Noboru Igarashi

Oxidative stress causes enhanced endothelial cell injury in human heme oxygenase-1 deficiency

The first known human case of heme oxygenase-1 (HO-1) deficiency is presented in this report. The patient is a six-year-old boy with severe growth retardation. He has been suffering from persistent hemolytic anemia characterized by marked erythrocyte fragmentation and intravascular hemolysis, with paradoxical increase of serum haptoglobin and low bilirubin. An abnormal coagulation/fibrinolysis system, associated with elevated thrombomodulin and von Willebrand factor, indicated the presence of severe, persistent endothelial damage. Electron microscopy of renal glomeruli revealed detachment of endothelium, with subendothelial deposition of an unidentified material. Iron deposition was noted in renal and hepatic tissue. Immunohistochemistry of hepatic tissue and immunoblotting of a cadmium-stimulated Epstein-Barr virus-transformed lymphoblastoid cell line (LCL) revealed complete absence of HO-1 production. An LCL derived from the patient was extremely sensitive to hemin-induced cell injury. Sequence analysis of the patients HO-1 gene revealed complete loss of exon-2 of the maternal allele and a two-nucleotide deletion within exon3 of the paternal allele. Growth retardation, anemia, iron deposition, and vulnerability to stressful injury are all characteristics observed in recently described HO-1 targeted mice. This study presents not only the first human case of HO-1 deficiency but may also provide clues to the key roles played by this important enzyme in vivo.

Molecular analysis of peroxisomal β-oxidation enzymes in infants with Zellweger syndrome and Zellweger-like syndrome: Further heterogeneity of the peroxisomal disorder

The biosynthesis of enzymes of peroxisomal beta-oxidation was investigated in an attempt to elucidate the mechanism of deficiencies of proteins of these enzymes in 3 infants with Zellweger syndrome and in a baby with Zellweger-like syndrome with clinical and biochemical findings consistent with Zellweger syndrome except that the peroxisomes were detected electronmicroscopically. Enzyme proteins of peroxisomal beta-oxidation, acyl-CoA oxidase, bifunctional protein and 3-ketoacyl-CoA thiolase were hardly detectable, in both syndromes. Total hepatic RNA extracted from the liver of one patient with each syndrome and three controls was translated in a rabbit reticulocyte lysate protein-synthesizing system in the presence of [35S]methionine. Translatable mRNAs for all of the peroxisomal beta-oxidation enzymes were detected in both patients at much the same levels seen in the controls. Pulse labelling and chase experiments of fibroblasts from the control revealed that the 72 kDa subunit of acyl-CoA oxidase was first synthesized, after which the 52 kDa and 21 kDa subunits were processed from the 72 kDa subunit. In the patient with Zellweger syndrome, little of the 52 kDa and 21 kDa subunits of acyl-CoA oxidase were synthesized. The mature form of peroxisomal 3-ketoacyl-CoA thiolase was also not processed from its precursor form, in this patient. We consider that Zellweger-like syndrome is a new variant form of a peroxisomal disorder in which biogenesis of peroxisomes is intact, while in contrast, the biogenesis of peroxisome is considered to be defective in those with Zellweger syndrome. Multiple defects of enzymes of beta-oxidation in Zellweger-like syndrome are assumed to be caused by a defect of transport or localization of these enzymes. Our molecular analyses indicate that the enzymes of peroxisomal beta-oxidation are synthesized in patients with Zellweger and Zellweger-like syndrome but that these enzymes are not processed normally and are degraded rapidly.

Zellweger-like syndrome with detectable hepatic peroxisomes: A variant form of peroxisomal disorder

A male infant with typical clinical and biochemical findings of Zellweger syndrome, but in whom hepatic peroxisomes were detected by electron microscopy, had profound hypotonia, hepatomegaly, typical facial appearance including large fontanelle and frontal bossing, convulsions, panaminoaciduria, and hyperammonemia. He died of liver failure at age 5 months. There were increased levels of very long chain fatty acids and trihydroxycoprostanic acid in serum, and increased excretion of dicarboxylic acids and tyrosine metabolites in the urine. Levels of peroxisomal enzymes, acyl coenzyme A oxidase, bifunctional protein, 3-ketoacyl coenzyme A thiolase, and dihydroxyacetone phosphate acyltransferase in the liver tissue from the patient were all deficient, findings consistent with Zellweger syndrome. However, immunocytochemical study and electron microscopic examination of the liver at autopsy revealed that hepatic peroxisomes were present at a level similar to that in a control subject. These observations suggest further heterogeneity in Zellweger syndrome and a different pathogenesis in this variant case.

Cytokine profiles of patients with enterohemorrhagic Escherichia coli O111-induced hemolytic-uremic syndrome

Proinflammatory cytokines are related to the pathogenesis of enterohemorrhagic Escherichia coli (EHEC) infection and hemolytic-uremic syndrome (HUS). We assessed the kinetics of the release of cytokines such as neopterin, interleukin (IL)-6, IL-8 and tumour necrosis factor (TNF)-α and the soluble forms of type I and II TNF receptors during EHEC O111-induced HUS (EHEC O111/HUS). Fourteen patients with EHEC O111/HUS were enrolled in this study. Serum concentrations of all cytokines other than TNF-α were significantly elevated in patients with severe HUS compared with those in patients with mild HUS. Although serum concentrations of TNF-α were not significantly higher in patients with severe HUS, most patients with acute encephalopathy showed elevated TNF-α levels. Serum concentrations of these cytokines rapidly and markedly increased, and massive hypercytokinaemia developed 1 day before the diagnosis of HUS in patients with severe HUS. Changes in the number of white blood cells and concentration of serum lactate dehydrogenase were significantly larger between the onset of hemorrhagic colitis and the time of the diagnosis of HUS in patients with severe HUS compared with those in patients with mild HUS. Proinflammatory cytokines play an important role in the pathogenesis of EHEC infection and development of severe complications, including HUS and encephalopathy. Monitoring the cytokine profile may be useful for assessing disease activity of EHEC O111 infections.

Acquired von Willebrand's syndrome with lupus-like serology.

We describe a 12-year-old boy with acquired von Willebrands syndrome, who also had various autoantibodies. He presented with recent hemorrhagic symptoms and a prolonged bleeding time. Hemostatic studies revealed severely reduced levels of factor VIII procoagulant activity (VIII:C), von Willebrands factor (vWF) antigen (vWF:Ag), and ristocetin cofactor activity (RCoF). An inhibitor that could be detected in the patients plasma moderately decreased the levels of vWF:Ag in normal plasma, but did not interfere with the measurement of VIII:C or RCoF. Following the infusion of cryoprecipitate, half-lives of VIII:C, vWF:Ag, and RCoF were markedly reduced. 1-Deamino-8-D-arginine vasopressin infusion induced normalization of the prolonged bleeding time and caused a marked increase in VIII:C, vWF:Ag, and RCoF. Prior to treatment, there was a uniform reduction of all the multimers of plasma vWF in sodium dodecyl sulfate agarose gel electrophoresis. Following prednisone therapy, clinical and hemostatic findings were improved, and the multimeric patterns of vWF were normalized. These findings suggest that the low levels of all three parameters of factor VIII and all the multimers of plasma vWF in the patient are caused by rapid elimination of factor VIII complex from the circulation.

Extensive serum biomarker analysis in patients with enterohemorrhagic Escherichia coli O111-induced hemolytic-uremic syndrome

Proinflammatory cytokines are related to the pathogenesis of enterohemorrhagic Escherichia coli infection and hemolytic-uremic syndrome (HUS). We employed an antibody array that simultaneously detects 174 serum cytokines. We identified five serum biomarkers, namely insulin growth factor-binding protein-2, angiopoietin-2, soluble interleukin-6 receptor, soluble tumor necrosis factor receptor type II, and matrix metalloprotease protein-3 whose levels increased with the development of HUS. Furthermore, the levels of these cytokines were significantly increased in severe HUS compared with mild HUS. These cytokines might play an important role in the pathogenesis of HUS and may also be used to predict the severity of HUS.

Serum tau protein as a marker of disease activity in enterohemorrhagic Escherichia coli O111-induced hemolytic uremic syndrome

Tau protein levels in cerebrospinal fluid (CSF) and serum are elevated in patients with various central nervous system diseases. We investigated whether serum tau protein levels are useful for predicting and assessing disease activity of acute encephalopathy (AE) in enterohemorrhagic Escherichia coli (EHEC) O111-induced hemolytic uremic syndrome (HUS; EHEC encephalopathy). Serum samples were obtained from 14 patients with EHEC O111/HUS, 20 patients with non-EHEC-related AE, and 20 age- and sex-matched healthy controls. CSF samples were obtained from 2 patients with EHEC encephalopathy and 20 patients with non-EHEC-related AE. Tau protein levels and levels of several proinflammatory cytokines were quantified by enzyme-linked immunosorbent assays. Results were compared with the clinical features of EHEC encephalopathy, including magnetic resonance image (MRI) findings. Serum tau levels in patients with EHEC encephalopathy were significantly elevated compared with those in patients with EHEC O111/HUS without encephalopathy, patients with non-EHEC-related AE, and healthy controls. The ratio of CSF tau levels to serum tau levels was >1.0 in all patients with non-EHEC-related AE but <1.0 in 2 patients with EHEC encephalopathy. Serum tau protein levels increased rapidly and markedly in patients with severe EHEC 0111/HUS and encephalopathy when HUS occurred, but were not elevated in mild patients, even in the HUS phase. Furthermore, changes in serum tau protein levels in patients with EHEC encephalopathy were consistent with abnormalities on brain MRI and were positively correlated with proinflammatory cytokine levels. Our results indicate that serum tau protein might be useful to predict and assess disease activity of EHEC encephalopathy.

Commotio Cordis Presenting as a Temporary Complete Atrioventricular Block in a 2-Year-Old Girl With Congenitally Corrected Transposition of the great Arteries

This report describes a 2-year-old girl with congenitally corrected transposition of the great arteries (ccTGA) who presented with transient complete atrioventricular (AV) block after a mild chest blow. Running around the house with her older sister, she fell to the floor. Her sister also fell and landed on her. The girl became cyanotic and pale and experienced a transient loss of consciousness. At arrival to the emergency department, she had regained consciousness, but she remained pale. An electrocardiogram (ECG) demonstrated complete AV block with a heart rate of 78 beats per minute (bpm). The ECG after admission showed a Wenckebach-type second-degree AV block. Day 2 after admission, a 12-lead ECG showed significant ST and T-wave abnormalities in the precordial leads, but the girl had no chest pain and a normal physical examination. Echocardiography demonstrated normal contractility of the systemic right ventricle. The first-degree AV block and the ST and T-wave abnormalities on the 12-lead ECG improved gradually without abnormal Q-waves. This is the first report of ccTGA in which a transient complete AV block naturally recovered after a presentation with commotio cordis.

Dopaminergic Instability in Children with Orthostatic Dysregulation

Effects of postural change on plasma catecholamine levels were examined in 53 children with orthostatic dysregulation (OD). Special interest was focused on plasma dopamine and dopamine‐β‐hydroxylase (DBH) activity, since one patient showed a prominent rise in plasma dopamine from 822 pmol/L to 126 nmol/L in postural change. She had a wide fluctuation of plasma dopamine from 209 pmol/L to 305 nmol/L during 12 hours of observation period, but plasma norepinephrine and epinephrine remained within the normal range as well as urinary excretion of catecholamines. Of 52 children with OD, 11 had a marked increase in plasma dopamine, whereas 11 showed a reduction of it by postural change. Plasma DBH activity was significantly decreased in the former (81±14 nmol/h/ml), while it was elevated in the latter (320±48 nmol/h/ml, P<0.001 vs the former), although DBH activity in individuals was not affected by postural change. These results indicate the involvement of dopaminergic instability as a cause of OD in childhood.

Correlation between pituitary growth hormone reserve and degree of growth failure in children with short stature

The correlation between a releasable pituitary growth hormone (GH) pool and degree of growth failure was examined in 30 children with GH deficiency (group I) and 19 children with normal short stature (group II). Based on the responsiveness of GH to GH-releasing hormone (GHRH), group I, with low GH responses (below 7 ng/ml) to both insulin and arginine, was classified into three subgroups; Ia (peak value less than 10 ng/ml, n=19), Ib (10–20 ng/ml, n=5) and Ic (above 20 ng/ml, n=6). Group II, with a GH response above 10 ng/ml to either insulin or arginine, was also divided into IIa (below 20 ng/ml, n=5) and IIb (above 20 ng/ml, n=14). Body length and growth velocity in Ia and Ib were significantly reduced vs Ic; bone age in Ia was retarded vs Ic; plasma somatomedin C (Sm-C) levels in Ia and Ib were decreased vs Ic, who had almost normal levels (0.90±0.55 U/ml). The incidence of other combined pituitary hormone deficiencies and previous perinatal distress was definitely high in Ia and Ib, but zero in Ic. In group II also, body length and growth velocity were significantly decreased in IIa vs IIb (P<0.01). These results indicate that [1] the pituitary reserve of GH estimated by GHRH is a good reflection of the degree of growth failure in GH-deficient children as well as in those of normal short stature, [2] hypothalamic GHRH deficiency tends to have a milder effect on growth retardation than pituitary GH deficiency, and [3] normal short children with a diminished GH reserve may be potential candidates for the GH treatment.