Urinary Loss Of Carnitine Research Articles

Carnitine Transporter Deficiency

Primary carnitine deficiency or carnitine transporter deficiency is an inborn error of fatty acid transportation caused by a defect in carnitine transport across plasma membrane and impairs the entry of long-chain fatty acids into the mitochondrial matrix. Leading to a variety of symptoms such as, cardiomyopathy, muscle weakness, non ketotic hypoglycemia and hepatic involvement. Acute episodes may be induced by metabolic stress such as prolong fasting, infections or vomiting. Pathophysiology: Primary carnitine deficiency is caused by a defect in the plasma membrane carnitine transporter, with urinary loss of carnitine causing systemic Intracellular carnitine deficiency that impairs the entry of long-chain fatty acids into the mitochondrial matrix. Consequently, impairs the production of ketone bodies due to defect of beta-oxidation and energy production. Signs and symptoms: The presentation of patient is varied, from asymptomatic to lethal cardiac manifestations. May be present with non ketotic hypoglycemia encephalopathy, seizure, apnea, myopathy, liver dysfunction and cardiomyopathy Diagnosis: Primary carnitine deficiency must be considered in every child with emergency situation such as seizures, apnea, coma, cardiomyopathy, or episode of hypoketotic hypoglycemia, encephalopathy and Reye syndrome presentation. Hyperuricemia and Elevated Ammonia and liver transaminases and serum CK levels may be observed in primary carnitine deficiency. On the other hand several patients with primary carnitine deficiency have been detected by newborn screening programs. Enzyme and Carnitine transport assay in cultured fibroblasts and Mutation analysis are methods for confirmation of diagnosis. Also in imaging study chest x Ray reveals cardiac enlargement and in the echocardiogram cardiac enlargement and increased thickness of the left ventricular wall. ECG reveals left ventricular hypertrophy and peaked T waves in primary carnitine deficiency. Treatment: In acute attack with hypoketotic hypoglycemic encephalopathy, start with 4cc/kg D/W 10% and then infusion of serum D/W 10%. IV carnitine 100 to 200 mg/kg/day in acute attack and continue with oral carnitine. Management of cardiomyopathy Oral carnitine therapy in primary carnitine deficiency improves, cardiac function, fasting ketogenesis , growth, and cognitive performance.

Inhibition of OCTN2-Mediated Transport of Carnitine by Etoposide

OCTN2 is a bifunctional transporter that reabsorbs filtered carnitine in a sodium-dependent manner and secretes organic cations into urine as a proton antiport mechanism. We hypothesized that inhibition of OCTN2 by anticancer drugs can influence carnitine resorption. OCTN2-mediated transport inhibition by anticancer drugs was assessed using cells transfected with human OCTN2 (hOCTN2) or mouse Octn2 (mOctn2). Excretion of carnitine and acetylcarnitine was measured in urine collected from mice and pediatric patients with cancer before and after administration of etoposide. Five of 27 tested drugs (50-100 μmol/L) inhibited hOCTN2-mediated carnitine uptake by 42% to 85% (P < 0.001). Of these inhibitors, etoposide was itself a transported substrate of hOCTN2 and mOctn2. Etoposide uptake by hOCTN2 was reversed in the presence of excess carnitine. This competitive inhibitory mechanism was confirmed in an in silico molecular docking analysis. In addition, etoposide inhibited the transcellular apical-to-basolateral flux of carnitine in kidney cells. Etoposide was also associated with a significant urinary loss of carnitine in mice (~1.5-fold) and in patients with cancer (~2.4-fold). Collectively, these findings indicate that etoposide can inhibit hOCTN2 function, potentially disturb carnitine homeostasis, and that this phenomenon can contribute to treatment-related toxicities.

Cisplatin-Induced Downregulation of OCTN2 Affects Carnitine Wasting

Carnitine is an essential cofactor for mitochondrial fatty acid oxidation that is actively reabsorbed by the luminal transporter Octn2 (Slc22a5). Because the nephrotoxic agent cisplatin causes urinary loss of carnitine in humans, we hypothesized that cisplatin may affect Octn2 function. Excretion of carnitine and acetylcarnitine was measured in urine collected from mice with or without cisplatin administration. The transport of carnitine was assessed in cells that were transfected with OCT1 or OCT2. The effect of cisplatin treatment on gene expression was analyzed using a mouse GeneChip array and validated using quantitative reverse transcriptase-PCR. In wild-type mice, urinary carnitine excretion at baseline was ∼3-fold higher than in mice lacking the basolateral cisplatin transporters Oct1 and Oct2 [Oct1/2(-/-) mice], indicating that carnitine itself undergoes basolateral uptake into the kidney. Transport of carnitine by OCT2, but not OCT1, was confirmed in transfected cells. We also found that cisplatin caused an increase in the urinary excretion of carnitine and acetylcarnitine in wild-type mice but not in Oct1/2(-/-) mice, suggesting that tubular transport of cisplatin is a prerequisite for this phenomenon. Cisplatin did not directly inhibit the transport of carnitine by Octn2 but downregulated multiple target genes of the transcription factor peroxisome proliferator activated receptor α, including Slc22a5, in the kidney of wild-type mice that were absent in Oct1/2(-/-) mice. Our study shows a pivotal role of Oct1 and Oct2 in cisplatin-related disturbances in carnitine homeostasis. We postulate that this phenomenon is triggered by deactivation of peroxisome proliferator activated receptor α and leads to deregulation of carnitine-shuttle genes.

Highlights of This Issue

Oxygen levels can dictate the expression of many genes, potentially altering immunogenic properties important for tumor cell–based vaccines. In this issue, Olin and colleagues demonstrate that in murine models of glioma and breast carcinoma, 5% O2-derived vaccines could enhance in vivo tumoricidal activity, resulting in extended survival. In the glioma cavity of mice treated with lysate vaccines from 5% O2, the induction of caspase-3 in glioma cells by multiple CD3+ lymphocytes was observed. These findings have important implications for the development of cancer vaccines, which are typically taken from cells cultured in atmospheric oxygen (about 20% O2).A better understanding of the functional differences between HIF-1α and HIF-2α in oral squamous cell carcinoma (OSCC) is needed. Zhu and colleagues evaluated correlations between HIF-1/2α expression and clinical-pathological characteristics of OSCC patients, investigated HIF-1α/2α-dependent target genes in OSCC cell lines, and examined the contribution of HIF-1α/2α to xenograft tumor growth, angiogenesis, and apoptosis. HIF-1α and HIF-2α correlated with different clinical-pathological parameters, stabilized at different oxygen levels, and regulated different genes in OSCC. However, both HIF-1α and HIF-2α had promoting roles in tumor angiogenesis and growth, suggesting that combined targeting of HIF-1α and HIF-2α may improve outcome.Carnitine (an essential cofactor for mitochondrial fatty acid oxidation) is reabsorbed by the luminal transporter OCTN2. In humans, cisplatin causes urinary loss of carnitine. To determine the effect cisplatin has on OCTN2 function, Lancaster and colleagues measured carnitine and acetylcarnitine excretion in urine collected from mice with/without cisplatin administration. Cisplatin caused increased urinary excretion of carnitine and acetylcarnitine in wild-type mice but not in Oct1/2(-/-) mice. Gene expression analysis on untreated/treated kidneys showed multiple genes in wild-type mice were downregulated compared to Oct1/2(-/-) mice, including those involved in carnitine shuttling. These findings highlight the role of Oct1/2 in cisplatin-related disturbances in carnitine homeostasis.Lung squamous cell carcinoma (SCC) has broad clinical, genetic, and morphologic heterogeneity. Wilkerson and colleagues developed a robust SCC subclassification to describe this variability. They analyzed gene expression arrays from over 400 patients to detect and validate novel gene expression subtypes. The subtypes were found to have significantly different survival outcomes, patient populations, biological processes, and similarities to normal lung cell types. Further, the subtypes stratified heterogeneous SCC into multiple discrete diseases, which could lead to more precise patient prognosis and management, and to advances in basic research.

Abstract 3529: Cisplatin-induced downregulation of OCTN2 affects carnitine (vitamin BT) wasting

Abstract Carnitine is an essential cofactor for mitochondrial fatty acid oxidation. It is normally maintained at a steady level in the blood, and is retained in the body by a rescue mechanism that involves renal tubular reabsorption of filtered carnitine by OCTN2, a luminal organic cation transporter. The physiological significance of OCTN2 has been confirmed by the identification of mutations that cause a potentially lethal autosomal recessive disease known as primary systemic carnitine deficiency. Heterozygosity for OCTN2 mutations is associated with an intermediate carnitine-deficiency phenotype, suggesting that even partial loss of transporter function may be detrimental. The nephrotoxic agent cisplatin was previously reported to cause urinary loss of carnitine in cancer patients, although the mechanism by which this occurs remains unknown. We hypothesized that cisplatin may influence the function of OCTN2 and/or catalytic enzymes involved in carnitine homeostasis. In wildtype mice, we found that the urinary excretion of carnitine and acetylcarnitine at baseline over a 24-hour period was about ∼3 fold higher than in mice lacking the organic cation transporters Oct1 and Oct2 [Oct1/2(−/−) mice] (P&amp;lt;0.001), indicating that carnitine may undergo carrier-mediated basolateral uptake in renal tubular cells. Transport of carnitine (50 nM) was confirmed in 293 Flp-In cells overexpressing OCT2 (P=0.0034), and carnitine (1 mM) itself caused approximately 40% inhibition of OCT2 function as assessed by changes in transport of [14C]tetraethylammonium, a known OCT2 substrate. In wildtype mice, a single dose of cisplatin (10 mg/kg, i.p) caused a time-dependent increase in urinary carnitine excretion on days 2 and 3 (2-3 fold), and a 7-14 fold increase in acetylcarnitine excretion on days 1-3 post-treatment. Cisplatin-induced changes in urinary carnitine and acetylcarnitine were not observed in Oct1/2(−/−) mice, suggesting that renal tubular transport of cisplatin by Oct1 and Oct2 is a prerequisite for treatment-related disturbances in carnitine excretion. One possible explanation for this phenomenon involves inhibition of OCTN2 activity by cisplatin. However, cisplatin and a mixture of cisplatin-glutathione conjugates (100 µM; 30 min) did not substantially affect carnitine transport in HEK293 cells stably transfected with OCTN2. Next, we considered the possibility that cisplatin affects the expression of Octn2 and/or carnitine-palmitoyltransferases (Cpt1/2) involved in the formation of acetylcarnitine. In wildtype mice, mRNA expression analysis in kidney biopsies following treatment with cisplatin indicated a dramatic downregulation of Octn2 as well as upregulation of Cpt1b in a time-dependent manner, which changes were absent in the Oct1/2(−/−) mice. This study suggests that carnitine wasting associated with cisplatin chemotherapy is at least partially dependent on drug-induced downregulation of OCTN2 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3529.

Pregnancy increases urinary loss of carnitine and reduces plasma carnitine in Korean women

This study compared plasma and urinary carnitine concentrations in pregnant and non-pregnant Korean women. The subjects were fifty pregnant women and thirty non-pregnant women aged 24-28 years. During the first trimester, dietary carnitine intakes in the pregnant women were much lower than in non-pregnant women (70.00 (SD 29.22) micromol/d), but over the course of pregnancy carnitine intake increased from 44.64 (SD 24.84) micromol/d during the first trimester to 96.11 (SD 36.56) micromol/d during the third trimester. Pregnant women had a significantly lower plasma carnitine concentration than non-pregnant women. Plasma concentrations of non-esterified carnitine, acid-soluble acylcarnitine and total carnitine were significantly lower during the second and third trimesters than the first. Plasma acid-insoluble acylcarnitine levels, which tended to be higher in the non-pregnant women compared with the pregnant women, increased significantly as gestation proceeded. The urinary excretion of non-esterified carnitine, acid-soluble acylcarnitine and total carnitine was significantly higher in the pregnant women during the first and second trimesters than in non-pregnant women and decreased significantly as gestation proceeded. We found that there was a significant decrease in plasma carnitine level even though dietary carnitine intake increased as gestation proceeded. The low urinary excretion of carnitine in late pregnancy may be caused by an increased demand during pregnancy.

Defective urinary carnitine transport in heterozygotes for primary carnitine deficiency

PurposePrimary carnitine deficiency is an autosomal recessive disorder caused by defective carnitine transport and manifests as nonketotic hypoglycemia or skeletal or heart myopathy. MethodsTo define the mechanisms producing partially reduced plasma carnitine levels in the parents of affected patients, we examined carnitine transport in vivo and in the fibroblasts of a new patient and his heterozygous parents. ResultsKinetic analysis of carnitine transport in fibroblasts revealed an absence of saturable carnitine transport in the proband's cells and a partially impaired carnitine transport in fibroblasts from both parents, whose cells retained normal Km values toward carnitine (6–9 μM) but reduced Vmax. At steady state, normal fibroblasts accumulated carnitine to a concentration that was up to 80 times the extracellular value (0.5 μM). By contrast, cells from the proband had minimal carnitine accumulation, and cells from both parents had intermediate values of carnitine accumulation. Plasma carnitine levels were slightly below normal in both heterozygous, yet clinically normal, parents and in the paternal grandfather and the maternal grandmother. To define the mechanism producing partially decreased carnitine levels, we studied urinary carnitine losses in heterozygous parents compared with controls. Urinary losses increased linearly (P < 0.05) with plasma carnitine levels in normal controls. When urinary carnitine losses were normalized to plasma carnitine levels, a significant difference was observed between controls and heterozygous individuals (P < 0.01). ConclusionsThese results indicate that fibroblasts from heterozygotes for primary carnitine deficiency have a decreased capacity to accumulate carnitine and that heterozygotes have increased urinary losses, which may contribute to their reduced plasma carnitine levels.

Effect of cyclosporine treatment on carnitine and myo-inositol in diabetic rats

1. The effect of long-term (20 wk) treatment of cyclosporine A (CyA) was studied in urine, blood, liver, kidney and pancreatic concentrations of acid-soluble carnitine and free myo-inositol in streptozotocin diabetic rats. 2. Diabetic rats excreted significantly higher concentrations of carnitine and myo-inositol; CyA prevented the urinary loss of carnitine but not myo-inositol. 3. Blood carnitine levels were not different between normal and diabetic rats, however, CyA significantly decreased these levels. Conversely, blood myo-inositol concentrations were higher in diabetic than in normal rats; CyA prevented this increase. 4. Hepatic concentrations of both carnitine and myo-inositol were increased in diabetic rats; CyA treatment caused even further increase. 5. Pancreas from diabetic rats contained less carnitine and myo-inositol compared to normal pancreas. CyA treatment did not affect pancreatic carnitine, but it normalized myo-inositol in diabetic rats. 6. The kidney carnitine or myo-inositol levels were not influenced either by diabetes or by CyA treatment. 7. These results suggest that CyA treatment causes changes in carnitine and myo-inositol concentrations in biologic fluids and certain tissues.

Plasma carnitine levels and urinary carnitine excretion during sepsis.

Carnitine is an indispensable factor for the beta-oxidation of medium- and long-chain fatty acids, and it plays a possible role in the oxidation of branched-chain amino acids. Plasma and urinary levels of free carnitine and short-chain acyl-carnitines were studied in 67 surgical patients, after non-septic surgical procedures or during sepsis. The septic state was associated with increased urinary excretion of free carnitine (p less than 0.001), as well as with lower plasma levels of short-chain acyl-carnitines (p less than 0.001); the latter feature correlated with the level of hypermetabolism, as evaluated by the metabolic rate and by the arterial-mixed venous O2 difference. In 26 patients during total parenteral nutrition D, L-acetyl-carnitine was administered (100 mg/kg/24 hrs, in continuous iv infusion) and was associated, in septic patients only, with a significant decrease in the respiratory quotient, suggesting enhanced oxidation of low respiratory quotient substrates (fatty acids and/or branched-chain amino acids). Carnitine supplementation during total parenteral nutrition might be of theoretical benefit in some clinical conditions, such as sepsis, in which the following conditions coexist enhanced utilization of substrates whose oxidation is partially or totally carnitine dependent; prolonged absence of exogenous intake of carnitine (as in long-term total parenteral nutrition); eventual impairment of carnitine synthesis due to hepatic dysfunction; increased, massive urinary loss of carnitine.