The human organic cation transporter OCT1 mediates high affinity uptake of the anticancer drug daunorubicin

Les anthracyclines tels que la doxorubicin et la daunorubicin sont une famille de médicaments anticancéreux hydrophiles qui doivent être transportés dans les cellules afin d’exercer leur action par intercalation à l’ADN dans le noyau cellulaire. Ceci mène à la perturbation du métabolisme de l’ADN et entraine la mort cellulaire. Les anthracyclines sont utilisés pour le traitement d’une variété de cancers incluant la leucémie, les lymphomes, le cancer du sein, le cancer des poumons et le cancer des ovaires. Étant donné que le transport actif des anthracyclines dans les cellules a partiellement été démontré, le transporteur spécifique impliqué dans ce processus n’est pas encore connu. En utilisant un modèle de cancer des ovaires, la lignée cellulaire TOV2223G, nous avons démontré que des substrats spécifiques au transporteur de cations organiques 1 (OCT1), notamment la ergothionéine, la thiamine et la phenformin, ont partiellement inhibé l’absorption de la daunorubicin en différence de la carnitine qui est un substrat de haute affinité des transporteurs CT2 et OCTN2. Ces résultats suggèrent que les transporteurs organiques spécifiques au transport de la carnitine ne sont pas impliqués dans le transport des anthracyclines. Ainsi, nos résultats ont démontré que l’absorption de la daunorubicin est orchestrée par le transporteur OCT1 dans les cellules TOV2223G (Km ~ 5 μM) et des concentrations micromolaires de choline ont complètement abolies l’absorption de la drogue. De plus, un ARN sh dirigé contre OCT1 a réprimé son expression protéique, ce qui a été confirmé par la technique d’immuno-buvardage en utilisant un anti-OCT1 anticorps. Les cellules déficientes en OCT1 n’ont pas été capables d’absorber la daunorubicin et ont été plus résistantes à l’action de la drogue par rapport aux cellules contrôle. La transfection des cellules HEK293T avec un plasmide construit de façon à faire exprimer OCT1 comme protéine de fusion avec la protéine fluorescente EYFP a montré que celle-ci est localisée dans la membrane plasmique. Les cellules transfectées ont été capables d’absorber cinq fois plus de daunorubicin comparé aux cellules contrôles. Cette étude est, selon nous, la première à démontrer que OCT1 est un transporteur de haute affinité des anthracyclines. Ainsi, nous avons émis l’hypothèse que des défauts de OCT1 peuvent contribuer à l’efficacité de la réponse des cellules cancéreuses à la chimiothérapie avec les anthracyclines.

The human organic cation transporter OCT1 mediates high affinity uptake of the anticancer drug daunorubicin

Les anthracyclines tels que la doxorubicin et la daunorubicin sont une famille de médicaments anticancéreux hydrophiles qui doivent être transportés dans les cellules afin d’exercer leur action par intercalation à l’ADN dans le noyau cellulaire. Ceci mène à la perturbation du métabolisme de l’ADN et entraine la mort cellulaire. Les anthracyclines sont utilisés pour le traitement d’une variété de cancers incluant la leucémie, les lymphomes, le cancer du sein, le cancer des poumons et le cancer des ovaires. Étant donné que le transport actif des anthracyclines dans les cellules a partiellement été démontré, le transporteur spécifique impliqué dans ce processus n’est pas encore connu. En utilisant un modèle de cancer des ovaires, la lignée cellulaire TOV2223G, nous avons démontré que des substrats spécifiques au transporteur de cations organiques 1 (OCT1), notamment la ergothionéine, la thiamine et la phenformin, ont partiellement inhibé l’absorption de la daunorubicin en différence de la carnitine qui est un substrat de haute affinité des transporteurs CT2 et OCTN2. Ces résultats suggèrent que les transporteurs organiques spécifiques au transport de la carnitine ne sont pas impliqués dans le transport des anthracyclines. Ainsi, nos résultats ont démontré que l’absorption de la daunorubicin est orchestrée par le transporteur OCT1 dans les cellules TOV2223G (Km ~ 5 μM) et des concentrations micromolaires de choline ont complètement abolies l’absorption de la drogue. De plus, un ARN sh dirigé contre OCT1 a réprimé son expression protéique, ce qui a été confirmé par la technique d’immuno-buvardage en utilisant un anti-OCT1 anticorps. Les cellules déficientes en OCT1 n’ont pas été capables d’absorber la daunorubicin et ont été plus résistantes à l’action de la drogue par rapport aux cellules contrôle. La transfection des cellules HEK293T avec un plasmide construit de façon à faire exprimer OCT1 comme protéine de fusion avec la protéine fluorescente EYFP a montré que celle-ci est localisée dans la membrane plasmique. Les cellules transfectées ont été capables d’absorber cinq fois plus de daunorubicin comparé aux cellules contrôles. Cette étude est, selon nous, la première à démontrer que OCT1 est un transporteur de haute affinité des anthracyclines. Ainsi, nous avons émis l’hypothèse que des défauts de OCT1 peuvent contribuer à l’efficacité de la réponse des cellules cancéreuses à la chimiothérapie avec les anthracyclines.

Creatine Supplementation Prevents the Accumulation of Fat in the Livers of Rats Fed a High-Fat Diet

The aim of the present study was to examine the effects of creatine supplementation on liver fat accumulation induced by a high-fat diet in rats. Rats were fed 1 of 3 different diets for 3 wk: a control liquid diet (C), a high-fat liquid diet (HF), or a high-fat liquid diet supplemented with creatine (HFC). The C and HF diets contained, respectively, 35 and 71% of energy derived from fat. Creatine supplementation involved the addition of 1% (wt:v) of creatine monohydrate to the liquid diet. The HF diet increased total liver fat concentration, liver TG, and liver TBARS and decreased the hepatic S-adenosylmethionine (SAM) concentration. Creatine supplementation normalized all of these perturbations. Creatine supplementation significantly decreased the renal activity of L-arginine:glycine amidinotransferase and plasma guanidinoacetate and prevented the decrease in hepatic SAM concentration in rats fed the HF diet. However, there was no change in either the phosphatidylcholine:phosphatidylethanolamine (PE) ratio or PE N-methyltransferase activity. The HF diet decreased mRNA for PPAR as well as 2 of its targets, carnitine palmitoyltransferase and long-chain acylCoA dehydrogenase. Creatine supplementation normalized these mRNA levels. In conclusion, creatine supplementation prevented the fatty liver induced by feeding rats a HF diet, probably by normalization of the expression of key genes of beta-oxidation. J. Nutr. 141: 1799-1804, 2011.

Carnitine deficiency in chronic critical illness.

PURPOSE OF REVIEW: New insight in mitochondrial physiology has highlighted the importance of mitochondrial dysfunction in the metabolic and neuroendocrine changes observed in patients presenting with chronic critical illness. This review highlights specifically the importance of carnitine status in this particular patient population and its impact on beta-oxidation and mitochondrial function. RECENT FINDINGS: The main function of carnitine is long chain fatty acid esterification and transport through the mitochondrial membrane. Carnitine depletion should be suspected in critically ill patients with risk factors such as prolonged continuous renal replacement therapy or chronic parenteral nutrition, and evidence of beta-oxidation impairments such as inappropriate hypertriglyceridemia or hyperlactatemia. When fatty acid oxidation is impaired, acyl-CoAs accumulate and deplete the CoA intramitochondrial pool, hence causing a generalized mitochondrial dysfunction and multiorgan failure, with clinical consequences such as muscle weakness, rhabdomyolysis, cardiomyopathy, arrhythmia or sudden death. In such situations, carnitine plasma levels should be measured along with a complete assessment of plasma amino acid, plasma acylcarnitines and urinary organic acid analysis. Supplementation should be initiated if below normal levels (20 μmol/l) of carnitine are observed. In the absence of current guidelines, we recommend an initial supplementation of 0.5-1 g/day. SUMMARY: Metabolic modifications associated with chronic critical illness are just being explored. Carnitine deficiency in critically ill patients is one aspect of these profound and complex changes associated with prolonged stay in ICU. It is readily measurable in the plasma and can easily be substituted if needed, although guidelines are currently missing.

Une cause de cardiomyopathie dilatée chez l'enfant: le déficit primaire en carnitine [A cause of dilated cardiomyopathy in child: primary carnitine deficiency].

AIM: The aim of this case report was to show the importance to research metabolic etiology, especially a carnitine deficiency in dilated cardiomyopathy of children. CASE REPORT: A three years old Togolese child presented muscular hypotonia, dyspnea. Examination showed left galop murmur and systolic murmur 2/6. Chest X-ray showed cardiomegaly (CTI: 0.66), electrocardiogram, a sinusal rythm, left ventricle hypertrophy and T wave abnormalities. Echocardiogram showed a markedly dilated left ventricle with reduced systolic function (EF: 0.43; reference range 0.55-0.80) and moderate mitral regurgitation. The inflammatory signs where negatives. Magnetic resonance imaging don't show signs of ischemic or myocarditis. The levels of free and total plasmatic carnitine decreased: 3μmol/L (N: 18-48μmol/L) and 5μmol/l (N: 29-70μmol/L) respectively. Mutation analysis of the gene SLC22A5 confirms the diagnosis of primary systemic carnitine deficiency. Treatment with oral carnitine was started at 200mg/kg per day. Within three weeks of treatment, we observed the decrease of all symptoms and the left ventricular size and function normalized (EF: 0.62). He has now been on oral carnitine for live. CONCLUSION: Primary carnitine deficiency is a cause of dilated cardiomyopathy in child. It must systematically be suspected when a child presents a primitive cardiomyopathy. The treatment with oral carnitine for live is simple, with excellent prognosis.

Effect of supplementation with MgCO3 and L-tryptophan on the welfare and on the carcass and meat quality of two halothane pig genotypes (NN and nn)

Sixty-one animals with different Halothane genes (homozygous halothane positive, n=34; and homozygous halothane negative, n=27) were fed with three diets (controlgroup, with no supplement; magnesium (Mg) group with 1.28g MgCO3/kg and tryptophan (Trp) group with 5g L-Trp/kg) during the last 5 days before slaughter. Animals were submitted to minimal stress ante mortem conditions. Pig behaviour was recorded at the experimental farm, raceway to the CO2 stunning system and during the stunning period. Corneal reflexes were recorded after stunning as well. There were no differences in feed intake among diets (p>0.05) during the 5 days of treatment. Thehalothane positive (nn) group had lower intake than the halothane negative (NN) group(p<0.01). The behaviour of the pigs in the raceway did not differ (p>0.05) among treatments or halothane genotype. A significant (p<0.001) interaction diet*halothane was found in the time to appear the first retreat attempt during the exposure to the CO2 system. In the nn group, the time of performing the first retreat attempt was later in the Mg (p<0.05) than the Control group. Moreover, in the Mg group, the nn had a later (p<0.05) first retreat attempt than the NN. Thus, Mg supplementation could have a positive effect on welfare of nn pigs. The nn had a lower proportion of animals thatshowed corneal reflexes after stunning than NN, indicating a higher effectiveness of thestunning method in nn pigs. Neither Mg nor Trp affected carcass quality and meat quality parameters, although significant differences were found between genotypes

Effect of supplementation with MgCO3 and L-tryptophan on the welfare and on the carcass and meat quality of two halothane pig genotypes (NN and nn)

Sixty-one animals with different Halothane genes (homozygous halothane positive, n=34; and homozygous halothane negative, n=27) were fed with three diets (controlgroup, with no supplement; magnesium (Mg) group with 1.28g MgCO3/kg and tryptophan (Trp) group with 5g L-Trp/kg) during the last 5 days before slaughter. Animals were submitted to minimal stress ante mortem conditions. Pig behaviour was recorded at the experimental farm, raceway to the CO2 stunning system and during the stunning period. Corneal reflexes were recorded after stunning as well. There were no differences in feed intake among diets (p>0.05) during the 5 days of treatment. The halothane positive (nn) group had lower intake than the halothane negative (NN) group (p<0.01). The behaviour of the pigs in the raceway did not differ (p>0.05) among treatments or halothane genotype. A significant (p<0.001) interaction diet*halothane was found in the time to appear the first retreat attempt during the exposure to the CO2 system. In the nn group, the time of performing the first retreat attempt was later in the Mg (p<0.05) than the Control group. Moreover, in the Mg group, the nn had a later (p<0.05) first retreat attempt than the NN. Thus, Mg supplementation could have a positive effect on welfare of nn pigs. The nn had a lower proportion of animals that showed corneal reflexes after stunning than NN, indicating a higher effectiveness of the stunning method in nn pigs. Neither Mg nor Trp affected carcass quality and meat quality parameters, although significant differences were found between genotypes

Effect of carbohydrate supplementation on walking performance in peripheral arterial disease: A preliminary physiologic study

Objective: In this preliminary study we tested the effect of short-term carbohydrate supplementation on carbohydrate oxidation and walking performance in peripheral arterial disease. Methods: Eleven patients with peripheral arterial disease and intermittent claudication and 8 healthy control subjects completed several weeks of baseline exercise testing, then were given supplementation for 3 days with a carbohydrate solution and placebo. Maximal walking time was assessed with a graded treadmill test. Carbohydrate oxidation during a submaximal phase of this test was measured with indirect calorimetry. At the end of baseline testing a biopsy specimen was taken from the gastrocnemius muscle, and the active fraction of pyruvate dehydrogenase complex activity was determined. Results: Carbohydrate supplementation resulted in a significant increase in body weight and carbohydrate oxidation during exercise in patients with intermittent claudication and control subjects. Maximal walking time decreased by 3% in control subjects, whereas it increased by 6% in patients with intermittent claudication (group X treatment interaction, P < .05). There was a wide range of performance responses to carbohydrate supplementation among patients with claudication (-3%-37%). This effect was greater in poorer performers, and was negatively correlated (P < .05) with muscle pyruvate dehydrogenase complex activity. Conclusion: Preliminary data suggest that carbohydrate oxidation during exercise might contribute to exercise intolerance in more dysfunctional patients with intermittent claudication and that carbohydrate supplementation might be an effective therapeutic intervention in these patients.

Effects of high-dose creatine supplementation on kidney and liver responses in sedentary and exercised rats

This study evaluated the effects of high-dose of short-term creatine supplementation (5g.kg(-1). day(-1) to 1 week) and long-term creatine supplementation (1g.kg(-1). day(-1) to 4-8 weeks) on kidney and liver structure and function of sedentary and exercised Wistar rats ( Exercise sessions consisted of swimming at 80% of maximal work load supported during 5 days per week with daily sessions of 60 minutes throughout the duration of the supplementation). Seventy-two animals ( 245 +/- 5g) were divided into four groups (n = 18): control diet Sedentary ( SED), Creatine diet Sedentary (CRE), control diet Exercised (EXE), and Creatine diet Exercised (EXECRE). Histological and blood biochemical studies were performed after one, four, and eight weeks of creatine supplementation and exercise ( n = 6). No differences were found when comparing SED, EXE and EXECRE groups for kidney and liver structure and function at one, four and eight weeks. However, the CRE group showed higher levels of creatinine (1.1 +/- 0.2 vs. 0.4 +/- 0.1 mg.dl(-1); p < 0.05), and urea ( 37 +/- 3 vs. 19 +/- 1 mg. dl(-1); p < 0.05) when compared with all others groups at four and eight weeks. At eight weeks, the CRE group presented increased levels of ALT (41 +/- 7 vs. 23 +/- 7 U.L(-1); p < 0.05), AST (89 +/- 6 vs. 62 +/- 5 U. L(-1); p < 0.05), GGT (8.0 +/- 0.9 vs. 3.9 +/- 1.0 U. L(-1); p < 0.05), and AP (125 +/- 10 vs. 69 +/- 9 U. L(-1); p < 0.05) also when compared with all others groups. Moreover, the CRE group demonstrated some structural alterations indicating renal and hepatic damage at four and eight weeks, respectively. These results suggest that long-term creatine supplementation (up to 4-8 weeks) may adversely affect kidney and liver structure and function of sedentary but not of exercised rats.

Leucine supplementation improves adiponectin and total cholesterol concentrations despite the lack of changes in adiposity or glucose homeostasis in rats previously exposed to a high-fat diet

Background: Studies suggest that leucine supplementation (LS) has a therapeutic potential to prevent obesity and to promote glucose homeostasis. Furthermore, regular physical exercise is a widely accepted strategy for body weight maintenance and also for the prevention of obesity. The aim of this study was to determine the effect of chronic LS alone or combined with endurance training (ET) as potential approaches for reversing the insulin resistance and obesity induced by a high-fat diet (HFD) in rats. Methods: Forty-seven rats were randomly divided into two groups. Animals were fed a control diet-low fat (n = 10) or HFD (n = 37). After 15 weeks on HFD, all rats received the control diet-low fat and were randomly divided according to treatment: reference (REF), LS, ET, and LS+ET (n = 7-8 rats per group). After 6 weeks of treatment, the animals were sacrificed and body composition, fat cell volume, and serum concentrations of total cholesterol, HDL-cholesterol, triacylglycerol, glucose, adiponectin, leptin and tumor necrosis factor-alpha (TNF-alpha) were analyzed. Results: At the end of the sixth week of treatment, there was no significant difference in body weight between the REF, LS, ET and LS+ET groups. However, ET increased lean body mass in rats (P = 0.019). In addition, ET was more effective than LS in reducing adiposity (P = 0.019), serum insulin (P = 0.022) and TNF-alpha (P = 0.044). Conversely, LS increased serum adiponectin (P = 0.021) levels and reduced serum total cholesterol concentration (P = 0.042). Conclusions: The results showed that LS had no beneficial effects on insulin sensitivity or adiposity in previously obese rats. On the other hand, LS was effective in increasing adiponectin levels and in reducing total cholesterol concentration.

Role of beta-alanine supplementation on muscle carnosine and exercise performance

ARTIOLI, G. G., B. GUALANO, A. SMITH, J. STOUT, and A. H. LANCHA, JR. Role of beta-Alanine Supplementation on Muscle Carnosine and Exercise Performance. Med. Sci. Sports Exerc., Vol. 42, No. 6, pp. 1162-1173, 2010. In this narrative review, we present and discuss the current knowledge available on carnosine and beta-alanine metabolism as well as the effects of beta-alanine supplementation on exercise performance. Intramuscular acidosis has been attributed to be one of the main causes of fatigue during intense exercise. Carnosine has been shown to play a significant role in muscle pH regulation. Carnosine is synthesized in skeletal muscle from the amino acids L-histidine and beta-alanine. The rate-limiting factor of carnosine synthesis is beta-alanine availability. Supplementation with beta-alanine has been shown to increase muscle carnosine content and therefore total muscle buffer capacity, with the potential to elicit improvements in physical performance during high-intensity exercise. Studies on beta-alanine supplementation and exercise performance have demonstrated improvements in performance during multiple bouts of high-intensity exercise and in single bouts of exercise lasting more than 60 s. Similarly, beta-alanine supplementation has been shown to delay the onset of neuromuscular fatigue. Although beta-alanine does not improve maximal strength or (V) over dotO(2max), some aspects of endurance performance, such as anaerobic threshold and time to exhaustion, can be enhanced. Symptoms of paresthesia may be observed if a single dose higher than 800 mg is ingested. The symptoms, however, are transient and related to the increase in plasma concentration. They can be prevented by using controlled release capsules and smaller dosing strategies. No important side effect was related to the use of this amino acid so far. In conclusion, beta-alanine supplementation seems to be a safe nutritional strategy capable of improving high-intensity anaerobic performance.

Effects of creatine supplementation on renal function: a randomized, double-blind, placebo-controlled clinical trial

Creatine (CR) supplementation is commonly used by athletes. However, its effects on renal function remain controversial. The aim of this study was to evaluate the effects of creatine supplementation on renal function in healthy sedentary males (18-35 years old) submitted to exercise training. A randomized, double-blind, placebo-controlled trial was performed. Subjects (n = 18) were randomly allocated to receive treatment with either creatine (CR) (similar to 10 g day(-1) over 3 months) or placebo (PL) (dextrose). All subjects undertook moderate intensity aerobic training, in three 40-min sessions per week, during 3 months. Serum creatinine, serum and urinary sodium and potassium were determined at baseline and at the end of the study. Cystatin C was assessed prior to training (PRE), after 4 (POST 4) and 12 weeks (POST 12). Cystatin C levels (mg L-1) (PRE CR: 0.82 +/- 0.09; PL: 0.88 +/- 0.07 vs. POST 12 CR: 0.71 +/- 0.06; PL: 0.75 +/- 0.09, P = 0.0001) were decreased over time, suggesting an increase in glomerular filtration rate. Serum creatinine decreased with training in PL but was unchanged with training in CR. No significant differences were observed within or between groups in other parameters investigated. The decrease in cystatin C indicates that high-dose creatine supplementation over 3 months does not provoke any renal dysfunction in healthy males undergoing aerobic training. In addition, the results suggest that moderate aerobic training per se may improve renal function.

Effects of medium supplementation and pH control on lactic acid production from brewer`s spent grain

A cellulose pulp obtained by chemical pre-treatment of brewer`s spent grain was saccharified by a commercial cellulase preparation and the produced hydrolysate (50 g/l glucose) was fermented to lactic acid by Lactobacillus delbrueckii. The effects of pH control and nutrient supplementation of the hydrolysate on fermentation performance were investigated. Addition of 5g/l yeast extract enhanced the lactic acid volumetric productivity that attained 0.53 g/l h, value 18% higher than that obtained from non-supplemented hydrolysate. Addition of the MRS broth medium components (except the carbon source) was still better, providing a productivity of 0.79 g/l h. In all the cases, the lactic acid yield factor was of 0.7 g/g glucose consumed, but the fermentations stopped after 24 h due to the pH drop from 6.0 to 4.2, resulting in large amounts of residual glucose (38-41 g/l). Fermentation runs pH-controlled at 6.0 gave better results than those where the initial pH was not further controlled. The best result, 35.54 g/l lactic acid (0.99 g/g glucose consumed) was obtained during the pH-controlled fermentation of hydrolysate medium supplemented with MRS components. The volumetric productivity at the end of this fermentation was 0.59 g/l h, with a maximum of 0.82 g/l h during the first 12 h. (c) 2008 Elsevier B.V. All rights reserved.

Establishment of the optimum initial xylose concentration and nutritional supplementation of brewer`s spent grain hydrolysate for xylitol production by Candida guilliermondii

The effects of initial xylose concentration and nutritional supplementation of brewer`s spent grain hydrolysate on xylitol production by Candida guilliermondii were evaluated using experimental design methodology. The hydrolysate containing 55, 75 or 95 g/l xylose, supplemented or not with nutrients (calcium chloride, ammonium sulfate and rice bran extract), was used as fermentation medium. The increase in xylitol yield and productivity was related to the increase of initial xylose concentration, but up to a certain limit. above of which the yeast performance was not improved. The hydrolysate supplementation with nutrients did not interfere with xylose-to-xylitol conversion. By using the statistic tool the best conditions for maximum xylitol production were found. which consisted in using the non-supplemented hydrolysate containing 70 g/l initial xylose concentration. Under these conditions, a xylitol yield of 0.78 g/g and productivity of 0.58 g/(l h) were achieved. (C) 2008 Elsevier Ltd. All rights reserved.

Effects of dietary supplementation of rumen-protected conjugated linoleic acid to grazing cows in early lactation

Conjugated linoleic acids (CLA) are potent anticarcinogens in animal and in vitro models as well as inhibitors of fatty acid synthesis in mammary gland, liver, and adipose tissue. Our objective was to evaluate long-term CLA supplementation of lactating dairy cows in tropical pasture on milk production and composition and residual effects posttreatment. Thirty crossbred cows grazing stargrass (Cynodon nlemfuensis Vanderyst var. nlemfuensis) were blocked by parity and received 150 g/d of a dietary fat supplement of either Ca-salts of palm oil fatty acids (control) or a mixture of Ca-salts of CLA (CLA treatment). Supplements of fatty acids were mixed with 4 kg/d of concentrate. Grazing plus supplements were estimated to provide 115% of the estimated metabolizable protein requirements from 28 to 84 d in milk (treatment period). The CLA supplement provided 15 g/d of cis-9, trans-11 and 22 g of cis-10, trans-12. Residual effects were evaluated from 85 to 112 d in milk (residual period) when cows were fed an 18% crude protein concentrate without added fat. The CLA treatment increased milk production but reduced milk fat concentration from 2.90 to 2.14% and fat production from 437 to 348 g/d. Milk protein concentration increased by 11.5% (2.79 to 3.11%) and production by 19% (422 to 504 g/d) in the cows fed CLA. The CLA treatment decreased milk energy concentration and increased milk volume, resulting in unchanged energy output. Milk production and protein concentration and production were also greater during the residual period for the CLA-treated cows. The CLA treatment reduced production of fatty acids (FA) of all chain lengths, but the larger effect was on short-chain FA, causing a shift toward a greater content of longer chain FA. The CLA treatment increased total milk CLA content by 30% and content of the trans-10, cis-12 CLA isomer by 88%. The CLA treatment tended to decrease the number of days open, suggesting a possible effect on reproduction. Under tropical grazing conditions, in a nutritionally challenging environment, CLA-treated cows decreased milk fat content and secreted the same amount of milk energy by increasing milk volume and milk protein production.