Thiamine-responsive megaloblastic anemia

Rogers, Lon E.; Porter, F. Stanley; Sidbury, James

doi:10.1016/s0022-3476(69)80031-4

Cited by 164 publications

(70 citation statements)

References 18 publications

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“…In literature many methods are reported for erotic acid including enzymatic methods [ 131, colorimetric methods without [14] and with preceding extraction [15,16], partition column chromatographic procedures [ 16,171, anion-[ 18,191 and cationexchange column chromatography [20], and an isotope dilution procedure [ 211. For erotic acid and orotidine both time consuming HPLC methods have been described, using anion-exchange columns, gradient elution and UV detection [ 22-251. In the method used by Cohen et al [9], HPLC of the urine is performed before and after treatment with OPRT and ODC, and the concentration of the compounds calculated by subtraction of the results obtained in the two analyses.…”

Section: Discussionmentioning

confidence: 99%

Urinary excretion of orotic acid, orotidine and other pyrimidines in a patient with purine nucleoside phosphorylase deficiency

Gennip

Grift

Bree

et al. 1979

Clinica Chimica Acta

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SummaryUrinary orotidine and erotic acid have been determined in a patient with purine nucleoside phosphorylase (PNP) deficiency under various dietary and therapeutic conditions. For this purpose a new procedure for the analysis of both compounds has been developed, consisting of prefractionation with Dowex 1X8, followed by two HPLC steps on a ti Bondapak NH, and a /J Bondapak C,, column. With this method normal as well as slightly elevated excretions of erotic acid have been found in our patient. No evidence was obtained for inhibition of OPRT by purine (deoxy)nucleosides as a cause of pyrimidine starvation.A significant increase of urinary orotidine was found after loading with allopurinol.For comparison excretory values in a patient with omithine transcarbamylase deficiency and also in a patient with erotic aciduria type I are shown. The possible cause of the slight increase in urinary erotic acid in our patient has been discussed.

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Section: Discussionmentioning

confidence: 99%

Urinary excretion of orotic acid, orotidine and other pyrimidines in a patient with purine nucleoside phosphorylase deficiency

Gennip

Grift

Bree

et al. 1979

Clinica Chimica Acta

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“…Thiamine is not synthesized by humans and other mammals and is obtained from dietary sources via absorption into intestinal epithelia. Deficiencies in cellular thiamine accumulation lead to cardiovascular and neurological disorders and are associated with the inherited condition of thiamine-responsive megaloblastic anemia (TRMA) 1 (1,(3)(4)(5)(6)(7). Recent molecular analyses have identified the protein product of the SLC19A2 gene in humans (chromosome 1q23.…”

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Cell Biology of the Human Thiamine Transporter-1 (hTHTR1)

Subramanian¹,

Marchant²,

Parker³

et al. 2003

Journal of Biological Chemistry

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The human thiamine transporter hTHTR1 is involved in the cellular accumulation of thiamine (vitamin B1) in many tissues. Thiamine deficiency disorders, such as thiamine-responsive megaloblastic anemia (TRMA), which is associated with specific mutations within hTHTR1, likely impairs the functionality and/or intracellular targeting of hTHTR1. Unfortunately, nothing is known about the mechanisms that control the intracellular trafficking or membrane targeting of hTHTR1. To identify molecular determinants involved in hTHTR1 targeting, we generated a series of hTHTR1 truncations fused with the green fluorescent protein and imaged the targeting and trafficking dynamics of each construct in living duodenal epithelial cells. Whereas the full-length fusion protein was functionally expressed at the plasma membrane, analysis of the truncated mutants demonstrated an essential role for both NH 2 -terminal sequence and the integrity of the backbone polypeptide for cell surface expression. Most notably, truncation of hTHTR1 within a region where several TRMA truncations are clustered resulted in intracellular retention of the mutant protein. Finally, confocal imaging of the dynamics of intracellular hTHTR1 vesicles revealed a critical role for microtubules, but not microfilaments, in hTHTR1 trafficking. Taken together, these results correlate hTHTR1 structure with cellular expression profile and reveal a critical dependence on hTHTR1 backbone integrity and microtubule-based trafficking processes for functional expression of hTHTR1.Thiamine (vitamin B1) is a water-soluble micronutrient that is essential for many cellular functions relating to growth and development. For example, thiamine pyrophosphate (the coenzyme form) is important for normal carbohydrate metabolism and energy production (1, 2). Thiamine is not synthesized by humans and other mammals and is obtained from dietary sources via absorption into intestinal epithelia. Deficiencies in cellular thiamine accumulation lead to cardiovascular and neurological disorders and are associated with the inherited condition of thiamine-responsive megaloblastic anemia (TRMA) 1 (1, 3-7). Recent molecular analyses have identified the protein product of the SLC19A2 gene in humans (chromosome 1q23.3) as the site of mutations that link with TRMA inheritance (2, 7-10). SLC19A2 encodes a saturable, high affinity human thiamine transporter known as hTHTR1 (2, 7-10), which is expressed in many tissues, including duodenal epithelium (11). The functional properties of hTHTR1 mimic the biochemical characteristics of thiamine uptake in intestinal preparations (reviewed in Refs. 12 and 13). The full-length hTHTR1 protein encodes a 497-amino acid polypeptide with 12 predicted transmembrane-spanning segments and cytoplasmic NH 2 -and COOH-terminal regions (2, 8). Several clinically identified mutations have been characterized in hTHTR1 (2, 7-10, 14), including ten prematurely truncated mutants, resulting from either point mutation, deletion, or insertion of nucleotides (reviewed in Refs. 7 and 15...

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“…[5][6][7][8][9][10][11][12][13] To date, orotic acid has been determined mostly by liquid chromatography (HPLC) with UV detection at 275 nm. 14 This approach is quite cumbersome in the subsequent integration and interpretation process.…”

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Rapid determination of orotic acid in urine by a fast liquid chromatography/tandem mass spectrometric method

Marca

Casetta²,

Zammarchi

2003

Rapid Comm Mass Spectrometry

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Quantification of orotic acid (uracil-6-carboxylic acid) in urine is an important tool to diagnose some inherited diseases, such as urea cycle disorder (OTCD) and hereditary orotic aciduria. New rapid analytical methods are necessary to provide high-throughput orotic acid analyses. A new analytical method has been developed for the rapid analysis of orotic acid in urine by liquid chromatography coupled with ion spray tandem mass spectrometry (LC/MS/MS). After a sample dilution 1:20, the analysis was performed in the selected reaction monitoring mode in which orotic acid was detected through the transition m/z 155 to 111. The retention time was 3.9 min in a 4.5-min analysis. The analysis of orotic acid (OA) is used in clinical chemistry to diagnose some diseases associated with urea cycle disorders; ornithine transcarbamylase deficiency (OTCD) 1 is the most common, and also hereditary orotic aciduria. 2 The urea cycle has three roles: it produces urea as the waste product incorporating nitrogen not used for biosynthetic purposes, it is involved in the biochemical reactions that degrade and synthesise arginine de novo, and it plays a fundamental role in pH homeostasis. 3 Ornithine transcarbamylase, a mitochondrial matrix enzyme, catalyses the synthesis of citrulline from carbamyl phosphate and ornithine. OTCD male patients show classical clinical symptoms such as vomiting, lethargy, and confusion; clinical laboratory indications are high values of glutamine and ammonia, low citrulline, and high urinary excretion of orotic acid. Also, female manifesting carriers generally have higher than normal values of urinary orotic acid excretion due to X-inactivation.Hereditary orotic aciduria is a pyrimidine synthesis defect caused by uridine-5 0 -monophosphate (UMP) synthase deficiency. 4 This multienzyme catalyses the synthesis of UMP starting from orotic acid via orotidine-5 0 -monophosphate. Some characteristic clinical features are anaemia, hematuria, pallor, diarrhoea, retarded and poor development, while laboratory indications include very high urinary orotic acid excretion.Therefore, determination of orotic acid levels in urine is becoming an assay routinely performed in clinical analysis. [5][6][7][8][9][10][11][12][13] To date, orotic acid has been determined mostly by liquid chromatography (HPLC) with UV detection at 275 nm. 14 This approach is quite cumbersome in the subsequent integration and interpretation process. Signals from a 'normal' specimen are quite difficult to quantify since the LC-UV peaks are very close to the baseline (lack of adequate sensitivity) and are sometimes not sufficiently resolved from other endogenous components owing to the nonspecific wavelength chosen for detection. In addition the chromatographic run is time demanding.In this paper we evaluate the possibility to take advantage of tandem mass spectrometry 15 (MS/MS) as the basis for a rapid method for determination of OA in urine with very simple preparation, which could be a candidate for highthroughput routine analyses. The main cr...

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Thiamine-responsive megaloblastic anemia

Cited by 164 publications

References 18 publications

Urinary excretion of orotic acid, orotidine and other pyrimidines in a patient with purine nucleoside phosphorylase deficiency

Urinary excretion of orotic acid, orotidine and other pyrimidines in a patient with purine nucleoside phosphorylase deficiency

Cell Biology of the Human Thiamine Transporter-1 (hTHTR1)

Rapid determination of orotic acid in urine by a fast liquid chromatography/tandem mass spectrometric method

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