Maleic acid is a biomarker for maleylacetoacetate isomerase deficiency; implications for newborn screening of tyrosinemia type 1.

Dried blood spot succinylacetone (SA) is often used as a biomarker for newborn screening (NBS) for tyrosinemia type 1 (TT1). However, false-positive SA results are often observed. Elevated SA may also be due to maleylacetoacetate isomerase deficiency (MAAI-D), which appears to be clinically insignificant. This study investigated whether urine organic acid (uOA) and quantitative urine maleic acid (Q-uMA) analyses can distinguish between TT1 and MAAI-D. We reevaluated/measured uOA (GC-MS) and/or Q-uMA (LC-MS/MS) in available urine samples of nine referred newborns (2 TT1, 7 false-positive), eight genetically confirmed MAAI-D children, and 66 controls. Maleic acid was elevated in uOA of 5/7 false-positive newborns and in the three available samples of confirmed MAAI-D children, but not in TT1 patients. Q-uMA ranged from not detectable to 1.16 mmol/mol creatinine in controls (n = 66) and from 0.95 to 192.06 mmol/mol creatinine in false-positive newborns and MAAI-D children (n = 10). MAAI-D was genetically confirmed in 4/7 false-positive newborns, all with elevated Q-uMA, and rejected in the two newborns with normal Q-uMA. No sample was available for genetic analysis of the last false-positive infant with elevated Q-uMA. Our study shows that MAAI-D is a recognizable cause of false-positive TT1 NBS results. Elevated urine maleic acid excretion seems highly effective in discriminating MAAI-D from TT1.

Rapid quantification of underivatized amino acids in plasma by hydrophilic interaction liquid chromatography (HILIC) coupled with tandem mass-spectrometry.

BACKGROUND: Amino acidopathies are a class of inborn errors of metabolism (IEM) that can be diagnosed by analysis of amino acids (AA) in plasma. Current strategies for AA analysis include cation exchange HPLC with post-column ninhydrin derivatization, GC-MS, and LC-MS/MS-related methods. Major drawbacks of the current methods are time-consuming procedures, derivative problems, problems with retention, and MS-sensitivity. The use of hydrophilic interaction liquid chromatography (HILIC) columns is an ideal separation mode for hydrophilic compounds like AA. Here we report a HILIC-method for analysis of 36 underivatized AA in plasma to detect defects in AA metabolism that overcomes the major drawbacks of other methods. METHODS: A rapid, sensitive, and specific method was developed for the analysis of AA in plasma without derivatization using HILIC coupled with tandem mass-spectrometry (Xevo TQ, Waters). RESULTS: Excellent separation of 36 AA (24 quantitative/12 qualitative) in plasma was achieved on an Acquity BEH Amide column (2.1×100 mm, 1.7 µm) in a single MS run of 18 min. Plasma of patients with a known IEM in AA metabolism was analyzed and all patients were correctly identified. CONCLUSION: The reported method analyzes 36 AA in plasma within 18 min and provides baseline separation of isomeric AA such as leucine and isoleucine. No separation was obtained for isoleucine and allo-isoleucine. The method is applicable to study defects in AA metabolism in plasma.

[Metabolic diseases in psychiatry]. / Stofwisselingsziekten binnen de psychiatrie.

Metabolic diseases can be associated with psychiatric symptoms. We present two case histories that demonstrate the importance of correctly diagnosing a metabolic disease as being the cause of psychiatric symptoms. We also discuss which symptoms or signals may indicate a metabolic disease.

Equine acquired multiple acyl-CoA dehydrogenase deficiency (MADD) in 14 horses associated with ingestion of Maple leaves (Acer pseudoplatanus) covered with European tar spot (Rhytisma acerinum).

This case-series describes fourteen horses suspected of equine acquired multiple acyl-CoA dehydrogenase deficiency (MADD) also known as atypical myopathy of which seven cases were confirmed biochemically with all horses having had access to leaves of the Maple tree (Acer pseudoplatanus) covered with European tar spot (Rhytisma acerinum). Assessment of organic acids, glycine conjugates, and acylcarnitines in urine was regarded as gold standard in the biochemical diagnosis of equine acquired multiple acyl-CoA dehydrogenase deficiency.

Acquired multiple Acyl-CoA dehydrogenase deficiency in 10 horses with atypical myopathy.

The aim of the current study was to assess lipid metabolism in horses with atypical myopathy. Urine samples from 10 cases were subjected to analysis of organic acids, glycine conjugates, and acylcarnitines revealing increased mean excretion of lactic acid, ethylmalonic acid, 2-methylsuccinic acid, butyrylglycine, (iso)valerylglycine, hexanoylglycine, free carnitine, C2-, C3-, C4-, C5-, C6-, C8-, C8:1-, C10:1-, and C10:2-carnitine as compared with 15 control horses (12 healthy and three with acute myopathy due to other causes). Analysis of plasma revealed similar results for these predominantly short-chain acylcarnitines. Furthermore, measurement of dehydrogenase activities in lateral vastus muscle from one horse with atypical myopathy indeed showed deficiencies of short-chain acyl-CoA dehydrogenase (0.66 as compared with 2.27 and 2.48 in two controls), medium-chain acyl-CoA dehydrogenase (0.36 as compared with 4.31 and 4.82 in two controls) and isovaleryl-CoA dehydrogenase (0.74 as compared with 1.43 and 1.61 nmol min(-1) mg(-1) in two controls). A deficiency of several mitochondrial dehydrogenases that utilize flavin adenine dinucleotide as cofactor including the acyl-CoA dehydrogenases of fatty acid beta-oxidation, and enzymes that degrade the CoA-esters of glutaric acid, isovaleric acid, 2-methylbutyric acid, isobutyric acid, and sarcosine was suspected in 10 out of 10 cases as the possible etiology for a highly fatal and prevalent toxic equine muscle disease similar to the combined metabolic derangements seen in human multiple acyl-CoA dehydrogenase deficiency also known as glutaric acidemia type II.

Elevated cholesterol precursors other than cholestanol can also be a hallmark for CTX.

Cerebrotendinous xanthomatosis (CTX) is an inborn error of bile acid synthesis in which hepatic conversion of cholesterol to cholic and chenodeoxycholic acids is impaired. Patients have abnormal bile alcohols in urine, normal to increased plasma cholesterol concentrations and increased concentrations of plasma cholestanol. Little is known about cholesterol precursors in CTX, however. We studied cholesterol and phytosterol profiles in two siblings with CTX during follow-up. While cholesterol concentrations were low in both patients, plasma cholestanol was 6-fold higher compared to control values. In addition, both siblings had a more than 100-fold increase in 7-dehydrocholesterol (7DHC) and 8-dehydrocholesterol (8DHC). Lathosterol, lanosterol and sitosterol were increased in both patients while concentrations of desmosterol and campesterol were normal. In addition, plasma lathosterol/cholesterol ratios were significantly elevated. After treatment with chenodeoxycholate, both patients showed a marked decrease in cholestanol, 7DHC, 8DHC, lathosterol, lanosterol and sitosterol. In addition, the lathosterol/cholesterol ratio normalized, indicating that overall cholesterol synthesis was sufficiently suppressed. This study shows that elevated cholesterol precursors, other than cholestanol, can be a hallmark for CTX.

The distribution of heavy metals during fluidized bed combustion of sludge (FBSC).

During combustion of wastewater treatment sludge, the inorganic constituents are converted into ash which contains the major fraction of the heavy metals present. The behaviour of heavy metals in combustion processes has been studied extensively for mostly coal combustion and waste incineration. For biomass and sludge, literature data are scarce and mostly limited to laboratory experiments. The present paper assesses the partitioning of eight heavy metals (Hg, As, Cd, Cu, Pb, Cr, Ni and Zn) in the different residues from a large-scale fluidized bed sludge combustor of 4.4 m i.d. The origin of the sludge is mostly from treating urban wastewaters (>90%), although some mixed sludge (urban+industrial, <10%) is also burnt. The different residues (bottom ash, fly ash, filter cake, scrubber effluent and stack emissions) were sampled and analysed during 33 weeks, spread over a period of 1 year. The mass balance of relevant heavy metals closes for 96.5%, inaccuracies being related to the unsteadiness of the process, the accuracy of the mass flow data monitored at the plant, and on collecting representative samples. It is also shown that all heavy metals under scrutiny, except Hg, are concentrated in the fly ash as collected in the electrostatic precipitator.

Prehospital interventions: time wasted or time saved? An observational cohort study of management in initial trauma care.

OBJECTIVE: Preclinical actions in the primary assessment of victims of blunt trauma may prolong the time to definitive clinical care. The aim of this study was to examine the duration of performed interventions and to study the effect of on-scene time (OST) and interventions performed before admission to hospital on hospital resuscitation time. METHODS: 147 consecutive patients with high-energy blunt trauma aged >/=15 years were studied prospectively. Prehospital time intervals and interventions were documented and compared with hospital data collected from continuous digital video registration. Analyses were performed with correction for injury severity and type of prehospital medical assistance (emergency medical services (EMS) versus physician-staffed helicopter emergency medical services (HEMS)). RESULTS: Primary survey and initial treatment were initiated and completed within 1 h of arrival of the EMS. 83% of this "golden hour" elapsed out of hospital and 81% (n = 224) of all interventions (n = 275) were carried out before admission to hospital. An increase in the number of prehospital interventions was associated with an increased OST (p<0.001). Subanalyses showed no such correlation in the HEMS group. The HEMS group had a longer mean OST than the EMS group (p<0.001) with relatively more prehospital interventions (p<0.001) and a shorter mean in-hospital primary survey time with fewer in-hospital interventions. Overall, OST and the number of prehospital interventions were not related to in-hospital primary survey time and interventions. CONCLUSION: For most trauma patients the initial life- and limb-saving care is achieved within the "golden hour". Prehospital treatment occupies most of the golden hour. More prehospital interventions were performed with HEMS than with EMS only, but the higher number of interventions did not result in a longer OST with HEMS. Although the numbers of subsequent in-hospital interventions may be lower, no reduction in time in hospital may be expected from the interventions performed before hospital admission.

[Chemotherapy during clozapine treatment. Increased risk of agranulocytosis?]. / Chemotherapie bij gebruik van clozapine; een verhoogde kans op agranulocytose?

In a 37-year-old female, a combined treatment consisting of chemotherapy and radiation was considered for cervical cancer. However, she was using clozapine for the treatment of schizophrenia. As both clozapine and chemotherapy can induce decrease of white blood cell counts, we had to decide if clozapine and chemotherapy could be safely co-prescribed. Hypotheses concerning the mechanisms underlying clozapine-induced decrease of white blood cell counts and case reports on combining chemotherapy and clozapine are discussed. After cessation of clozapine the psychosis recurred despite treatment with risperidone. The decision was made to administer radiotherapy only and to reinstate the treatment with clozapine. The radiotherapy treatment went according to plan and the psychosis receded.

Cluster Correspondence Analysis.

A method is proposed that combines dimension reduction and cluster analysis for categorical data by simultaneously assigning individuals to clusters and optimal scaling values to categories in such a way that a single between variance maximization objective is achieved. In a unified framework, a brief review of alternative methods is provided and we show that the proposed method is equivalent to GROUPALS applied to categorical data. Performance of the methods is appraised by means of a simulation study. The results of the joint dimension reduction and clustering methods are compared with the so-called tandem approach, a sequential analysis of dimension reduction followed by cluster analysis. The tandem approach is conjectured to perform worse when variables are added that are unrelated to the cluster structure. Our simulation study confirms this conjecture. Moreover, the results of the simulation study indicate that the proposed method also consistently outperforms alternative joint dimension reduction and clustering methods.

Saline is as effective as nitrogen scavengers for treatment of hyperammonemia.

Urea cycle enzyme deficiency (UCED) patients with hyperammonemia are treated with sodium benzoate (SB) and sodium phenylacetate (SPA) to induce alternative pathways of nitrogen excretion. The suggested guidelines supporting their use in the management of hyperammonemia are primarily based on non-analytic studies such as case reports and case series. Canine congenital portosystemic shunting (CPSS) is a naturally occurring model for hyperammonemia. Here, we performed cross-over, randomized, placebo-controlled studies in healthy dogs to assess safety and pharmacokinetics of SB and SPA (phase I). As follow-up safety and efficacy of SB was evaluated in CPSS-dogs with hyperammonemia (phase II). Pharmacokinetics of SB and SPA were comparable to those reported in humans. Treatment with SB and SPA was safe and both nitrogen scavengers were converted into their respective metabolites hippuric acid and phenylacetylglutamine or phenylacetylglycine, with a preference for phenylacetylglycine. In CPSS-dogs, treatment with SB resulted in the same effect on plasma ammonia as the control treatment (i.e. saline infusion) suggesting that the decrease is a result of volume expansion and/or forced diuresis rather than increased production of nitrogenous waste. Consequentially, treatment of hyperammonemia justifies additional/placebo-controlled trials in human medicine.

Particle-in-cell Monte Carlo simulations of an extreme ultraviolet radiation driven plasma.

A self-consistent kinetic particle-in-cell model has been developed to describe a radiation driven plasma. Collisions between charged species and the neutral background are represented statistically by Monte Carlo collisions. The weakly ionized plasma is formed when extreme ultraviolet radiation coming from a pulsed discharge photoionizes a low pressure argon gas. The presence of a plasma close to optical components is potentially dangerous in case the ions that are accelerated in the plasma sheath gain enough energy to sputter the optics. The simulations predict the plasma parameters and notably the energy at which ions impact on the plasma boundaries. Finally, sputter rates are estimated on the basis of two sputtering models.

The complex blood supply to the equine testis as a cause of failure in laparoscopic castration.

REASONS FOR PERFORMING STUDY: Intra-abdominal ligation/ transection of the spermatic cord may result in necrosis of the testis; castration of abdominal cryptorchids via laparoscopy has therefore become common. Notwithstanding some adaptations of the technique, a small percentage of operations fail, prompting research into the anatomical background and clinical relevance of the procedure. HYPOTHESIS: That an alternate blood supply may prevent complete necrosis of the testis after spermatic cord transection. OBJECTIVE: To establish the prevalence of the problem in normal and cryptorchid stallions. METHODS: In a preliminary study, the spermatic cords of 8 normal stallions were ligated and transected at different sites and in various manners. Five weeks later the testes were removed and the vitality of both the testes and epididymes was evaluated. In a prospective clinical trial, intra-abdominal spermatic cord transection was performed in 241 cryptorchid and normal stallions. In cases of surgical failure, the testes were removed and histology performed. RESULTS: Examination of the specimen removed from the 8 animals of the preliminary study revealed that all epididymes were completely or largely spared. All except one testis were completely necrotic. In the patients that underwent surgery all abdominally retained testes (n = 123) were necrotic, while 5 out of 88 inguinally retained and 8 out of 236 normally descended testes had partially survived. The pattern of survival differed between inguinally retained and normally descended testes. The epididymes of these 13 horses were (largely) vital. The (partial) survival of the epididymes and inguinally retained testes was ascribed to an alternate blood supply via anastomosing vessels derived from the cremasteric artery. A tributary from the external pudendal artery was considered responsible for the partial survival of normally descended testes. CONCLUSIONS AND POTENTIAL RELEVANCE: After intra-abdominal transection of the entire spermatic cord, 5.6% of inguinally retained and 3.4% of normally descended testes failed to become completely necrotic, as a result of an alternate blood supply via the cremasteric and/or external pudendal artery. Therefore, laparoscopic castration without orchidectomy cannot be recommended as a trustworthy method for castration of inguinal cryptorchids and normal stallions.

[Unsuccessful laparoscopic castration in a cryptorchid Frisian stallion]. / Mislukte laparoscopische castratie van een Friese klophengst.

A 2-year-old unilateral inguinal cryptorchid Frisian stallion was castrated by bilateral laparoscopic intra-abdominal spermatic cord transection. Because blood testosterone levels were still high on postoperative day 7, both testicles were removed using an open approach. Pathology revealed the presence of vital Leydig-cells in both testes. Failure of the laparoscopic castration in this horse was attributed to an alternate blood supply to the testes by vessels derived from the cremaster and/or external pudendal artery.

Pilot Experience with an External Quality Assurance Scheme for Acylcarnitines in Plasma/Serum.

The analysis of acylcarnitines (AC) in plasma/serum is established as a useful test for the biochemical diagnosis and the monitoring of treatment of organic acidurias and fatty acid oxidation defects. External quality assurance (EQA) for qualitative and quantitative AC is offered by ERNDIM and CDC in dried blood spots but not in plasma/serum samples. A pilot interlaboratory comparison between 14 European laboratories was performed over 3 years using serum/plasma samples from patients with an established diagnosis of an organic aciduria or fatty acid oxidation defect. Twenty-three different samples with a short clinical description were circulated. Participants were asked to specify the method used to analyze diagnostic AC, to give quantitative data for diagnostic AC with the corresponding reference values, possible diagnosis, and advice for further investigations.Although the reference and pathological concentrations of AC varied among laboratories, elevated marker AC for propionic acidemia, isovaleric acidemia, medium-chain acyl-CoA dehydrogenase, very long-chain acyl-CoA dehydrogenase, and multiple acyl-CoA dehydrogenase deficiencies were correctly identified by all participants allowing the diagnosis of these diseases. Conversely, the increased concentrations of dicarboxylic AC were not always identified, and therefore the correct diagnosis was not reach by some participants, as exemplified in cases of malonic aciduria and 3-hydroxy-3-methylglutaryl-CoA lyase deficiency. Misinterpretation occurred in those laboratories that used multiple-reaction monitoring acquisition mode, did not derivatize, or did not separate isomers. However, some of these laboratories suggested further analyses to clarify the diagnosis.This pilot experience highlights the importance of an EQA scheme for AC in plasma.

Altered Energetics of Exercise Explain Risk of Rhabdomyolysis in Very Long-Chain Acyl-CoA Dehydrogenase Deficiency.

Rhabdomyolysis is common in very long-chain acyl-CoA dehydrogenase deficiency (VLCADD) and other metabolic myopathies, but its pathogenic basis is poorly understood. Here, we show that prolonged bicycling exercise against a standardized moderate workload in VLCADD patients is associated with threefold bigger changes in phosphocreatine (PCr) and inorganic phosphate (Pi) concentrations in quadriceps muscle and twofold lower changes in plasma acetyl-carnitine levels than in healthy subjects. This result is consistent with the hypothesis that muscle ATP homeostasis during exercise is compromised in VLCADD. However, the measured rates of PCr and Pi recovery post-exercise showed that the mitochondrial capacity for ATP synthesis in VLCADD muscle was normal. Mathematical modeling of oxidative ATP metabolism in muscle composed of three different fiber types indicated that the observed altered energy balance during submaximal exercise in VLCADD patients may be explained by a slow-to-fast shift in quadriceps fiber-type composition corresponding to 30% of the slow-twitch fiber-type pool in healthy quadriceps muscle. This study demonstrates for the first time that quadriceps energy balance during exercise in VLCADD patients is altered but not because of failing mitochondrial function. Our findings provide new clues to understanding the risk of rhabdomyolysis following exercise in human VLCADD.

[Eosinophilic keratitis in a horse]. / Keratitis eosinofilica bij een paard.

In this case report a 20-year-old horse with unilateral eosinophilic keratitis is presented. The occurrence, clinical symptoms, histological findings and treatment of this ocular disorder are described and discussed against the background of relevant literature. Local administration of 0.1% dexamethason was successful.