Toshiyuki Hikita

Epidermal growth factor receptor tyrosine kinase is modulated by GM3 interaction with N-linked GlcNAc termini of the receptor

Epidermal growth factor receptor (EGFR) at membrane microdomains plays an essential role in the growth control of epidermal cells, including cancer cells derived therefrom. Ligand-dependent activation of EGFR tyrosine kinase is known to be inhibited by ganglioside GM3, but to a much lesser degree by other glycosphingolipids. However, the mechanism of the inhibitory effect of GM3 on EGFR tyrosine kinase has been ambiguous. The mechanism is now defined by binding of N-linked glycan having multiple GlcNAc termini to GM3 through carbohydrate-to-carbohydrate interaction, based on the following data: (i) EGFR (molecular mass, ≈170 kDa) has N-linked glycan with GlcNAc termini, as probed by mAb (J1) or lectin (GS-II); (ii) GS-II-bound EGFR also bound to anti-EGFR Ab as well as to GM3-coated beads; (iii) GM3 inhibitory effect on EGFR tyrosine kinase was abrogated in vitro by coincubation with glycan having multiple GlcNAc termini and in cell culture in situ incubated with the same glycan; and (iv) cells treated with swainsonine, which increased expression of complex-type and hybrid-type glycans with GlcNAc termini, displayed higher inhibition of EGFR kinase by GM3 than swainsonine-untreated control cells. A similar effect was observed with 1-deoxymannojirimycin, which increased hybrid-type structure in addition to major accumulation of high mannose-type glycan. These findings indicate that N-linked glycan with GlcNAc termini linked to EGFR is the target to interact with GM3, causing inhibition of EGF-induced EGFR tyrosine kinase.

Sphingosine-dependent Protein Kinase-1, Directed to 14-3-3, Is Identified as the Kinase Domain of Protein Kinase Cδ

Some protein kinases are known to be activated by d-erythro-sphingosine (Sph) or N,N-dimethyl-d-erythro-sphingosine (DMS), but not by ceramide, Sph-1-P, other sphingolipids, or phospholipids. Among these, a specific protein kinase that phosphorylates Ser60, Ser59, or Ser58 of 14-3-3β, 14-3-3η, or 14-3-3ζ, respectively, was termed “sphingosine-dependent protein kinase-1” (SDK1) (Megidish, T., Cooper, J., Zhang, L., Fu, H., and Hakomori, S. (1998) J. Biol. Chem. 273, 21834–21845). We have now identified SDK1 as a protein having the C-terminal half kinase domain of protein kinase Cδ (PKCδ) based on the following observations. (i) Large-scale preparation and purification of proteins showing SDK1 activity from rat liver (by six steps of chromatography) gave a final fraction with an enhanced level of an ∼40-kDa protein band. This fraction had SDK1 activity ∼50,000-fold higher than that in the initial extract. (ii) This protein had ∼53% sequence identity to the Ser/Thr kinase domain of PKCδ based on peptide mapping using liquid chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry data. (iii) A search for amino acid homology based on the BLAST algorithm indicated that the only protein with high homology to the ∼40-kDa band is the kinase domain of PKCδ. The kinase activity of PKCδ did not depend on Sph or DMS; rather, it was inhibited by these sphingoid bases, i.e. PKCδ did not display any SDK1 activity. However, strong SDK1 activity became detectable when PKCδ was incubated with caspase-3, which releases the ∼40-kDa kinase domain. PKCδ and SDK1 showed different lipid requirements and substrate specificity, although both kinase activities were inhibited by common PKC inhibitors. The high susceptibility of SDK1 to Sph and DMS accounts for their important modulatory role in signal transduction.

Sumatriptan as a treatment for cyclic vomiting syndrome: A clinical trial

Background and objective: Cyclic vomiting syndrome (CVS) is associated with migraine. This study aimed to evaluate the efficacy of sumatriptan in treating CVS. Methods: Twelve patients were enrolled in this trial. Sumatriptan was administered either subcutaneously [(age × 4 + 20)/100 × 3 mg] or by nasal spray (NS; 20 mg). Response to the treatment was classified as complete, effective, or noneffective. Results: Eleven patients, who presented with 35 attacks, were treated by subcutaneous injection of sumatriptan. The treatment was responsive in 19 attacks. The efficacy of sumatriptan was high in attacks that occurred in cases with a family history of migraine compared to those without (p = .0482). Five patients were treated with sumatriptan NS for six attacks. The treatment was completely responsive in two of six attacks. We observed no adverse effects associated with sumatriptan treatment in this trial. Conclusion: We conclude that sumatriptan has potential efficacy in treating of patients with CVS.

Effective prophylactic therapy for cyclic vomiting syndrome in children using valproate.

This trial sought to evaluate our experience using the antimigraine prophylactic drug, use of valproate for the prophylactic management of cyclic vomiting syndrome (CVS) in children. Thirteen children diagnosed with severe CVS were enrolled. Prophylactic therapy consisted of valproate administered at a dose of 10-40 mg/kg/day. Upon enrollment in the study, all patients underwent diagnostic tests to rule out organic causes of their symptoms. Vomiting was severe enough in all patients to cause dehydration requiring hospitalization for intravenous rehydration. Nine of 13 patients did not respond to numerous previous medical therapies like propranolol, amitriptyline, cyproheptadine, phenobarbital, phenytoin, and carbamazepine. Three of 13 patients required combination therapy with valproate and phenobarbital. Of the 13 patients, two showed complete resolution of their symptoms, nine had marked improvement in their symptoms, as evidenced by infrequent attacks of reduced severity, and two failed to respond to valproate therapy. Four patients experienced relapse with a decreased dosage of valproate. Side effects associated with long-term valproate administration were not observed. Valproate appears to be effective for the prophylactic management of severe CVS, with 85% of all patients achieving at least a reduction in the frequency of attacks.

Fatty acid compositions of colostrum, cord blood, maternal blood and major infant formulas in Japan

Lipid profiles in colostrum, cord blood, maternal blood and major infant formulas in Japan were analyzed. In the first part of the study, colostrum obtained from 36 normal delivery women and six kinds of infant formulas provided by three major milk companies were analyzed for their fatty acid composition using capillary gas‐lipid chromatography. Although enriched with docosahexaenoic acid (DHA), the percent composition of DHA in the six infant formulas (0.15–0.21%) was significantly lower than that in the colostrum (1.1 ± 0.54). Arachidonic acid (AA) and eicosapentaenoic acid (EPA) were present in the colostrum but not detectable in the infant formulas. It is recommended that although the exact amount of specific fatty acids needed in the infant diet was not completely known, to be as close as possible to natural breast milk, the level of DHA, EPA and AA should be raised in the infant formulas. In the second part of the study, 19 pairs of maternal and cord blood were analyzed for their lipid profile. All samples were from normal vaginal delivery. The measurement of cholesterol, triglycerides, phospholipids and free fatty acids was performed with commercially available enzymatic methods on an automated discrete random access analyzer. Total fatty acid was determined as described in the first part of the study. The results were analyzed with Spearmans rank correlation coefficient. No correlation could be found between maternal and fetal concentrations of cholesterol, triglycerides, phospholipids or total fatty acids. Correlation could be found in non‐esterified fatty acids, in palmitic acids, and oleic acid levels. It was concluded that the lipid transport and metabolism in the fetal‐placenta unit is complex and further delicate investigation is required.

Regional cerebral blood flow abnormalities in patients with kawasaki disease.

Purpose: Kawasaki disease (KD) is an acute febrile disorder of unknown etiology. Brain single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI) help in detecting regional cerebral blood flow abnormalities and brain damage. The usefulness of SPECT and MRI in patients with KD was evaluated. Materials and Methods: All 22 patients with KD underwent brain SPECT using Tc-99m-hexamethyl propylene amine oxime from 6 days to 3 years after onset, and 8 patients underwent brain MRI. Of the 22 patients, 4 had neurologic symptoms. Case 1 showed prolonged apnea; case 2, prolonged disturbance of consciousness; and cases 3 and 4 generalized tonic-clonic seizures. Initial brain SPECT showed localized hypoperfusion in 4 and 13 patients with and without neurologic symptoms, respectively. Results: All patients with neurologic symptoms underwent follow-up SPECT; localized hypoperfusion was detected between 1- and 6-month follow-up in 3 of these patients. Six patients without neurologic symptoms underwent follow-up SPECT. Localized hypoperfusion was detected at approximately 1- to 11-month follow-up in 4 of these patients. Diffusion-weighted imaging revealed abnormal high-intensity areas in the corpus callosum in case 1. Case 2 showed a bilateral chronic subdural hematoma with decreased size and ischemic changes, and case 3 showed bilateral hippocampal atrophy and left hippocampal sclerosis. Conclusions: Because the occurrence of localized hypoperfusion is possibly not restricted to only the acute phase in KD, brain SPECT and MRI should also be performed in KD patients with neurologic symptoms.

Severe infantile myotubular myopathy with complete atrioventricular block

© 2008 Japan Pediatric Society In 1966 Spiro et al. described the fi rst case of myopathy in which structures resembling fetal myotubes persisted into adult life. 1 In 1969 the fi rst severe infantile form was described. 2 X-linked severe infantile myotubular myopathy is a severe muscular disease characterized by neonatal hypotonia, severe global muscular weakness and respiratory distress in affected male subjects. 3 The MTM1 gene, which is mutated in X-linked severe infantile myotubular myopathy, is located on Xq28 and was identifi ed in 1996 using positional cloning. 4 The majority of patients die during the fi rst year of life. The cause of death is usually respiratory insuffi ciency caused by severe muscle hypotonia and weakness. Prolonged survival is observed in the milder form, or as a result of prolongation of ventilation support. 3 None of the studies in the literature discuss the complete atrioventricular block (CAVB) in this disorder. Here, we describe an autopsy case of X-linked severe infantile myotubular myopathy with CAVB.

A catalytic carbohydrate contributes to bacterial antibiotic resistance

Penicillins are widespread in nature and lethal to growing bacteria. Because of the severe threat posed by these antibiotics, bacteria have evolved a wide variety of strategies for combating them. Here, we describe one unusual strategy that involves the activity of a catalytic carbohydrate. We show that the cyclic oligosaccharide, β-cyclodextrin (βCD), can hydrolyze, and thereby inactivate, penicillin in vivo. Moreover, we demonstrate that this catalytic activity contributes to the antibiotic resistance of a bacterium that synthesizes this oligosaccharide in the laboratory. Taken together, these data not only expand our understanding of the biochemistry of penicillin resistance, but also provide the first demonstration of natural carbohydrate-mediated catalysis in a living system.

Immunochromatography test for rapid diagnosis of Mycoplasma pneumoniae infection

The sensitivity and specificity of a new rapid Mycoplasma pneumoniae antigen immunochromatography (IC) test, DK‐MP‐001, were determined using particle agglutination (PA) antibody response and loop‐mediated isothermal amplification (LAMP) gene detection as the gold standard. Of 165 patients, 59 were diagnosed with M. pneumoniae infection based on a ≥fourfold rise of serum PA antibody during the course of the illness. Of the first visit swabs, 60 were positive for M. pneumoniae on LAMP, and 49 were positive for M. pneumoniae antigen on IC test. Compared with PA antibody and LAMP, the sensitivity/specificity of the IC test were 81.4% (48/59) and 99.1% (105/106); and 81.7% (49/60) and 100% (105/105), respectively. IC test detected antigen in pharyngeal swabs more sensitively than in nasal swabs for the same subjects (P < 0.05). The IC test performs well enough to be used with pharyngeal swabs at the first examination.

Prevalence of abdominal migraine and recurrent abdominal pain in a Japanese clinic

Prevalence of abdominal migraine (AM) and recurrent abdominal pain (RAP) was evaluated in patients who visited Hikita Pediatric Clinic between May 2010 and April 2015. Patient data were collected prospectively using a questionnaire. Out of a total of 3611 cases, observed prevalence was 2.44% for repeated abdominal pain over a period of ≥3 months, 1.47% for RAP, and 0.19% for AM. Duration of abdominal pain was longer for AM than for non‐AM RAP. Certain clinical features were significantly different between AM and non‐AM RAP. No correlations were found among age at onset, frequency of attack, and duration of attack for various types of RAP. It was difficult to determine useful diagnostic criteria for distinguishing between AM and non‐AM RAP. They did not appear to be separate disease entities but, instead, lie on a disease spectrum. The present prevalence of AM (0.19%) was lower than that in many previous studies from countries other than Japan.