OBJECTIVETo investigate the mutations of SLC22A5 gene in patients with systemic primary carnitine deficiency (CDSP).

METHODSHigh liquid chromatography tandem mass spectrometry (HPLC/MS/MS) was applied to screen congenital genetic metabolic disease and eight patients with CDSP were diagnosed among 77 511 samples. The SLC22A5 gene mutation was detected using massarray technology and sanger sequencing. Using SIFT and PolyPhen-2 to predict the function of protein for novel variations.

RESULTSTotal detection rate of gene mutation is 100% in the eight patients with CDSP. Seven patients had compound heterozygous mutations and one patient had homozygous mutations. Six different mutations were identified, including one nonsense mutation [c.760C>T(p.R254X)] and five missense mutations[c.51C>G(p.F17L), c.250T>A(p.Y84N), c.1195C>T(p.R399W), c.1196G>A(p.R399Q), c.1400C>G(p.S467C)]. The c.250T>A(p.Y84N) was a novel variation, the novel variation was predicted to have affected protein structure and function. The c.760C>T (p.R254X)was the most frequently seen mutation, which was followed by the c.1400C>G(p.S467C).

CONCLUSIONThis study confirmed the diagnosis of eight patients with CDSP on the gene level. Six mutations were found in the SLC22A5 gene, including one novel mutation which expanded the mutational spectrum of the SLC22A5 gene.

Adult ; Amino Acid Sequence ; Base Sequence ; Cardiomyopathies ; diagnosis ; genetics ; metabolism ; Carnitine ; deficiency ; genetics ; metabolism ; DNA Mutational Analysis ; methods ; Female ; Gene Frequency ; Genotype ; Humans ; Hyperammonemia ; diagnosis ; genetics ; metabolism ; Infant, Newborn ; Male ; Muscular Diseases ; diagnosis ; genetics ; metabolism ; Mutation ; Organic Cation Transport Proteins ; genetics ; metabolism ; Reproducibility of Results ; Sensitivity and Specificity ; Sequence Homology, Amino Acid ; Solute Carrier Family 22 Member 5 ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Objective: To investigate the expressions of solute carrier family 22 member 14 (SLC22A14) and sperm-associated antigen 6 (SPAG6) in the sperm of idiopathic asthenospermia men. METHODS: We collected semen samples from 50 idiopathic asthenozoospermia patients and another 50 normal sperm donors, purified the sperm by discontinuous density centrifugation on Percoll gradients, and then determined the mRNA and protein expressions of SLC22A14 and SPAG6 by RT-PCR and Western blot, respectively. RESULTS: Compared with the normal controls, the idiopathic asthenozoospermia patients showed significantly decreased mRNA expressions of SLC22A14 (0.77 ± 0.08 vs 0.53 ± 0.10, P<0.01) and SPAG6 (0.78 ± 0.09 vs0.52 ± 0.10 , P<0.01) and protein expressions of SLC22A14 (0.80 ± 0.09 vs 0.55 ± 0.10 , P<0.01) and SPAG6 (0.78 ± 0.09 vs 0.56 ± 0.09, P<0.01). CONCLUSIONS T The expressions of SLC22A14 and SPAG6 are reduced in the sperm of the patients with idiopathic asthenospermia, which may be one of the important causes of asthenospermia.
Asthenozoospermia ; metabolism ; Blotting, Western ; Ejaculation ; Humans ; Male ; Microtubule Proteins ; genetics ; metabolism ; Organic Cation Transport Proteins ; genetics ; metabolism ; Proteomics ; RNA, Messenger ; metabolism ; Sperm Motility ; Spermatozoa ; metabolism

OBJECTIVETo identify potential mutation of SLC22A5 gene in a 5-month-old boy affected with primary carnitine deficiency and provide genetic counseling and prenatal diagnosis for the members of his family.

METHODSDNA was extracted from peripheral blood samples derived from the proband, his parents and elder sister, as well as amniotic fluid from his pregnant mother. All of the 10 exons of the SLC22A5 gene were amplified by PCR and subjected to Sanger sequencing. The amniotic fluid sample was also subjected to G-banded karyotyping and multiplex ligation-dependent probe amplification (MLPA).

RESULTSA homozygous mutation c.760C>T (p.R254X) of the SLC22A5 gene was detected in the proband. Heterozygous mutation c.760C>T (p.R254X) was also found in other family members including the fetus. The karyotyping and chromosomal microdeletion testing for the amniotic fluid sample were both normal.

CONCLUSIONThe newly identified homozygous nonsense c.760C>T (p.R254X) mutation of the SLC22A5 gene probably underlies the primary carnitine deficiency of the proband. Genetic counseling and prenatal diagnosis have been provided for this family.

Adult ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; Cardiomyopathies ; embryology ; genetics ; Carnitine ; deficiency ; genetics ; China ; Exons ; Female ; Genotype ; Humans ; Hyperammonemia ; embryology ; genetics ; Infant ; Male ; Molecular Sequence Data ; Muscular Diseases ; embryology ; genetics ; Organic Cation Transport Proteins ; genetics ; Pedigree ; Pregnancy ; Prenatal Diagnosis ; Solute Carrier Family 22 Member 5

To establish single- and double-transfected transgenic cells stably expressing hMATE1, hMATE1 cDNA was cloned by RT-PCR from human cryopreserved kidney tissue, and subcloned into pcDNA3.1(+) plasmid by virtue of both HindIII and Kpn I restriction enzyme sites. Subsequently, the recombined pcDNA3.1(+)- hMATE1 plasmid was transfected into MDCK, MDCK-hOCT1 or MDCK-hOCT2 cells using Lipofectamine 2000 Reagent. After a 14-day-cultivation with hygromycin B at the concentration of 400 µg · mL(-1), all clones were screened with DAPI and MPP+ as substrates to identify the best candidate. The mRNA content of hMATE1, the cellular accumulation of metformin with or without cimetidine as inhibitor, or transportation of cimetidine was further valuated. The results showed that all of the three cell models over expressed hMATE1 mRNA. The cellular accumulation of metformin in MDCK-hMATE1 was 17.6 folds of the control cell, which was significantly inhibited by 100 µmol · L(-1) cimetidine. The transcellular transport parameter net efflux ratios of cimetidine across MDCK-hOCT1/hMATE1 and MDCK-hOCT2/hMATE1 monolayer were 17.5 and 3.65, respectively. In conclusion, cell models with good hMATE1 function have been established successfully, which can be applied to study the drug transport or drug-drug interaction involving hMATE1 alone or together with hOCT1/2 in vitro.
Animals ; Biological Transport ; Cimetidine ; pharmacology ; DNA, Complementary ; Dogs ; Drug Interactions ; Humans ; Madin Darby Canine Kidney Cells ; Metformin ; pharmacology ; Organic Cation Transport Proteins ; genetics ; metabolism ; Transfection

OBJECTIVETo investigate the mutation and background of SLC22A5 in 6 patients with primary carnitine deficiency (PCD) who only presented as cardiomyopathy.

METHODGenomic DNA were abstracted from the blood of the patients and their parents. Using high-throughput sequencing to determine the mutation site.Using Sanger method to confirm the mutated alleles in PCD patients and detect the corresponding sequences in their patients. Using SIFT and PolyPhen to predict the function of protein for detected missense mutations.

RESULTThree different mutations were identified, including 2 nonsense mutations (R254X and R289X), 1 missense mutation (C113Y), R254X was the most frequently seen mutation. Four patients had compound heterozygous mutations and 2 patients had homozygous mutations. Their parents were found to have heterozygous mutations in corresponding alleles.

CONCLUSIONR254X, R289X and C113Y might be associated with primary carnitine deficiency.

Adolescent ; Base Sequence ; Cardiomyopathies ; genetics ; Carnitine ; deficiency ; genetics ; Child ; Child, Preschool ; DNA Mutational Analysis ; Female ; Genotype ; Heterozygote ; High-Throughput Nucleotide Sequencing ; Humans ; Hyperammonemia ; genetics ; Infant ; Male ; Muscular Diseases ; genetics ; Mutation ; Organic Cation Transport Proteins ; genetics ; Pedigree ; Solute Carrier Family 22 Member 5

OBJECTIVETo identify pathogenic mutation in a boy affected with riboflavin responsive-multiple acyl-CoA dehydrogenase deficiency (RR-MADD).

METHODSThe patient was initially diagnosed as primary carnitine deficiency (PCD) and has been treated with carnitine supplementation for 7 years. Clinical manifestations and characteristics of fibula muscle specimen were analyzed. Potential mutation in electron transfer flavoprotein dehydrogenase (ETFDH) gene (for the patient and his parents) and carnitine transfer protein gene (SLC22A5) (for the patient) was screened.

RESULTSElectronic microscopy of the muscle specimen has suggested lipid storage myopathy. Mutation analysis has found that the patient carried compound heterozygous mutations, c.250G>A and c.380T>C, in exon 3 of the ETFDH gene, whilst his father and mother were heterozygous for the c.380T>C and c.250G>A mutations, respectively. Screening of the SLC22A5 gene has yielded no clinically meaningful result. After the establishment of diagnosis of RR-MADD, the condition of the patient has improved greatly with supplementation of high doses of riboflavin along with continuous carnitine supplement.

CONCLUSIONThe c.250G>A (p.Ala84Thr) mutation of exon 3 of the ETFDH gene has been a hot spot in Southern Chinese population, whilst the c.380T>C (p.Leu127Pro) is rarely reported. Our case has suggested that therapeutic diagnosis cannot substitute genetic testing. The mechanism for having stabilized the patient with only carnitine supplementation for 7 years needs further investigation.

Adolescent ; Adult ; Base Sequence ; Child ; DNA Mutational Analysis ; Electron-Transferring Flavoproteins ; genetics ; metabolism ; Female ; Humans ; Iron-Sulfur Proteins ; genetics ; metabolism ; Male ; Molecular Sequence Data ; Multiple Acyl Coenzyme A Dehydrogenase Deficiency ; enzymology ; genetics ; metabolism ; Muscle, Skeletal ; metabolism ; Organic Cation Transport Proteins ; genetics ; metabolism ; Oxidoreductases Acting on CH-NH Group Donors ; genetics ; metabolism ; Riboflavin ; metabolism ; Solute Carrier Family 22 Member 5

OBJECTIVEMany children were found to have low free carnitine level in blood by tandem mass spectrometry technology. In some of the cases the problems occurred secondary to malnutrition, organic acidemia and other fatty acid oxidation metabolic diseases, and some of cases had primary carnitine deficiency (PCD). In the present article, we discuss the diagnosis of PCD and evaluate the efficacy of carnitine in the treatment of PCD.

METHODWe measured the free carnitine (C0) and acylcarnitine levels in the blood of 270 000 neonates from newborns screening program and 12 000 children with suspected clinical inherited metabolic diseases by tandem mass spectrometry. The mutations of carnitine transporter protein were tested to the children with low C0 level and the diagnosis was made. The children with PCD were treated with 100 - 300 mg/kg of carnitine.

RESULTSeventeen children were diagnosed with PCD, 6 from newborn screening program and 11 from clinical patients. Mutations were found in all of them. The average C0 level [(2.9 ± 2.0) µmol/L] in patients was lower than the reference value (10 µmol/L), along with decreased level of different acylcarnitines. The clinical manifestations were diverse. For the 6 patients from newborn screening, 4 were asymptomatic, 1 showed hypoglycaemia and 1 showed movement intolerance from 2 years of age. For the 11 clinical patients, 8 showed hepatomegaly, 7 showed myasthenia, 6 showed cardiomyopathy, 1 showed chronic abdominal pain, and 1 showed restlessness and learning difficulty. Among these patients, 14 cases were treated with carnitine. Their clinical symptoms disappeared 1 to 3 months later. The C0 level in the blood rose to normal, with the average from (4.0 ± 2.7) µmol/L to (20.6 ± 8.3) µmol/L (P < 0.01). However, the level was still lower than the average level of healthy children [(27.1 ± 4.5) µmol/L, P < 0.01].

CONCLUSIONSeventeen patients were diagnosed with PCD by the test levels of free carnitine and acylcarnitines in blood with tandem mass spectrometry, and gene mutation test. Large dose of carnitine had a good effect in treatment of the PCD patients.

Cardiomyopathies ; diagnosis ; drug therapy ; genetics ; Carnitine ; analogs & derivatives ; blood ; deficiency ; genetics ; Child, Preschool ; DNA Mutational Analysis ; Female ; Follow-Up Studies ; Humans ; Hyperammonemia ; diagnosis ; drug therapy ; genetics ; Infant ; Infant, Newborn ; Male ; Muscular Diseases ; diagnosis ; drug therapy ; genetics ; Mutation ; Neonatal Screening ; methods ; Organic Cation Transport Proteins ; deficiency ; genetics ; Reference Values ; Tandem Mass Spectrometry

OBJECTIVETo investigate the association of prostate cancer (PCa) with PDLIM5 (rs17021918, T), SLC22A3 (rs9364554, C) and NKX3-1 (rs1512268, A) in Chinese men.

METHODSWe included 124 PCa patients and 138 normal controls in this study, compared the alleles and genotypes of PDLIM5 (rs17021918, T) , SLC22A3 (rs9364554, C) and NKX3-1 (rs1512268, A) of the two groups, and explored the association of each of the genes with the age, body mass index (BMI), Gleason score, PSA level and tumor stage of the patients. We analyzed the gene-gene interaction using the multifactor dimensionality reduction method (MDR).

RESULTSThere were no statistically significant differences in the frequency distribution of the risk alleles and genotypes of PDLIM5, SLC22A3 and NKX3-1 between the case and control groups (P > 0.05), nor were the three gene loci significantly associated with the age, Gleason score, PSA level and pathological grade of the PCa patients (CP < 0.05). MDR analysis showed no interaction between PDLIM5 and NKX3-1, but tree-diagram analysis revealed a possible synergistic action of the two polymorphism loci.

CONCLUSIONPCa might not be associated with PDLIM5 (rs17021918,T), SLC22A3 (rs9364554,C) and NKX3-1 (rs1512268,A) in Chinese men. However, PDLIM5 and NKX3-1 might have a synergistic action on the risk PCa.

Adaptor Proteins, Signal Transducing ; genetics ; Aged ; Aged, 80 and over ; Alleles ; Case-Control Studies ; Genotype ; Homeodomain Proteins ; genetics ; Humans ; LIM Domain Proteins ; genetics ; Male ; Middle Aged ; Organic Cation Transport Proteins ; genetics ; Polymorphism, Single Nucleotide ; Prostatic Neoplasms ; genetics ; Risk Factors ; Transcription Factors ; genetics

Liver is regarded as one of the most important organs for drug clearance in the body, which mediates both the metabolism and biliary excretion of drugs. Transporters are a class of functional membrane proteins and control the movement of substances into or out of cells. Transporters, which are extensively expressed in the liver, play important roles in the drug hepatic disposition by regulating the uptake of drugs from blood into hepatocytes or the efflux of drugs and their metabolites into bile. In this review, the localization, functions and substrate selectivity of the major transporters in the liver will be summarized, and the impacts of these transporters on drug hepatic disposition, the potential drug-drug interactions as well as their genetic polymorphisms will also be reviewed.
ATP Binding Cassette Transporter, Sub-Family G, Member 2 ; ATP-Binding Cassette Transporters ; genetics ; metabolism ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Bile ; metabolism ; Biological Transport ; Drug Interactions ; Humans ; Liver ; metabolism ; Membrane Transport Proteins ; genetics ; metabolism ; Metabolic Clearance Rate ; Multidrug Resistance-Associated Proteins ; genetics ; metabolism ; Neoplasm Proteins ; genetics ; metabolism ; Organic Anion Transporters ; genetics ; metabolism ; Organic Anion Transporters, Sodium-Dependent ; metabolism ; Organic Anion Transporters, Sodium-Independent ; genetics ; metabolism ; Organic Cation Transport Proteins ; genetics ; metabolism ; Pharmacokinetics ; Polymorphism, Genetic ; Symporters ; metabolism

Acute renal failure with severe loin pain which develops after anaerobic exercise is rare. One of predisposing factors of exercise-induced acute renal failure is renal hypouricemia. Idiopathic renal hypouricemia is a genetic disorder characterized by hypouricemia with abnormally high renal tubular uric acid excretion. The mutation in SCL22A12 gene which encodes renal uric acid transporter, URAT1, is the known major cause of this disorder. We here described a 25-yr-old man showing idiopathic renal hypouricemia with G774A mutation in SCL22A12 who presented exercise-induced acute renal failure. There have been a few reports of mutational analysis in Korean idiopathic renal hypouricemia without acute renal failure. This is the first report of genetically diagnosed idiopathic renal hypouricemia with exercise-induced acute renal failure in Korea.
Acute Kidney Injury/*diagnosis/genetics ; Adult ; Amino Acid Substitution ; DNA Mutational Analysis ; Exercise ; Exons ; Humans ; Male ; Mutation ; Organic Anion Transporters/*genetics ; Organic Cation Transport Proteins/*genetics ; Renal Tubular Transport, Inborn Errors/etiology/*genetics ; Urinary Calculi/etiology/*genetics