings indicate that the response of animals to trans-10, cis-12 CLA appears to be metabolic weight dependent.

Kinetics of trans-10, cis-12-conjugated linoleic acid transfer to plasma and milk following an abomasal bolus in lactating dairy cows.

Trans-10, cis-12-conjugated linoleic acid (CLA) is a potent bioactive fatty acids (FA) that causes milk fat depression in lactating animals. FA are transferred to milk directly through chylomicrons and indirectly by recycling through other tissues. The objective of this study was to characterise the kinetics of trans-10, cis-12 CLA transfer to plasma and milk after a single bolus infusion. Five multiparous mid-lactation cows received a single abomasal bolus infusion of an enriched CLA mixture providing 15 g of trans-10, cis-12 CLA and 15 g of cis-9, trans-11 CLA over a 30-min period. Plasma concentration of trans-10, cis-12 and cis-9, trans-11 CLA peaked 2 h post-bolus, reaching 0·29 and 0·38 % of total plasma FA, respectively, and returned to pre-bolus values at 72 h post-infusion. Milk trans-10, cis-12 CLA yield and concentration peaked 14 h post-bolus (0·25 g/h) and was not detectable in milk after 86 h. Total apparent transfer of trans-10, cis-12 CLA to milk was 41 %, with 73 % transferred to milk through the direct pool (chylomicrons) and the remaining 27 % transferred through the indirect pool (tissue recycling). Compartmental modelling revealed the existence of a transient unavailable pool of trans-10, cis-12 CLA in extravascular tissues represented primarily by the mammary gland, which slowly exchanges with an available pool for secretion in milk fat and transfer to milk. In conclusion, trans-10, cis-12 CLA is predominantly transferred to milk through the direct pathway; however, how this CLA isomer is processed within the mammary gland requires further investigation.

Peroxisome proliferator-activated receptor gamma (PPARγ) agonist fails to overcome trans-10, cis-12 conjugated linoleic acid (CLA) inhibition of milk fat in dairy sheep

The trans-10, cis-12 conjugated linoleic acid (CLA) causes milk fat depression by downregulating expression of genes and transcription factors involved in lipogenesis and it has been proposed that peroxisome proliferator-activated receptor gamma (PPARγ) can be inhibited by trans-10, cis-12 CLA. The PPARγ is a nuclear receptor activated by natural or synthetic ligands and promotes expression of lipogenic genes and its effect on mammary lipogenesis and the interaction with trans-10, cis-12 CLA in lactating ewes was evaluated using thiazolidinedione (TZD), a chemical PPARγ agonist. A total of 24 lactating ewes were randomly assigned to one of the following treatments for 7 days: (1) Control (5 ml/day of saline solution); (2) TZD (4 mg/kg of BW/day in 5 ml of saline solution); (3) CLA (27 g/day with 29.9% of trans-10, cis-12); (4) TZD+CLA. Compared with Control, milk fat content was not changed by TZD, but was decreased 22.3% and 20.5% by CLA and TZD+CLA treatments. In the mammary gland, TZD increased PPARγ gene expression by 174.8% and 207.8% compared with Control and TZD+CLA treatments, respectively. Conjugated linoleic acid reduced sterol regulatory element-binding transcription protein 1 (SREBP1) gene expression 89.2% and 75.3% compared with Control and TZD+CLA, respectively, demonstrating that TZD fails to overcome CLA inhibition of SREBP1 signaling. In adipose tissue, the expression of SREBP1 and stearoyl CoA desaturase 1 (SCD1) genes were increased by the TZD+CLA treatment, compared with the other treatments. Conjugated linoleic acid decreased milk fat concentration and expression of lipogenic genes, while TZD had no effect on milk fat concentration, expression of lipogenic enzymes or regulators in the mammary gland and failed to overcome the inhibition of these by CLA. Therefore, CLA inhibition of milk fat synthesis was independent of the PPARγ pathway in lactating dairy ewes.

The trans-10,cis-12 conjugated linoleic acid increases triacylglycerol hydrolysis in yeast Saccharomyces cerevisiae.

The trans-10,cis-12 conjugated linoleic acid (CLA) is known for its antilipogenic effect but the mechanism is not fully clear. In this study, the potential of yeast (Saccharomyces cerevisiae) metabolism to offer evidence for the mechanism was investigated. The inhibitory effect of CLA on lipid accumulation was studied by analysing the transcript abundance of selected genes involved in triacylglycerol synthesis (LRO1, DGA1, ARE1 and ARE2) in the presence of the two bioactive CLA isomers: trans-10,cis-12 and the cis-9,trans-11 CLA. None of the enzymes was reduced in transcription but the expression of ARE2 was induced by trans-10,cis-12 CLA. However, the ARE2 overexpression did not contribute to lipid accumulation. The expression of the Δ9 desaturase gene, OLE1, was reduced by the cis-9,trans-11 but not by the trans-10,cis-12 isomer. In the TGL3/TGL4-knockout strain the triacylglycerol content also remained high in the CLA fed cells. Triacylglycerol hydrolysis rather than synthesis was the most probable reason for the reduced lipid content in yeast induced by CLA. This study revealed new aspects of the functionality of CLA in eukaryotic lipid metabolism. Yeast was proven to be an applicable model to study further the mechanism of trans-10,cis-12 CLA functionality on lipid metabolism.

Effect of conjugated linoleic acid and acetate on milk fat synthesis and adipose lipogenesis in lactating dairy cows

During biohydrogenation-induced milk fat depression (MFD), nutrients are spared from milk fat synthesis and are available for other metabolic uses. Acetate is the major carbon source spared and it may increase lipid synthesis in adipose tissue during MFD. The objective of this study was to compare the effect of trans-10,cis-12 conjugated linoleic acid (CLA) and the amount of acetate spared during CLA-induced MFD on adipose tissue lipogenesis. Nine multiparous, lactating, ruminally cannulated Holstein cows (244 ± 107 d in milk; 25 ± 8.4 kg of milk/d; mean ± standard deviation) were randomly assigned to treatments in a 3 × 3 Latin square design. Experimental periods were 4 d followed by a 10-d washout. Treatments were control (CON), ruminal infusion of acetate (AC; continuous infusion of 7 mol/d adjusted to pH 6.1 with sodium hydroxide), or abomasal infusion of CLA (10 g/d of both trans-10,cis-12 CLA and cis-9,trans-11 CLA). Dry matter intake, milk yield, and milk protein yield and percentage were not affected by treatments. Compared with CON, milk fat yield decreased 23% and fat percent decreased 28% in CLA, and milk fat yield increased 20% in AC. Concentration and yield of milk de novo synthesized fatty acids (<C16) were reduced and concentration of preformed fatty acids (>C16) was increased by CLA, compared with CON. Yield of de novo synthesized fatty acids and palmitic acid was increased by AC, compared with CON. Lipogenesis capacity of adipose tissue explants was decreased 72% by CLA, but was not affected by AC. Acetate oxidation by adipose explants was not affected by treatments. Treatments had no effect on expression of key lipogenic factors, lipogenic enzymes, and leptin; however, expression of fatty acid binding protein 4 was reduced in CLA compared with CON. Additionally, hormone-sensitive lipase and perilipin 1 were decreased by CLA and acetate. Plasma glucose and glucagon concentrations were not affected by treatments; however, CLA increased nonesterified fatty acids 17.7%, β-hydroxybutyrate 16.1%, and insulin 27.8% compared with CON, and AC increased plasma β-hydroxybutyrate 18%. In conclusion, during CLA-induced MFD in low-producing cow adipose tissue was sensitive to the anti-lipogenic effects of CLA, while spared acetate did not stimulate adipose lipogenesis. However, acetate may play an important role in stimulating lipogenesis and improving energy status in the mammary gland under normal conditions.

The effect of conjugated linoleic acid, acetate, and their interaction on adipose tissue lipid metabolism in nonlactating cows

During biohydrogenation-induced milk fat depression, adipose tissue lipogenesis may be increased through nutrients spared from milk fat synthesis. However, the direct effect of trans-10,cis-12 conjugated linoleic acid (CLA) and the indirect effect of spared nutrients on adipose tissue lipogenesis during milk fat depression is not clear. The objective of this study was to determine the direct effect of CLA, spared acetate, and their interaction on adipose tissue lipogenesis using nonlactating dairy cows as an experimental model, which allows separation of the effect of CLA and nutrient sparing. Eight ruminally cannulated, multiparous nonlactating and pregnant Holstein cows were randomly assigned to treatments in a 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments. Factors were CLA and acetate, and treatments were control (CON), rumen acetate infusions (Ac; continuous infusion of 7 mol/d adjusted to pH 6.1 with sodium hydroxide), abomasal infusion of trans-10,cis-12 CLA (CLA; 10 g/d of both trans-10,cis-12 CLA and cis-9,trans-11 CLA), and Ac + CLA (Ac + CLA). Dry matter intake was not affected by Ac, but tended to decrease by CLA. Plasma trans-10,cis-12 CLA concentration as a percentage of fatty acids was increased by 0.3 percentage points by CLA. No treatment effect was observed on plasma glucose and β-hydroxybutyrate, but an interaction was observed of CLA and Ac on plasma insulin and nonesterified fatty acids. Insulin was increased 24% by CLA, but not by Ac + CLA, and nonesterified fatty acids were increased 55% by Ac + CLA, but not by CLA alone. Lipogenesis and oxidation capacity of adipose tissue explants were not affected by treatments. Adipose expression of key lipogenic factors (peroxisome proliferator-activated receptor γ2 and sterol response element binding protein 1c) were reduced by CLA, by the interaction of Ac and CLA (sterol response element binding protein 1c), and tended to be reduced with Ac (S14 and peroxisome proliferator-activated receptor γ1). Expression of several adipose lipogenic enzymes (fatty acid synthase, acetyl-CoA carboxylase, and stearoyl-CoA desaturase 1) was reduced by CLA and Ac. An interaction was observed of Ac and CLA for fatty acid binding protein 4, which was decreased by Ac, but not Ac + CLA. In conclusion, in the nonlactating cow, adipose tissue is sensitive to the anti-lipogenic effects of trans-10,cis-12 CLA at the transcription level and acetate does not stimulate lipogenesis.

Conjugated linoleic acid supplementation during the transition period increased milk production in primiparous and multiparous dairy cows

Trans-10, cis-12 conjugated linoleic acid (CLA), a bioactive fatty acid, has the potential to alter energy metabolism in lactating dairy cows by marginally reducing milk fat synthesis in the mammary gland. The objective of this study was to determine the effects of pre- and postpartum CLA supplementation on lactation performance in a commercial dairy setting. Multiparous (mp) and primiparous (pp) Holstein cows managed in an automatic milking system were blocked by expected calving month, and randomly assigned to either a CLA supplementation (CLA-ME; mp n=99, pp n=38) or control (CTL; mp n=97, pp n=38) group. Cows were supplemented 100g/d of lipid encapsulated CLA methyl esters containing 10g each of cis-9, trans-11 and trans-10, cis-12 CLA, mixed 50:50 with soybean meal, and delivered via an automated mineral dispenser. Supplementation continued from −21 d pre-calving through 30 d in milk (DIM) for mp or 70 DIM for pp. Daily milk yield, fat and protein content, body weight, and rumination minutes were recorded and averaged by wk for the first 100 DIM. Milk yield was increased (P=0.04) during the supplementation period (44.7 vs. 46.6±0.64kg/d) for mp CLA-ME cows and was increased (P=0.03) during the post-supplementation period (31.3 vs. 34.2±0.91kg/d) for pp CLA-ME cows. Milk fat content was decreased (P=0.03) during the supplementation period for mp CLA-ME cows and post-supplementation period for pp CLA-ME cows (P=0.02); but only tended to be decreased (P=0.09) in the supplementation for pp CLA-ME cows. Fat yield was not altered (P>0.10) in either period for mp and pp CLA-ME. Supplementation with CLA did not change milk protein content or yield during the supplementation period for either mp or pp CLA-ME. During the post-supplementation period protein content was decreased (P=0.02); however, yield remained unchanged for mp CLA-ME cows while protein content was unchanged and yield increased for pp CLA-ME cows. Body weight and rumination minutes were unchanged by CLA supplementation. Intervals to first estrus, first service, and conception were similar for CTL and CLA-ME mp and pp cows. First service conception rate was similar for mp CTL and CLA-ME cows; but, tended to be increased (P=0.10) in pp CLA-ME compared to CTL. These results suggest that transition period CLA supplementation altered energy metabolism to increase milk production during the supplementation period for mp cows and post-supplementation period for pp cows without adversely affecting reproductive performance in mp cows and improving conception in pp cows.

Trans-10,cis-12 conjugated linoleic acid (CLA) interferes with lipid droplet accumulation during 3T3-L1 preadipocyte differentiation

In this study, we hypothesize that the biologically active isomers of conjugated linoleic acid (CLA), cis-9,trans-11 (c9,t11) and trans-10,cis-12 (t10,c12) CLA, have different effects on early and late stages 3T3-L1 preadipocyte differentiation. Both c9-t11 and t10-c12CLA stimulated early stage pre-adipocyte differentiation (day 2), while t10-c12CLA inhibited late differentiation (day 8) as determined by lipid droplet numbers and both perilipin-1 levels and phosphorylation state. At day 8, the adipokines adiponectin, chemerin and adipsin were all reduced in t10-c12CLA treated cells versus control cells. Immunofluorescence microscopy showed perilipin-1 was present solely on lipid droplets on day 8 in t10-c12 treated 3T3-L1 cells, whereas preilipin-1 was also located in the perinuclear region in control and c9-t11 treated cells. The t10-c12CLA isomer also decreased levels of hormone-sensitive lipase and inhibited lipolysis. These findings indicate that the decrease in lipid droplets caused by t10-c12CLA is the result of an inhibition of lipid droplet production during adipogenesis rather than a stimulation of lipolysis. Additionally, treatment with Gö6976 blocked the effect of t10-c12CLA on perilipin-1 phosphorylation, implicating PKCα in perilipin-1 phosphorylation, and thus a regulator of triglyceride catabolism. These data are supported by evidence that t10-c12CLA activated PKCα. These are the first data to show that CLA isomers can affect lipid droplet dynamics in adipocytes through PKCα.

Does trans-10, cis-12 conjugated linoleic acid affect the intermediary glucose and energy expenditure of dairy cows due to repartitioning of milk component synthesis?

The overall goal of this study was to evaluate if intermediary energy metabolism of cows fed with trans-10, cis-12 conjugated linoleic acid (CLA) was modified such that milk-energy compounds were produced with less intermediary energy expenditure as compared to control cows. Published data on supplemented CLA were assembled. The extent was calculated to which the trans-10, cis-12 CLA isomer has an impact on glucose and energy conversion in the mammary gland by modifying glucose equivalent supply and energy required for fatty acid (FA) and fat synthesis, and if this will eventually lead to an improved glucose and energy status of CLA-supplemented high-yielding dairy cows. A possible relationship between CLA supplementation level and milk energy yield response was also studied. Calculations were conducted separately for orally and abomasally administered CLA and based on energy required for supply of glucose equivalents, i.e. lactose, glycerol and NADPH2. Further, modifications of milk FA profile due to CLA supplementation were considered when energy expenditures for FA and fat synthesis were quantified. Differences in yields between control and CLA groups were transformed into glucose energy equivalents. Only abomasal infusion (r(2) = 0.31) but not oral CLA administration (r(2) = 0.11) supplementation to dairy cow diets resulted in less glucose equivalent energy. Modifications of milk FA profiles also saved energy but the relationship with CLA supplementation was weaker for abomasal infusion (r(2) = 0.06) than oral administration (r(2) = 0.38). On average, 10 g/d of abomasally infused trans-10, cis-12 CLA saved 1.1 to 2.3 MJ net energy expressed as glucose equivalents, whereas both positive and negative values were observed when the trans-10, cis-12 CLA was fed to the cows. This study revealed a weak to moderate dose-dependent relationship between the amount of trans-10, cis-12 CLA administered and the amount of energy in glucose equivalents and energy for the synthesis of milk fat conserved from milk ingredient synthesis. Because abomasal infusion of the trans-10, cis-12 CLA more consistently conserved energy in glucose equivalents compared with oral CLA intake, rumen protection of the fed CLA products appears incomplete. Milk fat synthesis showed an energy saving with a weak dose-dependent relationship when CLA was supplemented orally or by abomasal infusion.

Comparison of the nutritional regulation of milk fat secretion and composition in cows and goats

A study with 2 ruminant species (goats and cows) with inherent differences in lipid metabolism was performed to test the hypothesis that milk fat depression (MFD) due to marine lipid supplements or diets containing high amounts of starch and plant oil is caused by different mechanisms and that each ruminant species responds differently. Cows and goats were allocated to 1 of 3 groups (4 cows and 5 goats per group) and fed diets containing no additional oil (control) or supplemented with fish oil (FO) or sunflower oil and wheat starch (SOS) according to a 3×3 Latin square design with 26-d experimental periods. In cows, milk fat content was lowered by FO and SOS (−31%), whereas only FO decreased milk fat content in goats (−21%) compared with the control. Furthermore, FO and SOS decreased milk fat yield in cows, but not in goats. In both species, FO and SOS decreased the secretion of <C16 and C16 fatty acids (FA), and FO lowered >C16 FA output. However, SOS increased milk secretion of >C16 FA in goats. Compared with the control, SOS resulted in similar increases in milk trans-10,cis-12 conjugated linoleic acid (CLA) in both species, but caused a 2-fold larger increase in trans-10 18:1 concentration in cows than for goats. Relative to the control, responses to FO in both species were characterized by a marked decrease in milk concentration of 18:0 (−74%) and cis-9 18:1 (−62%), together with a ~5-fold increase in total trans 18:1, but the proportionate changes in trans-10 18:1 were lower for goats. Direct comparison of animal performance and milk FA responses to FO and SOS treatments demonstrated interspecies differences in mammary lipogenesis, suggesting a lower sensitivity to the inhibitory effects of trans-10,cis-12 CLA in goats and that ruminal biohydrogenation pathways are more stable and less prone to diet-induced shifts toward the formation of trans-10-containing intermediates in goats compared with cows. Even though a direct cause and effect could not be established, results suggest that regulation of milk fat synthesis during FO-induced MFD may be related to a shortage of 18:0 for endogenous mammary cis-9 18:1 synthesis, increase in the incorporation of trans FA in milk triacylglycerols, and limitations in the synthesis of FA de novo to maintain milk fat melting point. However, the possible contribution of biohydrogenation intermediates with putative antilipogenic effects in the mammary gland, including trans-9,cis-11 CLA, trans-10 18:1, or cis-11 18:1 to FO-induced MFD cannot be excluded.

Production of conjugated linoleic acid by heterologous expression of linoleic acid isomerase in oleaginous fungus Mortierella alpina.

To increase the commercial value of oleaginous fungus Mortierella alpina by incorporation of trans-10,cis-12 conjugated linoleic acid (CLA) into the polyunsaturated fatty acids (PUFAs) of M. alpina via Propionibacterium acnes isomerase (PAI) conversion. The PAI gene and the codon optimization version were heterologously expressed in M. alpina via Agrobacterium tumefaciens-mediated transformation (ATMT). Coding usage modification significantly improved the translation of PAI transcripts and trans-10,cis-12 CLA was produced up to 1.2 mg l(-1), which corresponds to approx. 0.05% of the total fatty acid (TFA). Since PAI prefers free linoleic acid as a substrate rather than any other forms, 5 μM long-chain acyl CoA synthetase inhibitor was added and the trans-10,cis-12 CLA content increased approx. 24-fold to 29 mg l(-1), reaching up to 1.2% (w/w) of the TFA in M. alpina. Heterologous expression of PAI in M. alpina by ATMT methods is a practicable way in biosynthesis of CLA and this system may be a feasible platform for industrial production of CLA.

Phosphatidylcholine with cis-9,trans-11 and trans-10,cis-12 Conjugated Linoleic Acid Isomers: Synthesis and Cytotoxic Studies

Novel phosphatidylcholines and lysophosphatidylcholines with cis-9,trans-11 and trans-10,cis-12 conjugated linoleic acid (CLA) were synthesized in high yields (75–99 %). The in vitro cytotoxic activities of these compounds against three human cancer cell lines (HL-60, MCF-7, and HT-29) were evaluated. The results revealed that there are differences in the activity between phosphatidylcholine with cis-9,trans-11 and trans-10,cis-12 CLA acyl groups. 1,2-Di(9Z,11E)-octadecadienoyl-sn-glycero-3-phosphocholine was the most potent cytotoxic agent among all tested CLA derivatives and its IC50 (concentration of a compound that inhibits the proliferation of 50 % of the cancer cell population) was 29.4 µM against HL-60. Moreover, phosphatidylcholines with CLA acyls exhibited much lower cytotoxicity against non-cancer cells (Balb/3T3) than free CLA isomers.

Effects of dietary pomegranate byproduct silage supplementation on performance, carcass characteristics and meat quality of growing lambs

In an experiment with 24 male growing Florina (Pelagonia) lambs, effects of dietary pomegranate byproduct silage (PS) on performance, carcass characteristics and meat chemical and fatty acid composition were determined. In the 9 week experiment, lambs were allocated to one of three dietary treatments (PS0, PS120, and PS240) of 8 lambs each. Lambs had an initial body weight (BW) of 18.8±2.28kg, and were fed one of three isonitrogenous (crude protein 171g/kg, dry matter – DM basis) and isoenergetic (net energy for gain 5.62MJ/kg, DM basis) total mixed rations (TMRs) ad libitum. The PS was added to the TMR at inclusion levels (as mixed basis) of 0, 120, and 240g/kg DM for treatments PS0, PS120, and PS240, respectively. No differences (P>0.05) occurred among PS treatments in final BW (34.3kg), BW gain (0.24kg/day), DM intake (0.94kg/day), and FCR (3.95kg DM intake/kg BW gain). Moreover, carcass characteristics were not affected (P>0.05) with increased PS feeding, except for the fat color, fat firmness, wetness and overall acceptability of carcasses, which improved (P<0.05). Fat content of meat was linearly increased (P<0.001) with increasing PS in lambs TMRs. PS supplementation caused linear (P<0.05) and quadratic (P<0.01) effect on 11-Hexadecenoic concentration and a linear decrease (P<0.05) in 6-9-Hexadecenoic and cis-11, 14-eicosadienoic acids concentrations, in intramuscular fat. A linear increase (P<0.05) was observed in linoleic acid and trans-10, cis-12 conjugated linoleic acid (CLA) (P<0.05) concentration, with increasing PS inclusion levels and a quadratic (P<0.05) and linear (P<0.01) effect on α-linolenic acid concentration. Meat total phenolic content increased linearly (P<0.05) and antioxidant activity was linearly (P<0.05) and quadratic affected (P<0.05), with maximum levels observed in intermediated group (PS120). Pomegranate byproduct silage supplementation, at levels up to 240g/kg DM of TMR, in isonitrogenous and iso (net energy) energetic diets for growing lambs did not affect their performance and carcass quantitative characteristics, but improved qualitative carcass characteristics. More significantly, dietary PS supplementation improved the nutritional and functional qualities, as well as the antioxidant potential of meat, as indicated by the increase in essential fatty acids, linoleic, α-linolenic acid and trans-10, cis-12 CLA, in intramuscular fat and the increase in total phenolic content and antioxidant activity.

High-level exogenous trans10, cis12 conjugated linoleic acid plays an anti-lipogenesis role in bovine mammary epithelial cells.

Primary bovine mammary epithelial cells (BMECs) were treated by 0, 37.5, 75, 112.5, 150 μmol/L trans10, cis12 conjugated linoleic acid (CLA) to evaluate the effects of different level trans10, cis12 CLA on lipogenesis in BMEC. Addition of 75-150 μmol/L trans10, cis12 CLA reduced significantly the triacylglycerol (TAG) content (P<0.05), but did not have inhibiting action on cell proliferation (P>0.05). Treatment with 150 μmol/L trans10, cis12 CLA for 48 h resulted in a 17.1% reduction (P<0.0001) of medium chain fatty acids (MCFA, C14<C<C16), a 26.5% reduction (P<0.0001) of unsaturated fatty acids (UFA) and a corresponding reduction of the mRNA abundance of acetyl coenzyme A (acetylCoA) carboxylase (ACC) (P=0.046), fatty acid synthase (FAS) (P=0.017) and stearoylCoA desaturase1 (SCD1) (P=0.002). Another finding was that trans10, cis12 CLA elevated expression of diacylglycerol acyltransferase2 (DGAT2) (P=0.020) and long chain acylCoA synthetases (ACSL) (P=0.032). In conclusion, higher trans10, cis12 CLA, not low trans10, cis12 CLA, inhibited milk fat synthesis and changed fatty acid composition by regulating the expression of FAS, ACC, SCD1, DGAT2 and ACSL.