Decreased Urinary Sodium-to-urinary Creatinine Ratio Identifies Sodium Depletion in Pediatric Acute Gastroenteritis.

UNLABELLED: In acute gastroenteritis (AG) fecal losses may cause depletion of sodium (NaD) which may not be recognized because of normal plasma Na (pNa) concentrations. We studied the incidence of this state of normonatremic sodium depletion (NNaD) and the suitability of the urinary Na/urinary creatinine ratio (uNa/uCr) for diagnosing NNaD. PATIENTS: 16 AG- and 16 healthy control children aged 0.8-15.0 years. METHODS: Prospective cross sectional pilot study. Measurements of Na, K and creatinine in plasma (p) and urine (u). Calculation of uNa/uCr Ratio, fractional excretion of Na (FENa) and uNa/uK ratio as the hitherto best known parameters of prerenal Na depletion, respectively. RESULTS: pNa concentrations were normal in 15/16 AG patients (93.8%) with only one subnormal value of 133 mmol/L, and a mean value of 137.9±2.3 mmol/L not different from the normal control group (139.4±2.2 mmol/L). Also, mean uNa concentrations and uNa/uK ratios did not differ between both groups. However, uNa/uCr ratios were below normal in 13/16 AG children (81.3%) but normal in all healthy controls with a significantly lower mean value in the AG group (12.6±8.8 vs. 31.2±8.3 mmol/mmol; p<0.0001). Similarly, 14/16 AG patients (87.5%) had a decreased FENa<0.5% with a mean FENa value significantly lower than in controls (0.36±0.28% vs. 0.95±0.26%, p<0.0001). The good agreement between FENa and uNa/uCr results was also reflected by a high correlation coefficient of r=0.9333. CONCLUSIONS: The majority of AG patients was found to have NNaD as determined by uNa/uCr and FENa. Calculation of uNa/uCr may be useful for diagnosing NNaD in AG.

Severe anemia in 3 toddlers with gastric lactobezoar.

BACKGROUND: Anemia in toddlers may result from many disorders including excessive feeding with cow's milk. Another sequel of age-inadequate cow's milk nutrition may be gastric lactobezoar (GLB), a dense lump of coagulated milk and mucus in the stomach. PATIENTS: 3 toddlers presented with a history of excessive intake of full cream cow's milk, abdominal distension, vomiting, dehydration, fatigue, marked pallor and tachycardia. DIAGNOSTIC WORKUP: Diagnostic imaging revea-led large GLBs as the likely origin of the abdominal symptoms. Laboratory evaluation showed severe anemia with depleted iron stores and signs of protein catabolism. Non-cow's milk-induced causes of anemia including defects of erythropoiesis, hemoglobin structure, RBC-enzymes and blood coagulation, hemolysis, immune disorders, infection, inflammation, extraintestinal hemorrhage, nephropathy were - according to the available data - unlikely to cause the anemia in our patients. Thus their anemia is thought to be due to age-inadequate cow's milk nutrition leading to 1) low intake, decreased absorption/bioavailability and increased intestinal loss of iron, and 2) GLB which induced blood loss following mechanical irritation of the gastric mucosa and vomiting causing high gastric pH and decrease in duodenal iron absorption. CONCLUSION: The anemia in our patients is due to both exaggerated feeding with cow's milk and adverse effects of GLBs. This hypothesis is supported by the finding that, after erythrocyte transfusion, iron substitution, age-adapted nutrition and GLB-dissolution, the anemia did not recur. We propose to include GLB in the differential diagnosis of anemia in cow's milk fed small children.

Preoperative stress and anxiety in day-care patients and inpatients undergoing fast-track surgery.

BACKGROUND: To investigate preoperative levels of stress and anxiety in day-care patients and inpatients undergoing surgical interventions. METHODS: Before induction of anaesthesia, the degree of stress and anxiety was assessed in 135 patients using stress and anxiety questionnaires, bio-feedback, physiological measures, and serum levels for stress variables. Questionnaire responses and physiological measures such as arterial pressure, heart rate, skin conductance, cortisol, and catecholamine levels were compared for day-care patients and inpatients. RESULTS: Significant preoperative anxiety was reported by 34 (45.3%) inpatients and 23 (38.3%) day-care patients. Personal responses in stress and anxiety questionnaires and mean values of arterial pressure and heart rate did not differ significantly in day-care patients when compared with inpatients. Correlation between deviations in plasma cortisol concentrations from normal diurnal distribution and anxiety scores and stress scores was also similar, and the relative increase in preoperative stress variables and measures observed in day-care patients and inpatients was also comparable. Bio-feedback measurements revealed significantly higher preoperative skin conductance (P<0.001) in day-care patients than in inpatients, indicating increased vegetative stress responses. CONCLUSIONS: Preoperative anxiety and stress are common in surgical patients. Questionnaires and bio-feedback measurements may help to assess the degree of patients' burdens. Surgeons should be aware of the personal anxiety of patients and consider patient preferences when deciding who should undergo fast-track surgery in day-care.

Amino acids in CSF and plasma in hyperammonaemic coma due to arginase1 deficiency.

UNLABELLED: We report the CSF and plasma amino acid concentrations and their ratios in a male patient with arginase1 deficiency with an unusual early presentation at 34 days of age. He developed hyperammonaemic coma (ammonia >400 µmol/L; normal <90 µmol/L) on postnatal day 35. CSF and plasma concentrations were assayed by ion-exchange chromatography on day 36. Arginine was increased both in plasma (971 µmol/L; controls (mean ± 2SD) 50 ± 42) and in CSF (157 µmol/L; controls 19 ± 8.6), resulting in a normal CSF/plasma ratio of 0.16 (controls 0.41 ± 0.26). Interestingly, glutamine was disproportionately high in CSF (3114 µmol/L; controls 470 ± 236) but normal in plasma (420 µmol/L; controls 627 ± 246); the ratio exceeded unity (7.4; controls 0.76 ± 0.31). The CSF/plasma ratios of most neutral amino acids were elevated but not those of the imino- and of the dibasic amino acids lysine and ornithine. The mechanism leading to the increase of most neutral amino acids in brain is not known. CONCLUSION: A normal glutamine in plasma does not exclude an increased concentration in CSF; it could be useful to ascertain by MRS that a high CSF glutamine concentration truly reflects a high concentration in brain tissue for better understanding its pathogenesis.

The relation of cerebrospinal fluid and plasma glycine levels in propionic acidaemia, a 'ketotic hyperglycinaemia'.

The characteristic elevation of plasma glycine concentrations observed in propionic acidaemia (PA) and other 'ketotic hyperglycinaemias' has been attributed to secondary inhibition of the hepatic glycine cleavage system (GCS) by accumulating CoA derivatives of branched-chain amino acid metabolites. In nonketotic hyperglycinaemia (NKH), cerebrospinal fluid (CSF) and plasma glycine levels and their ratio are increased due to primary deficiency of central nervous system (CNS) as well as hepatic GCS. Whether the GCS in the CNS is also inhibited in PA is unclear, as there are scant data available on CSF glycine levels in this disorder. We studied the relation of CSF and plasma glycine levels in 6 paired samples from 4 PA patients, including one PA patient with bacterial meningitis who underwent ventriculoperitoneal shunting and multiple CSF analyses (n = 26). In contrast to the CSF glycine levels which were generally elevated in all four PA patients, the CSF/plasma glycine concentration ratios in paired samples were normal (0.016-0.029), with the exception of a single sample (0.132) with extremely high CSF protein concentration (2010 mg/L) during the course of meningitis indicating a disturbed blood-brain barrier. This finding of normal CSF/plasma glycine ratio in PA suggests that the observed elevations of CSF glycine levels are a reflection of the concurrent hyperglycinaemia resulting from secondary inhibition of hepatic GCS, but that brain GCS is not affected, in contrast to the situation in NKH. The neurological sequelae in PA are therefore unlikely to be related to disturbed glycine metabolism.

High frequency of missense mutations in glycogen storage disease type VI.

Deficiency of liver glycogen phosphorylase in glycogen storage disease (GSD) type VI results in a reduced ability to mobilize glucose from glycogen. Six mutations of the PYGL gene, which encodes the liver isoform of the enzyme, have been identified in the literature. We have characterized eight patients from seven families with GSD type VI and identified 11 novel PYGL gene defects. The majority of the mutations were missense, resulting in the substitution of highly conserved residues. These could be grouped into those that were predicted to affect substrate binding (p.V456M, p.E673K, p.S675L, p.S675T), pyridoxal phosphate binding (p.R491C, p.K681T), or activation of glycogen phosphorylase (p.Q13P) or that had an unknown effect (p.N632I and p.D634H). Two mutations were predicted to result in null alleles, p.R399X and [c.1964_1969inv6;c.1969+1_+4delGTAC]. Only 7 of the 23 (30%) reported PYGL alleles carry nonsense, splice site or frameshift mutations compared to 68-80% of affected alleles of the highly homologous muscle glycogen phosphorylase gene, PYGM, that underlie McArdle disease. There was heterogeneity in the clinical symptoms observed in affected individuals. These varied from hepatomegaly and subclinical hypoglycaemia, to severe hepatomegaly with recurrent severe hypoglycaemia and postprandial lactic acidosis. We conclude that deficiency of liver glycogen phosphorylase is predominantly the result of missense mutations affecting enzyme activity. There are no common mutations and the severity of clinical symptoms varies significantly.

Copper-induced hepatotoxicosis with hepatic stellate cell activation and severe fibrosis in North Ronaldsay lambs: a model for non-Wilsonian hepatic copper toxicosis of infants.

Copper-sensitive North Ronaldsay sheep represent a possible model for certain hepatic-overload syndromes of infancy and childhood that are clinically, pathologically and genetically distinct from Wilson's disease. The purpose of this study was to simulate in artificially reared lambs the syndrome produced by copper exposure in susceptible human infants. Twenty four North Ronaldsay lambs were assigned to three groups of eight animals, namely, an unsupplemented control group and two trial groups given milk replacer to which copper (CuSO4) had been added at the rate of 5 mg/litre and 10 mg/litre. Four lambs from each group were killed at 40 or 69 days. Livers were fixed in 10% formalin and analysed for copper by mass spectrometry. Paraffin wax-embedded sections were stained with rhodanine for copper and labelled immunohistochemically for alpha smooth muscle actin (ASMA). At 40 days the maximum amounts of copper in the livers of both copper-supplemented groups was 1466-1605 microg/g dry weight (control group 172-201 microg/g Cu dry weight). Histochemically, copper was demonstrated within hepatocytes, together with marked apoptosis. At 69 days there was a florid pericellular fibrosis complemented by strong ASMA immunolabelling, confirming phenotypic modulation of hepatic stellate cells. Such primary copper-induced fibrogenesis confirms the unique status of this animal model in respect of childhood copper toxicosis.

Lack of seasonal variation in flow-mediated dilatation of the brachial artery in women with primary Raynaud's phenomenon and healthy controls.

BACKGROUND: The aim of this study was to examine endothelium function and seasonal variations of endothelium function in women with primary Raynaud's phenomenon (RP) and healthy controls. PATIENTS AND METHODS: After a fast of at least 8 hours we studied 21 patients with primary RP (mean age 31.1 years, mean duration of RP 9.1 years) and 22 controls (mean age 27.8 years) by use of high resolution brachial artery sonography in winter (December/January 2000) and summer (July/August 2001). To exclude circadian variations all examinations were performed in the late afternoon only. All subjects were non-smokers. Confounding factors like serum glucose, HbAlc, and lipid concentrations were analyzed immediately before the investigations. Nicotine contamination was randomly analyzed in hair samples in 8 subjects of each study group. Flow mediated dilatation (FMD%) and nitroglycerin induced dilatation (NID%) were calculated by putting the basal vessel diameter as 100%. RESULTS: Basal, flow-mediated, and nitroglycerin-induced absolute diameters of the brachial artery did not differ significantly between the study groups (p = 0.85). The test conditions (basal, postocclusive, nitroglycerin-induced) always let to the same vessel response in winter and summer (p = 0.61) and there was no significant influence between these test conditions and the study groups (p = 0.07). Compared to patients FMD% was slightly reduced in controls in summer (p = 0.09). Analysis of variance excluded a significant relation between study group and season (p = 0.43). For NID% too, no statistically significant differences were found. CONCLUSIONS: We were not able to show impaired or seasonally variant flow-mediated or nitroglycerin-induced dilatation of the brachial artery in patients with primary RP. Our results argue against the presence of a more generalized endothelium dysfunction detectable with high resolution ultrasound of the brachial artery in patients with primary RP.

Colonic mucosal vasoactive intestinal peptide receptors in malformations of the enteric nervous system are reduced compared with morphologically normal innervated colon.

Vasoactive intestinal peptide (VIP) is the most important peptidergic transmitter in intestinal relaxation. VIPergic nerves are reduced in aganglionosis (AG). Corresponding findings in intestinal neuronal dysplasia (IND) are sparse. It is unknown whether superficial mucosal VIP receptors are reduced in AG, IND, or hypoganglionosis (HYP) compared to concentrations in morphologically normal innervated colon (MNIC). Cryostat sections from 38 colonic biopsies (23 patients with AG, IND, or HYP, 15 with MNIC) were incubated with radioactive iodinated (125)I-VIP. Receptors were analyzed by autoradiography. Radioactive-marked receptors trigger the precipitation of metallic silver as silver grains within a photographic emulsion. Grains were quantified by image analysis, calculating the percent covered cell surface. Statistical analysis was done by Mann-Whitney and Kruskal-Wallis tests (significance #E5/E5#<0.05). VIP receptors covered 4.31% of the cell surface in MNIC. The values were significantly reduced in AG (2.72%; #E5/E5#=0.012) and IND (2.93%; #E5/E5#=0.008). The two HYP biopsies showed the lowest values (1.83%). Aganglionic colon could be distinguished from healthy proximal segments and IND from habitual constipation. In AG and IND, even the superficial mucosal VIPergic innervation seems to be impaired. The reduction of mucosal VIP receptors in developmental faults of the enteric nervous system may thus be an indicator of a sensomotor disturbance.

Copper-associated liver disease in North Ronaldsay sheep: a possible animal model for non-Wilsonian hepatic copper toxicosis of infancy and childhood.

Indian childhood cirrhosis (ICC), endemic Tyrolean infantile cirrhosis (ETIC) and idiopathic copper toxicosis (ICT), are clinically and pathologically indistinguishable liver disorders of infants and young children linked with exogenous copper and with increasing evidence for a genetic predisposition. North Ronaldsay sheep are a primitive breed which have adapted to a copper impoverished environment (<5 ppm) and display an abnormal sensitivity to copper poisoning when transferred to a copper replete (11 ppm) habitat. The aetiological parallels prompted a study of copper-associated liver disease in North Ronaldsay sheep (RCT) to see if the pathology could contribute to the understanding of the childhood disorder. A retrospective study was performed in which the livers of 22 mainland-bred North Ronaldsay sheep were compared with three island-bred sheep and categorized for liver copper content and pathomorphology. It was found that all the mainland sheep had accumulated liver copper (>300 microg/g), in contrast to the island sheep, although 10 sheep with increased liver copper (mean 600 SD 270 microg/g) showed no evidence of liver damage. A further 10 sheep with liver copper (mean 1276 SD 508 microg/g) exhibited periportal to panlobular histochemical copper retention, a periportal and/or panlobular pericellular fibrosis, a mixed inflammatory infiltrate and cholangioplasia. Steatosis was absent and regeneration was in abeyance. Finally, two sheep (liver copper >2000 microg/g) had a more active hepatitis with a florid pericellular, panlobular fibrosis and cirrhosis. Electron microscopy identified large numbers of collagen-producing hepatic stellate (Ito) cells in periportal regions. The pathological findings were sufficiently reminiscent of ICC, ETIC and ICT to warrant further exploration of RCT as a putative animal model. The North Ronaldsay sheep liver may be a useful tool for the investigation of copper-induced fibrogenesis.

Congenital sodium diarrhea is an autosomal recessive disorder of sodium/proton exchange but unrelated to known candidate genes.

BACKGROUND & AIMS: Congenital sodium diarrhea (CSD) is caused by defective sodium/proton exchange with only 6 sporadic cases reported. The genetics of the disease have not been established. We studied 5 infants with secretory diarrhea, identified in a circumscribed rural area in Austria, to define the mode of transmission and the involvement of candidate genes known to encode for sodium/proton exchangers (NHEs). METHODS: We collected clinical and laboratory data from 5 affected patients, analyzed the pedigrees of their families, and performed homozygosity mapping and multipoint linkage analysis studies in 4 candidate regions known to contain NHE genes. RESULTS: The diagnosis of CSD in 4 of 5 patients was based on daily fecal sodium excretion between 98 and 190 mmol/L, hyponatremia, metabolic acidosis, and low-to-normal urinary sodium concentrations. Pedigree analysis of the affected 2 CSD families revealed parental consanguinity and a common single ancestor 5 generations ago. Homozygosity mapping and/or multipoint linkage analysis excluded the NHE1 locus on chromosome 1, NHE2 locus on chromosome 2, NHE3 locus on chromosome 5, and NHE5 locus on chromosome 16 as potential candidate genes for CSD in this pedigree. Results on NHE4 were inconclusive because the precise chromosomal location of this NHE gene in humans is currently unknown. CONCLUSIONS: Our data indicate that CSD is an autosomal recessive disorder but is not related to mutations in the NHE1, NHE2, NHE3, and NHE5 genes encoding for currently known sodium/proton exchangers.

Homozygous deletion of arginine-173 in the CYP11B2 gene in a girl with congenital hypoaldosteronism. Corticosterone methyloxidase deficiency type II.

The first child of consanguineous parents presented with failure to thrive and feeding problems at age 6 weeks. Important laboratory findings were low plasma sodium and elevated potassium and renin. Salt wasting was caused by an enzymatic defect in the terminal aldosterone biosynthesis. The biochemical diagnosis of corticosterone methyloxidase (CMO) deficiency type II was established on the basis of plasma multisteroid analysis, showing a pathologic increase of 18-OH-corticosterone/aldosterone ratio. Sequence analysis of the CYP11B2 gene which encodes aldosterone synthase (P450c11Aldo), the enzyme required for the terminal steps in aldosterone biosynthesis, revealed a hitherto undescribed homozygous deletion of codon 173. CYP11B2 is polymorphic at this position, encoding arginine or lysine. Both parents were heterozygous carriers of the mutation. Amino acid residue 173 in P450c11Aldo is positioned in alpha-helix D. We presume that the secondary structure of the enzyme is changed by the single amino acid deletion. This report describes a novel mutation in the CYP11B2 gene, the third known mutation associated with CMO deficiency type II.

Cellular localization of insulin-like growth factor II mRNA in the human fetus and the placenta: detection with a digoxigenin-labeled cRNA probe and immunocytochemistry.

IGF-II plays a major role in the regulation of human fetal growth and development. However, more extensive information on the cellular sites of IGF-II synthesis in the fetus would provide more insight into its role in fetal organogenesis. Thus we have determined the sites of IGF-II synthesis in 18-26-wk gestation human fetal tissues using in situ hybridization with a digoxigenin-labeled cRNA probe to localize IGF-II mRNA in fetal liver, kidney, adrenal gland, cerebral cortex, costal cartilage, skeletal muscle, and lung, and in placental tissue. In human fetal tissues it has to date been impossible to clearly assign IGF-II mRNA to epithelial cells of entodermal origin. Besides their already known localization in cell matrix and a variety of mesodermal cell types, strong IGF-II mRNA-positive signals were detected in epithelial cells in the liver (hepatocytes), bronchial and bronchiolar epithelium, undifferentiated renal tubular epithelium, mature glomerular epithelium, pelvic urothelium, and adrenal epithelial cells of the zona persistens. To identify the cellular location of immunoreactive IGF-II, we also performed immunocytochemical studies in tissues of the same fetuses. Every tissue studied except the cerebral cortex contained immunoreactive cells; however, immunostaining was generally weaker than in situ hybridization signals. Our data show that the distribution of IGF-II in human fetal tissue is much more widespread than hitherto thought. A digoxigenin-labeled detection system for IGF-II is more capable of detecting the cellular expression pattern of IGF-II than radioactive probes and is suitable for analysis of routinely prepared paraffin-embedded material.

Expression of IGFBP-2, -3, and -4 mRNA during differentiation of Caco-2 colon epithelial cells.

Local production of insulin-like growth factor (IGF)-binding proteins (IGFBP) determines the availability of the IGF to the cell and thus regulates IGF action. To find out whether specific patterns of IGFBP gene expression and IGFBP secretion were related to cell growth vs. cell differentiation, expression of IGFBP during long-term culture (21 days, n = 5) of the colon carcinoma cell line Caco-2 was investigated at the mRNA and protein levels. Markers of cell proliferation (increase in DNA, RNA, and protein content) and of differentiation [alkaline phosphatase (AP) activity; creatine kinase (CK) activity] were measured in parallel during long-term culture. IGFBP-2 mRNA expression correlated significantly with markers of proliferation (P < 0.05), whereas IGFBP-3 mRNA expression or IGFBP-3 secretion correlated with markers of differentiation (AP: r = 0.83, P < 0.001; CK: r = 0.45, P < 0.01). Similarly, IGFBP-4 mRNA expression correlated significantly with markers of differentiation (AP: r = 0.34, P < 0.05; CK: r = 0.35, P < 0.05). We hypothesize that IGFBP-3 and -4 are related to differentiation of Caco-2 cells, whereas IGFBP-2 is related to proliferation in Caco-2 cells.