Progressive atrophy of the cerebrum in 2 Japanese sisters with microcephaly with simplified gyri and enlarged extraaxial space.

This is a case report that describes 2 sisters with microcephaly, simplified gyri, and enlarged extraaxial space. Clinical features of the cases include dysmorphic features, congenital microcephaly, failure of postnatal brain growth, neonatal onset of seizures, quadriplegia, and severe psychomotor delay. Neuroradiological imaging demonstrated hypoplasia of bilateral cerebral hemispheres with enlarged extraaxial spaces, simplified gyral patterns without a thickened cortex, hypoplastic corpus callosum, and enlarged lateral ventricles, with a reduction in gray and white matter volume during the prenatal and neonatal periods. Repeat MRI revealed progressive atrophy of the cerebral gray and white matter, with enlarged lateral ventricles, although the sizes of the bilateral basal ganglia, thalamus, and infratentorial structures were relatively preserved. These neuroradiological findings imply that this disease is caused by the gene involved in neuronal and glial proliferation in the ventricular zone and in tangential neuronal migration from the ganglionic eminence. The nature of the progressive degeneration of the hemispheric structures should be clarified.

The cytokine and chemokine profiles in rhabdomyolysis in a patient with Gaucher disease type II.

This report describes a patient with Gaucher disease type II who developed severe rhabdomyolysis. We treated him successfully and measured various cytokine and chemokine levels sequentially to elucidate the pathophysiology of rhabdomyolysis. The serum levels of interleukin-6, -8, -10, granulocyte colony-stimulating factor, interferon-gamma, and monocyte chemoattractant protein-1 were markedly elevated in the early phase of rhabdomyolysis. These findings indicate that cytokines and chemokines are related to the massive myolysis and regenerating process. A viral infection may have triggered rhabdomyolysis through exaggerated activation of macrophages in our patient. The profiles of cytokines and chemokines should be examined in further cases to increase our understanding of the pathophysiology of rhabdomyolysis.

Influence of the cyclooctadepsipeptides PF1022A and PF1022E as natural products on the design of semi-synthetic anthelmintics such as emodepside.

The 24-membered cyclooctadepsipeptide (CODP) PF1022A, the active metabolite of the fungus imperfectus Mycelia sterilia (Rosellinia sp.) isolated from the plant Camellia japonica in Japan, is described as a powerful broad-spectrum anthelmintic natural product with low toxicity in animals. Further CODPs such as PF1022B, C, D and E have been isolated from the same culture and their structures have been established. Both PF1022A and PF1022E serve as valuable starting materials for the synthesis of semi-synthetic CODP derivatives with improved intrinsic anthelmintic potency and broad-spectrum activity. It was found that in most cases the di-substituted PF1022A derivatives showed a greater (or equal) activity by oral application against the gastrointestinal nematode Haemonchus contortus compared to the corresponding mono-substituted PF1022A analogues as exemplified by emodepside. In order to get additional information on the bioactive conformation, emodepside was transformed into its mono- and tetra-thionated derivatives by isosteric replacement. In the light of the increased efficacy of these derivatives against H. contortus or Trichostrongylus colubriformis, it has been suggested that the asymmetric conformation clearly influences the anthelmintic activity of CODPs. Although useful synthetic pathways are available today for the preparation of the semi-synthetic CODP emodepside, the fermentative production of its bis-para-nitro and bis-para-amino precursors could be the process used for its industrial-scale production in the future.

Identification of thyroid hormone transporters in humans: different molecules are involved in a tissue-specific manner.

We have recently identified that rat organic anion transporters, polypeptide2 (oatp2) and oatp3, both of which transport thyroid hormones. However, in humans the molecular organization of the organic anion transporters has diverged, and the responsible molecule for thyroid hormone transport has not been clarified, except for human liver-specific transporter (LST-1) identified by us. In this study we isolated and characterized a novel human organic anion transporter, OATP-E from human brain. The isolated complementary DNA encodes a polypeptide of 722 amino acids with 12 transmembrane domains. A rat counterpart, oatp-E, was also identified. Homology analysis and the phylogenetic tree analysis revealed that OATP-E/oatp-E is a subfamily of the organic anion transporter. Human OATP-E transported 3,3',5-triiodo-L-thyronine (K(m), 0.9 microM), thyronine, and rT(3) in a Na(+)-independent manner. Although the clone was isolated from the brain, OATP-E messenger RNA was abundantly expressed in various peripheral tissues. The rat counterpart, oatp-E, also transported 3,3',5-triiodo-L-thyronine. In addition, in this study we revealed that human OATP, which is exclusively expressed in the brain, transported 3,3',5-triiodo-L-thyronine (K(m), 6.5 microM), T(4) (K(m), 8.0 microM), and rT(3). These data suggest that in humans, several different molecules are involved in transporting thyroid hormone: OATP in the brain, LST-1 in the liver, and OATP-E in peripheral tissues.

A non-chromatographic non-extraction radioimmunoassay for serum aldosterone.

A direct radioimmunoassay for serum aldosterone was developed using a highly specific sntibody and 8-anilino-1-naphthalene sulfonic acid as a blocking agent to inhibit the binding of aldosterone to serum proteins. 125I-labeled aldosterone was used as the labeled antigen and polyethylene glycol was used to separate antibody-bound and free aldosterone. The minimum detectable concentration was 1.5 pg/tube. There were excellent correlations between the present method and other methods, i.e., 1) a method using tritiated aldosterone, 2) a method using dichloromethane extraction before assay, and 3) a commercial kit method. The intra-assay coefficient of variation was 6.9%, and the inter-assay coefficient of variation was 10.7%. The values found in normal human serum were comparable with those reported using other methods. The present radioimmunoassay eliminates both extraction of aldosterone from serum and chromatographic separation and requires only 0.1 ml of serum for assay.

Measurement of plasma renin activity by a simple solid phase radioimmunoassay.

A solid phase RIA in which an antibody adsorbed onto polystyrene balls was developed to determine PRA. Complete inhibition of converting enzyme and angiotensinase during enzymatic reaction was achieved by phenyl methyl sulfonyl fluoride and EDTA combination. Pepstatin A was found to be an effective agent to block angiotensin I generation during the RIA, and the sample can be directly incubated at room temperature for RIA without any special treatment to inhibit renin activity. The intra- and interassay coefficients of variation (CV) were 5.5-6.9% and 3.7-8.2%, respectively. The recovery was 91.9-117% of added angiotensin I. The assay is sufficiently sensitive and reliable for routine use and correlates acceptably (r = 0.996) with an established RIA. The antibody-adsorbed balls were compared to the soluble antibody with respect to thermodynamic parameters. It was found that the apparent equilibrium constant of the antibody-adsorbed ball was reduced to approximately 1/2 sec of soluble antibody, which was predominately due to the decrease in unitary entropy change by adsorption.

17Beta-hydroxysteroid dehydrogenase types 1 and 2 in human placenta: an immunohistochemical study with correlation to placental development.

In estrogen metabolism, the enzymatic properties of the 17beta-hydroxysteroid dehydrogenase (17betaHSD) isozymes play very important roles in steroid hormone metabolism in various tissues, including the placenta. 17betaHSD type 1 catalyzes primarily the reduction of estrone (E1) to estradiol (E2), whereas 17betaHSD type 2 catalyzes primarily the oxidation of E2 to E1. In this study, we examined immunohistochemical localization of 17betaHSD types 1 and 2 in human placenta (31 cases) ranging from 4-40 weeks gestation. The immunoreactivity of 17betaHSD type 1 was exclusively detected in syncytiotrophoblast from 4 weeks gestation to term placenta. Immunoreactivity of 17betaHSD type 2 first appeared in endothelial cells of intravillous vessels at 12 weeks gestation, and the number of 17betaHSD type 2-positive endothelial cells markedly increased up to 19 weeks, then reached a plateau. We quantitatively evaluated the 17betaHSD type 2-positive endothelial cells in chorionic villi and determined the ratio of 17betaHSD type 2-positive endothelial cells using immunohistochemistry of CD34, an endothelial antigen, in serial mirror tissue sections and subsequent image analysis using CAS 200. CD34 was detected from 4 weeks gestation, and its positive areas continued to increase toward term. The 17betaHSD type 2-positive area per CD34-positive area markedly increased from 13 weeks gestation and reached a plateau at 19 weeks gestation, in which almost all endothelial cells were positive for 17betaHSD type 2. 17BetaHSD type 2, therefore, is considered to prevent the passage of excessive estrogens into the fetal circulation at endothelial cells of the intravillous fetal capillaries by catalyzing the inactivation ofE2 to E1.

17beta-hydroxysteroid dehydrogenase type 1 and 2 expression in the human fetus.

The present study investigates the expression patterns of 17beta-hydroxysteroid dehydrogenase (17betaHSD) isozymes in human fetal tissues to understand how estrogenic activity is regulated in the human fetus. Using enzyme assay, high 17betaHSD activity was detected in the placenta and liver, and low levels of 17betaHSD activity were also present in the gastrointestinal tract and kidney. After Northern blot analysis, we detected the messenger ribonucleic acid for 17betaHSD type 1 (17betaHSD1) only in the placenta, whereas that for 17betaHSD type 2 (17betaHSD2) was detected in the placenta, liver, gastrointestinal tract, and urinary tract at 20 gestational weeks. In RT-PCR analysis of the messenger ribonucleic acid transcripts, 17betaHSD 1 was predominantly expressed in the placenta, brain, heart, lung, and adrenal, whereas 17betaHSD2 expression was predominantly detected in the liver, gastrointestinal tract, and kidney. In addition, we detected 17betaHSD2 immunoreactive protein in surface epithelial cells of the stomach, absorptive epithelial cells of the small intestine and colon, hepatocytes of the liver, and interstitial cells surrounding the urinary tubules of the renal medulla. 17betaHSD2 in these tissues may be functioning in the prevention of in utero exposure of the fetus to excessive estradiol from the maternal circulation and amniotic fluids.

Hemodynamic, renal, and hormonal responses to alpha-human atrial natriuretic peptide in patients with congestive heart failure.

Hemodynamic, renal, and hormonal effects of intravenous bolus injection of 50 micrograms synthetic alpha-human atrial natriuretic peptide (alpha-hANP) were studied in eight patients with congestive heart failure. alpha-hANP caused significant reductions in mean blood pressure and systemic vascular resistance. These responses were sustained up to 90 minutes and not accompanied by reflex tachycardia. Cardiac index and stroke volume index increased significantly at 90 minutes and pulmonary capillary wedge pressure, pulmonary arterial pressure, and mean right atrial pressure remained unchanged. Urine volume, urinary sodium excretion, creatinine clearance, and fractional excretion of sodium increased significantly, but fractional excretion of potassium and phosphate did not change. Elevated plasma renin activity, plasma aldosterone, and norepinephrine were suppressed after the injection of alpha-hANP. The bolus injection of this peptide has moderately hypotensive, vasorelaxant, and natriuretic effects in patients with congestive heart failure.

Effects of valproate on biogenesis and function of liver mitochondria.

The mitochondrial protein content of liver increased, and mitochondria enlarged after prolonged administration of valproic acid (VPA: N-dipropylacetic acid) to rats. The mitochondria showed low specific activities of membrane-bound enzymes, decreased content of cytochrome aa3, and decreased respiration in states 3 and 4. In vitro studies of amino acid incorporation suggested a decrease in protein synthesis in liver mitochondria from VPA-treated rats. Prolonged administration of VPA seems to impair mitochondrial structure and function.

Mutation in the caveolin-3 gene causes a peculiar form of distal myopathy.

The authors describe a patient with sporadic distal myopathy associated with reduced caveolin-3 in muscle fibers in which the muscle atrophy was restricted to the small muscles of the hands and feet. Gene analysis disclosed a heterozygous 80 G-->A substitution in the caveolin-3 gene that was identical to that of reported cases of elevated serum creatine kinase. This patient further demonstrated possible clinical heterogeneity of myopathies with mutations in the caveolin-3 gene.

Abnormal kainic acid receptor density and reduced seizure susceptibility in dystrophin-deficient mdx mice.

Duchenne muscular dystrophy is characterized by a defect in dystrophin, which often causes mental retardation in addition to progressive muscular weakness. As dystrophin is localized in synaptic regions of the CNS, cognitive abnormalities associated with Duchenne muscular dystrophy are attributable to synaptic dysfunction. We report that dystrophin-deficient mdx mice were more resistant to kainic acid-induced seizures but not to GABA antagonist-induced seizures compared with the control mice. The kainic-acid receptor density in the brain was significantly lower in the mdx than in the control, although the density of muscarinic cholinergic receptors, another important neurotransmitter receptor for cognitive function, was normal. Moreover, mdx had significantly lower Timm staining intensity in the mossy fibers, which originate from the dentate granule cells and terminate on the pyramidal cells in the CA3 of the hippocampus. These results suggest that an instability of neurotransmitter receptors, such as kainate-type glutamate receptors, on synaptic membranes due to the disruption of dystrophin complex induces inefficient neurotransmission in Duchenne muscular dystrophy patients.

Cell shape change and cytosolic Ca2+ in human umbilical-vein endothelial cells stimulated with thrombin.

We quantified thrombin-induced endothelial cells shape change and investigated the role of Ca2+ in such shape change. We used the fluorescent Ca2+ indicator, fura2, to measure both shape change as cell size and intracellular free Ca2+ ([Ca2+]i), in cultured human umbilical-vein endothelial cells (HUVEC). Thrombin induced concentration-dependent decreases in cell size (percentage of cell size at 6 min after stimulation with 0.01 U/ml, 0.1 U/ml, or 1 U/ml thrombin) was 90.1 +/- 1.5%, 78.1 +/- 2.4%, and 40.9 +/- 2.4%, respectively. Thrombin also increased [Ca2+]i in a concentration-dependent manner. Both depletion of extracellular Ca2+, and also the addition of W5, a calmodulin antagonist, inhibited thrombin-induced size reduction. These results indicate an association between shape change and [Ca2+]i mobilization in human endothelial cells stimulated by thrombin.

Gonadal suppression by a GnRH analogue does not alter somatic growth in rats with complete GH deficiency.

Sexual dimorphism of somatic growth in rats appears to reflect differing actions of sex steroids. However, mechanisms of gonadal steroid effects on the somatotropic axis are incompletely understood. To evaluate whether GH is involved in the effects of long-term gonadal suppression on somatic growth in rats, a GnRH agonistic analogue (GnRHa) was administered to normal Sprague-Dawley rats (controls) and to a strain of rats with complete growth hormone deficiency (GHD; n=4-6 in each group). Subcutaneous injection of GnRHa (2 mg/kg) or saline were given within 48 h after birth and repeated every 3 weeks. GnRHa treatment significantly reduced serum gonadal steroid levels in rats of both sexes with small testes in males and impaired development of internal genitalia in females. GnRHa-treated control females became significantly heavier (P<0.01 ANOVA for repeated measures) than saline-treated rats beginning at 8 weeks. However, female GHD rats with GnRHa treatment did not differ in body weight from rats receiving saline. In male rats, GnRHa treatment did not change body weight in either control or GHD rats. Serum IGF-I concentrations did not differ between treatment groups in GHD and control rats of either sex. Hepatic GH binding was reduced significantly by GnRHa treatment in female control rats (P<0.01), but not in female GHD rats. These data suggest that sexual dimorphism in body size and its modulation by estrogens are independent of circulating IGF-I levels suggesting non-endocrine IGF-I-mediated mechanisms, and that GH-induced somatic growth is modulated by estrogens, but not androgens, in rats.

Detection of apoptosis in acute promyelocytic leukemia cells in vivo during differentiation-induction with all-trans retinoic acid in combination with chemotherapy.

In two patients with acute promyelocytic leukemia (APL) treated with all-trans retinoic acid (ATRA) in combination with chemotherapy, we demonstrated that cells with apoptotic morphology were recognized in a small fraction of whole blood cells (0.2-0.4%) at the regression phase of leukocytosis with mostly maturing aberrant granulocytes of APL clone origin. Moreover, in a light-density cell fraction ( < 1.077) of peripheral blood or marrow cells obtained from three patients, DNA fragmentation was detected by agarose gel electrophoresis temporarily. These findings may suggest that, during the treatment with ATRA, a small fraction of APL cells maturing toward the stage of terminal differentiation underwent apoptosis in vivo, which might be enriched by the combination of chemotherapy to ATRA administration.

Oral-facial-digital syndrome with hypothalamic hamartoma, postaxial ray hypoplasia of the limbs, and vagino-cystic communication: a new variant?

We report on a 20-month-old girl with hypothalamic hamartoma, left cerebral atrophy, tongue nodules, oral frenula, micrognathia, hypoplasia of the left ulna, the fibulae, and right tibia, polysyndactyly of the hands and feet, vagino-cystic drainage with hydrometrocolpos, megaloureters, and hydronephrosis, agenesis of urethra, complex partial seizures, and central precocious puberty. The differential diagnosis is discussed. We conclude that the malformation complex in this girl is an oral-facial-digital syndrome, but is different from any of the 11 known subtypes.

Trisomy 22 mosaicism syndrome and Ullrich-Turner stigmata.

Mosaic trisomy 22, ascertained in three unrelated patients, was found to be associated with body asymmetry and signs of the Ullrich-Turner syndrome including short stature, ptosis, webbed neck, nevi, cubitus valgus, dysplastic nails, malformed great vessels, and abnormal ovaries. These anomalies in trisomy 22 mosaicism have not been emphasized heretofore. In each of our patients, trisomy 22 mosaicism was found only in fibroblasts. In one patient, the trisomy resulted from a paternal first meiotic nondisjunction, and in the 46,XX cells, both chromosomes 22 were of paternal origin.

Glycogen storage disease type Ia: molecular diagnosis of 51 Japanese patients and characterization of splicing mutations by analysis of ectopically transcribed mRNA from lymphoblastoid cells.

Glycogen storage disease type Ia (GSD-Ia) is an autosomal recessive disorder of glycogen metabolism caused by a deficiency of glucose-6-phosphatase (G6Pase) that is expressed in the liver, kidney, and intestinal mucosa. Clinical manifestations include short stature, hepatomegaly, hypoglycemia, hyperuricemia, and lactic acidemia. To elucidate a spectrum of the G6Pase gene mutations and their frequencies, we analyzed mutations in 51 unrelated Japanese patients with GSD-Ia. The most prevalent mutation was g727t, accounting for 88 of 102 mutant alleles examined, followed by R170X mutation, which accounted for 6 mutant alleles, and R83H mutation which was observed in 3 mutant alleles. In addition, 3 different, novel mutations, IVS1-1g