H. J. Bremer

Large neutral amino acids block phenylalanine transport into brain tissue in patients with phenylketonuria

Large neutral amino acids (LNAAs), including phenylalanine (Phe), compete for transport across the blood-brain barrier (BBB) via the L-type amino acid carrier. Accordingly, elevated plasma Phe impairs brain uptake of other LNAAs in patients with phenylketonuria (PKU). Direct effects of elevated brain Phe and depleted LNAAs are probably major causes for disturbed brain development and function in PKU. Competition for the carrier might conversely be put to use to lower Phe influx when the plasma concentrations of all other LNAAs are increased. This hypothesis was tested by measuring brain Phe in patients with PKU by quantitative 1H magnetic resonance spectroscopy during an oral Phe challenge with and without additional supplementation with all other LNAAs. Baseline plasma Phe was approximately 1,000 micromol/l and brain Phe was approximately 250 micromol/l in both series. Without LNAA supplementation, brain Phe increased to approximately 400 micromol/l after the oral Phe load. Electroencephalogram (EEG) spectral analysis revealed acutely disturbed brain activity. With concurrent LNAA supplementation, Phe influx was completely blocked and there was no slowing of EEG activity. These results are relevant for further characterization of the LNAA carrier and of the pathophysiology underlying brain dysfunction in PKU and for treatment of patients with PKU, as brain function might be improved by continued LNAA supplementation.

A metabolic disorder similar to Zellweger syndrome with hepatic acatalasia and absence of peroxisomes, altered content and redox state of cytochromes, and infantile cirrhosis with hemosiderosis.

A patient with a cerebro-hepato-renal syndrome was investigated. The visceral manifestations were those of the Zellweger syndrome (ZS); however, the child exhibited muscular hypertonia and survived into the 2nd year of life. Ultramicroscopically, hepatocytes were lacking peroxisomes, but, contrary to findings in one patient with ZS [2], contained smooth endoplasmic reticulum. No catalase was found by histochemistry or spectroscopy. Mitochondria showed normal succinate and glutamate respiration, and normal coupling of respiration to the phosphorylation potential. The cytochrome (cyt) content was diminished to one-third with an abnormally inversed redox patterns of the respiratory chain in the controlled state, cyt b being 5%, cyt c 23% reduced. The oxygen affinity of cyt a3 was normal. These findings exclude a defect in the nonheme iron protein region of the respiratory chain as described in ZS [2], but point to a functional abnormality of cyt b in our patient.

Alpha-ketoadipic aciduria, a new inborn error of lysine metabolism; biochemical studies☆

Investigation of a psychomotorically retarded girl showed excretion of abnormal amounts of alpha-ketoadipic acid, alpha-hydroxyadipic acid, alpha-aminoadipic acid, 1,2-butenedicarboxylic acid and elevation of plasma alpha-aminoadipic acid levels. The identity of these metabolities was established by various methods. The excretion of alpha-aminoadipic acid correlated to the lysine intake. Degradation studies with cultured fibroblasts indicate a defect in the oxidative decarboxylation of alpha-ketoadipic acid (see Clin. Chim. Acta, 58 (1975) 271.

Fatty acid composition of plasma lipids in Nigerian children with protein-energy malnutrition

The fatty acid (FA) composition of the main plasma lipids was analysed in eight well-nourished, generally healthy Nigerian children aged 14.1±7.2 months and in 17 malnourished children (8 marasmus, 9 kwashiorkor) aged 14.6±3.8 months within the first 2 days of admission at the Dept. of Child Health, University of Benin. In comparison to the control group, the malnourished children showed a marked decrease of polyunsaturated FA with low linoleic acid, mainly in sterolesters (STE), and severely reduced linoleic acid metabolites, including arachidonic acid, in all lipid fractions. ω-3-FA were not altered except for a reduction of docosapentaenoic and docosahexaenoic acids in phospholipids. Clearly increased values were found for saturated FA in STE and for the non-essential monoenoic FA in all lipid classes. This pattern indicates the presence of essential fatty acid deficiency in the malnourished children. There was no significant difference between marasmus and kwashiorkor. Eight malnourished children were followed up in the early phase of recovery during hospital treatment 14.0±3.1 days after obtaining the first sample. Linoleic acid had increased again in STE, but its metabolites were as low or even lower than before. An impaired activity of delta-6-desaturase, the rate limiting enzyme of linoleic acid metabolism, in suggested by elevated substrate-product-ratios of this enzyme in untreated children with protein energy malnutrition and in the early phase of recovery, which may be due to low insulin levels, protein and zinc deficiency. The trientetraen-ratio (20∶3ω9/20∶4ω6) thus is not a reliable indicator of essential FA status in protein-energy malnutrition.

Glutathione and associated antioxidant systems in protein energy malnutrition: Results of a study in nigeria

Marasmus and kwashiorkor are manifestations of protein energy malnutrition. The pathophysiology of these disorders is poorly understood. We studied a number of blood antioxidants [glucose-6-phosphate dehydrogenase (G6PDH), glutathione reductase (GR) and its cofactor flavin adenine dinucleotide (FAD), the tripeptide glutathione as the major nonprotein thiol], serum albumin, and retinol-binding protein in 12 children suffering from kwashiorkor with all classical symptoms, in 13 patients with clinically severe marasmus, in 19 marasmic but active children, and in 23 controls. Significant changes were observed for erythrocyte glutathione and correspondingly for nonprotein thiols in whole blood (0.72 +/- 0.29 mM thiols in controls, 0.50 +/- 0.22 mM in marasmus, 0.35 +/- 0.23 mM in severe marasmus, and 0.22 +/- 0.13 mM in kwashiorkor). These differences were paralleled by a decrease in serum albumin concentration so that the molar ratio of nonprotein thiols/albumin had an average value of approximately 1.5 in all groups. The erythrocyte glutathione-reducing system, represented by G6PDH and glutathione reductase, showed only slight differences among the four groups of children; the supposition that kwashiorkor occurs predominantly in children with aberrant G6PDH could not be substantiated. Unexpectedly, erythrocyte FAD, an index of riboflavin status, was normal in most malnourished patients. Discussed is the prospect of administering glutathione in kwashiorkor patients.

Influence of dietary (n-3)-polyunsaturated fatty acids on leukotriene B4 and prostaglandin E2 synthesis and course of experimental tuberculosis in guinea pigs

SummaryIn the present study eicosanoid synthesis was studied in macrophages of guinea pigs fed different amounts of (n-6)- and (n-3)-polyunsaturated fatty acids (PUFA). Three groups of weanling guinea pigs were fed by isocaloric diets differing only in their contents of PUFA: controls with 2.8 Cal% of linoleic acid (LA; 18:2(n-6)); (n-6)-rich fed animals with 15.4 Cal% of LA; and (n-3)-rich fed animals with 10.1 Cal% of LA, 1.4 Cal% of eicosapentaenoic acid (20:5(n-3)) and docosahexaenoic acid (22:6(n-3)). After 13 weeks half the number of animals from each group was infected i.m. by 180 colony forming units ofMycobacterium tuberculosis strain H37Rv. Seven weeks after infection the release of leukotriene (LT)B4 and prostaglandin (PG)E2 was quantified in calcium ionophore stimulated whole blood, peritoneal macrophage cultures and alveolar macrophages by immunoassays after high performance liquid chromatography. Synthesis of LTB4 and PGE2 was found to be reduced in (n-3)-rich fed guinea pigs (p<0.05), and equivalent between controls and (n-6)-rich fed animals. Controls and (n-6)-rich fed animals showed the same mycobacterial counts in the spleen whereas (n-3)-rich fed guinea pigs demonstrated an increased number of mycobacteria (p<0.05). Our results demonstrate that an increased dietary intake of (n-3)-PUFA suppress LTB4 and PGE2 synthesis. The increased number ofM. tuberculosis found in the spleens of (n-3)-rich fed animals could represent persistence of the experimental infection. It may be speculated that a functional relationship exists between the two findings.ZusammenfassungIn der vorliegenden Studie wurde die Eikosanoidsynthese in Makrophagen von Meerschweinchen untersucht, die mit unterschiedlichen Gehalten (n-6)- und (n-3)-mehrfach ungesättigten Fettsäuren (PUFA) gefüttert wurden. Drei Gruppen entwöhnter Meerschweinchen wurden mit isokalorischen Diäten gefüttert, die sich nur in ihrem Gehalt an PUFA unterschieden: Kontrollen mit 2,8 Cal% Linolsäure (LA. (18:2(n-6)); Tiere mit (n-6)-angereichertem Futter mit 15,4 Cal% LA und Tiere mit (n-3)-angereichertem Futter mit 10,1 Cal% LA, 1,4 Cal% Eikosapentaensäure (20:5(n-3)) und 0,9 Cal% Dokosahexaensäure (22:6(n-3)). Nach 13 Wochen wurde die Hälfte der Tiere aus jeder Gruppe mit 180 Kolonie-bildenden Einheiten des StammesMycobacterium tuberculosis H37Rv i.m. infiziert. Sieben Wochen nach Infektion wurde die Freisetzung von LTB4 und PGE2 in Kalzium-Ionophor stimuliertem Vollblut, Peritonealmakrophagen und Alveolarmakrophagen mittels Immunoassays nach Hochdruckflüssigkeitschromatographie quantifiziert. Die Synthese von LTB4 und PGE2 war reduziert bei den (n-3)-reich gefütterten Meerschweinchen (p<0,05) und äquivalent bei den Kontrollen und (n-6)-reich gefütterten Tieren. Kontrolltiere und (n-6)-reich gefütterte Meerschweinchen wiesen die gleiche Zahl von Mykobakterien in der Milz auf, während sich bei den (n-3)-gefütterten Tieren eine erhöhte Zahl von Mykobakterien zeigte (p<0,05). Unsere Ergebnisse zeigen, daß eine höhere diätetische Zufuhr von (n-3)-PUFA die Synthese von LTB4 und PGE2 unterdrückt. Die erhöhte Zahl vonM. tuberculosis in den Milzen von (n-3)-gefütterten Tieren spricht für eine Persistenz der experimentellen Tuberkulose unter diesen Bedingungen. Möglicherweise existiert zwischen beiden Befunden ein funktioneller Zusammenhang.

Serum-selenium concentrations in patients with maple-syrup-urine disease and phenylketonuria under dieto-therapy

Serum-selenium concentrations were measured in 2 dietetically treated patients with maple-syrup-urine disease, in 11 dietetically treated patients with phenylketonuria, in 37 healthy children of different ages and in 183 healthy adults. The estimations were performed by instrumental neutron activation analysis. The values of the dietetically treated patients were much lower than those of healthy children of the same age group. Within 8 to 12 weeks the serum-selenium concentrations decreased from normal values before therapy to very low values under dieto-therapy. During infancy the serum-selenium concentrations of healthy individuals show an increase to the adult range of values.

Akrodermatitis enteropathica — eine Zinkstoffwechselstörung mit Zinkmalabsorption

The intestinal resorption of zinc using 65ZnCl2 was estimated in 3 patients with acrodermatitis enteropathica, 2 healthy controls, and 3 heterozygotes. After oral application of 65Zn the whole body activity was measured by a whole body counter for 34 days. The 65Zn resorption of the patients amounted to 16, 42 and 30% of the applied dose, whereas the resorption values of the heterozygotes and the controls were in the range of 58 and 77%. The elimination of 65Zn from the body amounted to about 0.7% of the applied dose with no difference between controls and patients with acrodermatitis enteropathica. Before therapy the serum-zinc levels of patients were markedly decreased. After oral application of high doses of zinc aspartate (2×400 mg/day) all clinical symptoms disappeared within a week.The results point at a causal connection between zinc and the pathogenesis of acrodermatitis enteropathica. Ultrastructural alterations of the Paneth cells of the intestine are also shown in this disease [12] as have also been seen in Paneth cells of zinc deficient rats [Beitr. Path. 145, 336 (1972)].ZusammenfassungBei 3 Patienten mit Akrodermatitis enteropathica wurde mit Hilfe der Ganzkörpermessung nach oraler Applikation von 65Zn eine verminderte intestinale Zinkresorption gedunden. Dagegen war die Zinkelimination aus dem Körper normal. Die Zinkkonzentration im Serum war bei den Patienten stark erniedrigt. Alle klinischen Symptome verschwanden nach sehr hohen oralen Zinkdosen. Diese Befunde sprechen um so mehr für eine ursächliche Rolle des Zinks in der Pathogenese der Akrodermatitis enteropathica, als bei dieser Krankheit ultrastrukturelle Veränderungen in den Panethschen Zellen nachzuweisen sind [12], die auch beim Zinkmangel der Ratten gefunden wurden [Beitr. Path. 145, 336 (1972)].

Plasma glutathione peroxidase after selenium supplementation in patients with reduced selenium state

The plasma glutathione peroxidase (GSHPx) activity was measured in normal adults and children and in patients with reduced selenium state because of dietary treatment of metabolic diseases (phenylketonuria or maple-syrup-urine disease) before and after selenium supplementation. Besides GSHPx (measured with t-butyl hydroperoxide, cumene hydroperoxide and hydrogen peroxide as acceptor substrates) the activity of glutathione S-transferase was estimated in plasma. Plasma GSHPx activity in healthy children was significantly lower than in healthy adults. In 11 dietetically treated patients with phenylketonuria or maple-syrup-urine disease the plasma GSHPx was reduced to about 17% of the values of healthy children of the same age. No glutathione S-transferase activity could be found in plasma of children in normal or reduced Se state.During administration of yeast rich in Se (200μg Se/d) for 90 days 2 healthy adults showed no significant change of plasma GSHPx activity. During Se supplementation (75–100μg Se/d) for 120–163 days 5 dietetically treated patients with PKU or MSUD exhibited a significant increase of plasma GSHPx activity within 2 days. The values reached a plateau after 1 to 3 weeks of supplementation and remained at this level within the following 4 to 5 months. Therefore, the activity of plasma glutathione peroxidase can be used as an indicator of short-term changes of selenium intake in selenium deficient individuals.

Alpha-ketoadipic aciduria: Degradation studies with fibroblasts

Observation of one patient with alpha-ketoadipic aciduria initiated degradation studies with radiolabelled lysine metabolites in fibroblasts in order to localise the metabolic defect. Liberation of 14-CO-2 from alpha-D,L-(1-14-C) aminoadipate and alpha-(1-14-C) ketoadipate was considerably less in the patients fibroblasts than in the patients fibroblasts than in normal controls, whereas 14-CO-2 production from (1,5-14-C) glutarate was in the normal range. These results indicate a defect in the oxidative decarboxylation of alpha-ketoadipate as the probable cause of alpha-ketoadipic aciduria; Cultured amniotic fluid cells from pregnancies of the 15th and 16th week of gestation degrade alpha-(1-14-C) ketoadipate with a similar activity to fibroblast cultures from normal humans after birth.