Effects of l-carnitine in the distillers dried grains with solubles diet of sows on reproductive performance and antioxidant status of sows and their offspring

Distillers dried grains with solubles (DDGS) are highly susceptible to lipid oxidation because DDGS contain about 10% crude fat, which is largely composed of polyunsaturated fatty acids. l-carnitine serves an important function in fatty acids β-oxidation, and also has antioxidant properties. The objective of this study was to examine the effects of l-carnitine in the DDGS diet of gestating and lactating sows on reproductive performance, milk composition and antioxidant status of sows and their offspring. One hundred and twenty sows (Landrace×Large white, mean parity 4.2, initial BW 230 kg) were randomly allotted to 1 of 4 dietary treatments (n=30 sows/treatment). Treatments were arranged as a 2×2 factorial with two levels of dietary DDGS (0 v. 250 g/kg in gestating diets and 400 g/kg in lactating diets) and two levels of dietary l-carnitine (0 v. 100 mg/kg in gestating diets and 0 v. 200 mg/kg in lactating diets). Distillers dried grains with solubles had no significant effect on litter size but significantly reduced the birth weights and weaning weights of piglets (P<0.05). Distillers dried grains with solubles reduced the antioxidant enzyme activities (P<0.05) and increased the malondialdehyde level in the plasma of sows on day 60 of gestation (P=0.004) and day 14 of lactation (P=0.008). The compositions of colostrum and milk were not affected by inclusion of DDGS and dietary l-carnitine (P>0.05). Supplementing the diets with l-carnitine had no significant effect of total litter size (P>0.05) but increased the number of piglets born alive and piglets weaned, birth weight and weaning weight of piglets and litter weight at birth and weaning (P<0.05). l-carnitine supplementation also increased the concentration of l-carnitine in milk and l-carnitine status of piglets (P<0.05). The antioxidant enzyme activities of new born and weaning piglets were increased (P<0.05) by maternal dietary l-carnitine but this did not extend to finishing pigs. In conclusion, including DDGS in the sows diet could induce oxidative stress, which may be associated with the reduced individual birth and weaning weight of piglets. Dietary l-carnitine supplementation improved the antioxidant and l-carnitine status of sows, which may be associated with the improved reproduction and piglet performance and the antioxidant status of piglets at birth and weaning. There were no interactions between DDGS and l-carnitine.     

Effect of dehydroepiandrosterone sulfate on carnitine acetyl transferase activity and l-carnitine levels in oophorectomized rats

Alteration in energy metabolism of postmenopausal women might be related to the reduction of dehydroepiandrosterone sulfate (DHEAS). DHEA and DHEAS decline with age, leveling at their nadir near menopause. DHEA and DHEAS modulate fatty acid metabolism by regulating carnitine acyltransferases and CoA. The purpose of this study was to determine whether dietary supplementation with DHEAS would also increase tissue l-carnitine levels, carnitine acetyltransferase (CAT) activity and mitochondrial respiration in oophorectomized rats. Plasma l-carnitine levels rose following oophorectomy in all groups (P<0.0001). Supplementation with DHEAS was not associated with further elevation of plasma l-carnitine levels, but with increased hepatic total and free l-carnitine (P=0.021 and P<0.0001, respectively) and cardiac total l-carnitine concentrations (P=0.045). In addition, DHEAS supplementation increased both hepatic and cardiac CAT activities (P<0.0001 and P=0.05 respectively). CAT activity positively correlated with the total and free carnitine levels in both liver and heart (r=0.764, r=0.785 and r=0.700, r=0.519, respectively). Liver mitochondrial respiratory control ratio, ADP:O ratio and oxygen uptake were similar in both control and supplemented groups. These results demonstrate that in oophorectomized rats, dietary DHEAS supplementation increases the liver and heart l-carnitine levels and CAT activities. In conclusion, DHEAS may modulate l-carnitine level and CAT activity in estrogen deficient rats. The potential role of DHEAS in the regulation of fatty acid oxidation in postmenopausal women is worthy of investigation.     

Distinct transport activity of tetraethylammonium from l-carnitine in rat renal brush-border membranes

We investigated the contribution of the Na⁺/l-carnitine cotransporter in the transport of tetraethylammonium (TEA) by rat renal brush-border membrane vesicles. The transient uphill transport of l-carnitine was observed in the presence of a Na⁺ gradient. The uptake of l-carnitine was of high affinity (K_m=21 μM) and pH dependent. Various compounds such as TEA, cephaloridine, and p-chloromercuribenzene sulfonate (PCMBS) had potent inhibitory effects for l-carnitine uptake. Therefore, we confirmed the Na⁺/l-carnitine cotransport activity in rat renal brush-border membranes. Levofloxacin and PCMBS showed different inhibitory effects for TEA and l-carnitine uptake. The presence of an outward H⁺ gradient induced a marked stimulation of TEA uptake, whereas it induced no stimulation of l-carnitine uptake. Furthermore, unlabeled TEA preloaded in the vesicles markedly enhanced [¹⁴C]TEA uptake, but unlabeled l-carnitine did not stimulate [¹⁴C]TEA uptake. These results suggest that transport of TEA across brush-border membranes is independent of the Na⁺/l-carnitine cotransport activity, and organic cation secretion across brush-border membranes is predominantly mediated by the H⁺/organic cation antiporter.     

Production of l-carnitine from d-carnitine by dried cells of Agrobacterium sp. 525a

d-Carnitine was converted into l-carnitine with dried cells of Agrobacterium sp. 525a grown on d-carnitine medium. During the conversion reaction, NAD⁺ concentration decreased. o-Phenanthroline inhibited the NAD⁺ degrading activity in dried cells, and the formation of l-carnitine increased 2.8 times. About 190 mM of l-carnitine was converted from 0.5 M of d-carnitine under the best conditions.     

Mechanism of the inhibitory effect of zwitterionic drugs (levofloxacin and grepafloxacin) on carnitine transporter (OCTN2) in Caco-2 cells

l-Carnitine plays an important role in lipid metabolism by facilitating the transport of long-chain fatty acids across the mitochondrial inner membrane followed by fatty acid beta-oxidation. It is known that l-carnitine exists as a zwitterion and that member of the OCTN family play an important role in its transport. The aims of this study were to characterize l-carnitine transport in the intestine by using Caco-2 cells and to elucidate the effects of levofloxacin (LVFX) and grepafloxacin (GPFX), which are zwitterionic drugs, on l-carnitine uptake. Kinetic analysis showed that the half-saturation Na⁺ concentration, Hill coefficient and K_m value of l-carnitine uptake in Caco-2 cells were 10.3 ± 4.5 mM, 1.09 and 8.0 ± 1.0 μM, respectively, suggesting that OCTN2 mainly transports l-carnitine. LVFX and GPFX have two pK_a values and the existence ratio of their zwitterionic forms is higher under a neutral condition than under an acidic condition. Experiments on the inhibitory effect of LVFX and GPFX on l-carnitine uptake showed that LVFX and GPFX inhibited l-carnitine uptake more strongly at pH 7.4 than at pH 5.5. It was concluded that the zwitterionic form of drugs plays an important role in inhibition of OCTN2 function.     

A high-fat diet increases l-carnitine synthesis through a differential maturation of the Bbox1 mRNAs

l-carnitine is a key molecule in both mitochondrial and peroxisomal lipid metabolisms. l-carnitine is biosynthesized from gamma-butyrobetaine by a reaction catalyzed by the gamma-butyrobetaine hydroxylase (Bbox1). The aim of this work was to identify molecular mechanisms involved in the regulation of l-carnitine biosynthesis and availability. Using 3′ RACE, we identified four alternatively polyadenylated Bbox1 mRNAs in rat liver. We utilized a combination of in vitro experiments using hybrid constructs containing the Bbox1 3′ UTR and in vivo experiments on rat liver mRNAs to reveal specificities in the different Bbox1 mRNA isoforms, especially in terms of polyadenylation efficiency, mRNA stability and translation efficiency. This complex maturation process of the Bbox1 mRNAs in the liver was studied on rats fed a high-fat diet. High-fat diet selectively increased the level of three Bbox1 mRNA isoforms in rat liver and the alternative use of polyadenylation sites contributed to the global increase in Bbox1 enzymatic activity and l-carnitine levels. Our results show that the maturation of Bbox1 mRNAs is nutritionally regulated in the liver through a selective polyadenylation process to adjust l-carnitine biosynthesis to the energy supply.     

The effect of dietary l-carnitine supplementation on cold tolerance and growth of the ornamental cichlid fish Pelvicachromis pulcher—preliminary results

The effect of dietary l-carnitine supplementation on cold tolerance, growth and survival of the ornamental cichlid Pelvicachromis pulcher was tested under laboratory conditions. Fish were reared for a period of 82 days on a diet containing different levels of added l-carnitine: 0, 500 (3.1 mmol), 1000 (6.2 mmol) and 2000 (12.4 mmol) mg/kg. At the end of the growth period the fish were exposed to a cold shock. Fish which received dietary l-carnitine supplementation (at all tested levels) exhibited significantly better survival rates following exposure to a cold shock, and readapted quicker to normal temperatures compared to the control group which had no l-carnitine added to their diet. The addition of l-carnitine to the diet at a level of 1000 mg/kg seems to yield the best protection against exposure to cold shock.Growth differences among the treated fish were not significant, although the fish which received 500 mg/kg supplementation exhibited slightly better growth.     

Protective action of l-carnitine on cardiac mitochondrial function and structure against fatty acid stress

Cardiovascular risks are frequently accompanied by high serum fatty acid levels. Although recent studies have shown that fatty acids affect mitochondrial function and induce cell apoptosis, l-carnitine is essential for the uptake of fatty acids by mitochondria, and may attenuate the mitochondrial dysfunction and apoptosis of cardiocytes. This study aimed to elucidate the activity of l-carnitine in the prevention on fatty acid-induced mitochondrial membrane permeability transition and cytochrome c release using isolated cardiac mitochondria from rats. Palmitoyl-CoA-induced mitochondrial respiration that was observed with l-carnitine was inhibited with oligomycin. The palmitoyl-CoA-induced mitochondrial membrane depolarization and swelling were greatly inhibited by the presence of l-carnitine. In ultrastructural observations, terminally swollen and ruptured mitochondria with little or no distinguishable cristae structures were induced by treatment with palmitoyl-CoA. However, the severe morphological damage in cardiac mitochondria was dramatically inhibited by pretreatment with l-carnitine. Treatment with l-carnitine also attenuated 4-hydroxy-l-phenylglycine- and rotenone-induced mitochondrial swelling even when the l-carnitine could not protect against the decrease in oxygen consumption associated with these inhibitors. Furthermore, l-carnitine completely inhibited palmitoyl-CoA-induced cytochrome c release. We concluded that l-carnitine is essential for cardiac mitochondria to attenuate the membrane permeability transition, and to maintain the ultrastructure and membrane stabilization, in the presence of high fatty acid β-oxidation. Consequently, the cells may be protected against apoptosis by l-carnitine through inhibition of the fatty acid-induced cytochrome c release.     

In vitro effects of l-carnitine and glutamine on motility,acrosomal abnormality,and plasma membrane integrity of rabbit sperm during liquid-storage

This study was designed to evaluate the in vitro effects of l-carnitine and glutamine (Gln) on the sperm quality parameters of liquid-stored rabbit semen maintained up to 24 h at 5 °C. Pooled and extended ejaculates were divided into two equal portions. l-Carnitine doses of 0.5, 1 and 2 mM were added to the first portion, and glutamine was added at the same doses to the second portion. All samples were cooled to 5 °C and examined at 0, 6, 12 and 24 h of liquid storage. Supplementation of the semen extender with three different doses of l-carnitine provided significant increases in the percentage of motile sperm at 12 h (P < 0.01), and 24 h (P < 0.001) and enabled significant protection of the sperm plasma membrane (P < 0.01) at 12 and 24 h of cool-storage, in comparison to the control samples. Only the 2 mM dose of l-carnitine significantly (P < 0.01) decreased the rate of acrosomal damage when compared to the control group. Furthermore, all doses of Gln caused a significant (P < 0.01) decrease in acrosomal damage at 6 h, and provided significant improvement (P < 0.01) in sperm motility, acrosomal and plasma membrane integrities at 12 and 24 h of liquid storage, when compared to the controls. In conclusion, the supplementation of liquid-stored rabbit semen with l-carnitine and Gln provided a protection for sperm against cool storage-induced functional and structural damages.     

Factors affecting the biotransformation of trimethylammonium compounds into l-carnitine by Escherichia coli

The biotransformation of d-carnitine and crotonobetaine into l-carnitine with wild and transformed E. coli strains under batch and continuous operation was optimised. In batch, the best conditions for the transformed strain were 30% oxygen saturation, a temperature of 41 °C and a minimal medium, whereas anaerobic cultures in either complex or minimal media at 37 °C and pH 7.5 were optimal for the wild strain. Studies on the expression of the enzymes involved in trimethylammonium metabolism showed that l-carnitine dehydratase activity was always higher than that of d-carnitine racemase. Experiments with the transformed strain in continuous cell-recycle reactors showed that, despite the higher productivity that could be achieved (0.65–1.2 g/L h), plasmid-bearing cells were segregated even when a selective medium was used. This fact was also confirmed by studying the evolution of the d-carnitine racemase level. Immobilization of the transformed strain in κ-carrageenan gels allowed continuous operation for l-carnitine production with no plasmid loss. In continuous processes with cell-retention systems, the wild strain showed higher productivity and stability than the transformed strain. Moreover, crotonobetaine was a better substrate for both strains used. Recycling with hollow-fiber cartridges provided the highest biomass level even though the l-carnitine dehydratase/biomass ratio was lower. However, membrane composition and cut-off had less influence on reactor performance as similar levels of productivity were attained. In spite of this, continuous processes attained a l-carnitine production as high as 11.5 g/L h as a result of the high enzyme induction and biomass levels.     

Ameliorative effects of l-carnitine on rats raised on a diet supplemented with lead acetate

Lead intoxication has been a major health hazard in humans. It affects people at all ages. Its toxicity is associated with various organs of the body and affects different metabolic pathways. Based on histological data, l-carnitine reduced the severity of tissue damage produced as a result of exposure of rats to lead acetate. The main objective of this study was to evaluate the underlying mechanism of protection offered by l-carnitine against lead acetate intoxication using male Sprague–Dawley rats.Forty male Sprague–Dawley rats were randomly divided into four groups with ten rats in each. The first group (G1) served as the control group and animals received standard diet only. The second group (G2) received lead acetate in their diet. The third group (G3) was the l-carnitine treated group and received the normal standard diet supplemented with l-carnitine. While the fourth group (G4) had a diet supplemented with both lead acetate and l-carnitine. At the end of each experiment, blood (serum and whole blood) were collected from each animal and analyzed for the following parameters: serum testosterone levels, serum nitric oxide and serum malondialdehyde. This is in addition to looking at the enzymatic activities of two important enzymes (superoxide dismutase and catalase) and on (glutathione reductase) which are indicative of the antioxidant activities in the whole blood. The results indicated that l-carnitine will counteract the undesirable effects of lead intoxication. It exerted its antioxidant potential by reducing the production of ROS and scavenging free radicals by maintaining and protecting the level of the of antioxidant enzymes SOD, CAT and glutathione peroxidase.Conclusion:l-Carnitine may play an important role in reversing the undesirable effects of lead intoxication. Future studies should be conducted to see whether such an effect is applicable in humans exposed to lead poising.     

Carnitine and acetyl-carnitine,natural substances endowed with interesting pharmacological properties.

The inotropic and anti-fatigue effects of d, l-carnitine, l-carnitine and d, l-acetyl-carnitine in laboratory animals were studied. In the isolated rabbit heart these substances at the concentration level of 10^?5 g/l increase significantly the contractile force depressed by anoxia, without changes in ECG, heart rate and coronary flow. The following rank order of activity was observed: d, l-acetyl-carnitine > l-carnitine > d, l-carnitine. Sodium acetate + d, l-carnitine mixture exerts the same inotropic effect as d, l-carnitine alone, showing the important role of the acetyl-bond. In rats submitted to a fatigue test (rotarod) only d, l-acetyl-carnitine induces a significant anti-fatigue effect. Since endogenous carnitine depletion is linked with many pathological states, particularly of the heart, exogenous carnitine and especially its acetyl-derivative might represent an important drug treatment with high tolerance advantages.     

l-Carnitine attenuates angiotensin II-induced proliferation of cardiac fibroblasts: role of NADPH oxidase inhibition and decreased sphingosine-1-phosphate generation

The heart is unable to synthesize l-carnitine and is strictly dependent on the l-carnitine provided by the blood stream; however, additional studies are needed to better understand the mechanism of l-carnitine supplementation to the heart. The aim of this study was to evaluate the effects of l-carnitine on angiotensin II (Ang II)-induced cardiac fibroblast proliferation and to explore its intracellular mechanism(s). Cultured rat cardiac fibroblasts were pretreated with l-carnitine (1-30 mM) then stimulated with Ang II (100 nM). Ang II increased fibroblast proliferation and endothelin-1 expression, which were partially inhibited by l-carnitine. l-Carnitine also attenuated Ang II-induced NADPH oxidase activity, reactive oxygen species formation, extracellular signal-regulated kinase phosphorylation, activator protein-1-mediated reporter activity and sphingosine-1-phosphate generation. In addition, l-carnitine increased prostacyclin (PGI₂) generation in cardiac fibroblasts. siRNA transfection of PGI₂ synthase significantly reduced l-carnitine-induced PGI₂ and its anti-proliferation effects on cardiac fibroblasts. Furthermore, blockading potential PGI₂ receptors, including immunoprecipitation (IP) receptors and peroxisome proliferator-activated receptors alpha (PPARα) and delta , revealed that siRNA-mediated blockage of PPARα considerably reduced the anti-proliferation effect of l-carnitine. In summary, these results suggest that l-carnitine attenuates Ang II-induced effects (including NADPH oxidase activation, sphingosine-1-phosphate generation and cell proliferation) in part through PGI₂ and PPARα-signaling pathways.     

Effect of orally administered l-carnitine on selected biochemical indicators of lactating Tuj-ewes

The effect of orally administered l-carnitine on biochemical parameters was examined in lactating Tuj-ewes. Ewes were orally given 500 mg of l-carnitine daily for 3 weeks. To evaluate the changes on selected blood indicators (total protein, albumin, glucose, triglyceride, cholesterol, urea, aspartate amino transferase, alanine amino transferase, lipase, triiodothyronine and thyroxine), blood samples were collected at the beginning of the study, and at the end of 1st, 2nd and 3rd week of study. Oral administration of supplemental carnitine significantly decreased serum triglyceride (P < 0.05), glucose (P < 0.05), cholesterol (P < 0.05) and triiodothyronine (P < 0.05) concentrations. In addition, serum thyroxine (P < 0.001) and albumin (P < 0.01) concentrations were significantly elevated as a result of oral carnitine treatment. These results suggest that supplemental l-carnitine improves selected biochemical indicators in Tuj-ewes.     

A diet supplemented with l-carnitine improves the sperm quality of Piétrain but not of Duroc and Large White boars when photoperiod and temperature increase

It has been reported that a diet supplemented with l-carnitine can improve sperm quality in some mammalian species. Against this background, the current study seeks to determine the effects of feeding l-carnitine (625 mg·day⁻¹) on boar semen characteristics (ejaculate volume, sperm concentration, sperm viability, acrosome and mitochondrial sheath integrity, sperm motility, sperm morphology, and osmotic resistance of spermatozoa) in three different porcine breeds (Sus domesticus) (Piétrain, Duroc, and Large White) exposed to natural environmental changes in temperature and photoperiod over a 20-wk period (February to July 2007). One hundred twenty boars (40 per breed) were randomly separated into two groups (60 boars each): the first (20 boars per breed) was fed a control diet and the second (also 20 males per breed) the same diet supplemented with l-carnitine (625 mg·day⁻¹). Whereas the l-carnitine supplement did not affect ejaculate volume, concentration, motility, viability, or the osmotic resistance of spermatozoa, it did improve sperm morphology in Piétrain boars by reducing the percentage of immature spermatozoa when the temperature and the photoperiod increased. Conversely, no effect on sperm morphology from supplementing feed with l-carnitine was observed in both Duroc and Large White breeds. We can therefore conclude that the addition of l-carnitine to the diet of males may maintain the level of normal sperm morphology in Piétrain boars when a drop in sperm quality occurs (due to increases in photoperiod and temperature), without affecting the other sperm quality parameters.     

Metabolism of labeled carnitine in the rat

In two series of rats, the concentration of carnitine in plasma was 39.9 and 37.8 μmol/ liter, in skeletal muscle tissue 2.97 and 3.26 μmol/g dry wt and the urinary excretion 3.2 and 2.4 μmol/24 h. The renal clearance of carnitine was calculated to 88 and 76 ml/24 h. L-[Me-¹⁴C]Carnitine and DL-[Me-¹⁴C]carnitine have been administered to rats. Only labeled l-carnitine has been found on chromatographic analysis of plasma, urine, and muscle tissue. The specific radioactivity of carnitine in plasma, urine, and muscle tissue has been followed for up to 16 days. A two-compartment metabolic model has been used to interpret the result of the experiment with labeled l-carnitine and the rate constants and compartment sizes have been calculated. The total body content of carnitine was 57 μmol (about 35 μmol/100 g body wt) and the daily turnover was about 7% of the body pool. The daily synthesis of carnitine in the rat is estimated to about 2 μmol/100 g body wt.     

l-carnitine supplementation as a potential antioxidant therapy for inherited neurometabolic disorders

In recent years increasing evidence has emerged suggesting that oxidative stress is involved in the pathophysiology of a number of inherited metabolic disorders. However the clinical use of classical antioxidants in these diseases has been poorly evaluated and so far no benefit has been demonstrated. l-Carnitine is an endogenous substance that acts as a carrier for fatty acids across the inner mitochondrial membrane necessary for subsequent beta-oxidation and ATP production. Besides its important role in the metabolism of lipids, l-carnitine is also a potent antioxidant (free radical scavenger) and thus may protect tissues from oxidative damage. This review addresses recent findings obtained from patients with some inherited neurometabolic diseases showing that l-carnitine may be involved in the reduction of oxidative damage observed in these disorders. For some of these diseases, reduced concentrations of l-carnitine may occur due to the combination of this compound to the accumulating toxic metabolites, especially organic acids, or as a result of protein restricted diets. Thus, l-carnitine supplementation may be useful not only to prevent tissue deficiency of this element, but also to avoid oxidative damage secondary to increased production of reactive species in these diseases. Considering the ability of l-carnitine to easily cross the blood–brain barrier, l-carnitine supplementation may also be beneficial in preventing neurological damage derived from oxidative injury. However further studies are required to better explore this potential.     

Hypoxia-Induced Lipid Peroxidation in Rat Brain and Protective Effect of Carnitine and Phosphocreatine

The exposure to hypobaric hypoxia increased lipid peroxidation (as indicated by thiobarbituric acid-reactive substances [TBARS] in rat brain. Plasma lactate/pyruvate ratio was used as a marker of hypoxia. We compared the protective effect of -tocopherol with the effect of l-carnitine or phosphocreatine. Rats pretreated with -tocopherol, l-carnitine, or phosphocreatine had lower TBARS levels after the exposure to hypobaric hypoxia. However, lactate/pyruvate ratio was improved only in rats pretreated with l-carnitine or phosphocreatine. We conclude from our data that, contrary to -tocopherol, protective effects of l-carnitine and phosphocreatine administrations are due to their regulation of metabolic reactions during hypobaric hypoxia rather than to their scavenger activity.     

CaiT of Escherichia coli,a new transporter catalyzing L-carnitine/gamma -butyrobetaine exchange

l-Carnitine is essential for beta-oxidation of fatty acids in mitochondria. Bacterial metabolic pathways are used for the production of this medically important compound. Here, we report the first detailed functional characterization of the caiT gene product, a putative transport protein whose function is required for l-carnitine conversion in Escherichia coli. The caiT gene was overexpressed in E. coli, and the gene product was purified by affinity chromatography and reconstituted into proteoliposomes. Functional analyses with intact cells and proteoliposomes demonstrated that CaiT is able to catalyze the exchange of l-carnitine for gamma-butyrobetaine, the excreted end product of l-carnitine conversion in E. coli, and related betaines. Electrochemical ion gradients did not significantly stimulate l-carnitine uptake. Analysis of l-carnitine counterflow yielded an apparent external K(m) of 105 microm and a turnover number of 5.5 s(-1). Contrary to related proteins, CaiT activity was not modulated by osmotic stress. l-Carnitine binding to CaiT increased the protein fluorescence and caused a red shift in the emission maximum, an observation explained by ligand-induced conformational alterations. The fluorescence effect was specific for betaine structures, for which the distance between trimethylammonium and carboxyl groups proved to be crucial for affinity. Taken together, the results suggest that CaiT functions as an exchanger (antiporter) for l-carnitine and gamma-butyrobetaine according to the substrate/product antiport principle.     

Expression and localization of organic cation/carnitine transporter OCTN2 in Caco-2 cells

l-Carnitine is derived both from dietary sources and biosynthesis. Dietary carnitine is absorbed in the small intestine and then distributed to other organs. Previous studies using Caco-2 cells demonstrated that the transport of l-carnitine in the intestine involves a carrier-mediated system. The purpose of this study was to determine whether the uptake of l-carnitine in Caco-2 cells is mediated by the recently identified organic cation/carnitine transporter (OCTN2). Kinetics of l-[(3)H]carnitine uptake were investigated with or without specific inhibitors. l-Carnitine uptake in mature cells was sodium dependent and linear with time. K(m) and V(max) values for saturable uptake were 14.07 +/- 1.70 micro M and 26.3 +/- 0.80 pmol. mg protein(-1). 6 min(-1), respectively. l-carnitine uptake was inhibited (P < 0.05-0.01) by valproate and other organic cations. Anti-OCTN2 antibodies recognized a protein in the brush-border membrane (BBM) of Caco-2 cells with an apparent molecular mass of 60 kDa. The OCTN2 expression was confirmed by double immunostaining. Our results demonstrate that l-carnitine uptake in differentiated Caco-2 cells is primarily mediated by OCTN2, located on the BBM.