Hitoshi Sato

THERAPEUTIC BASIS OF GLYCYRRHIZIN ON CHRONIC HEPATITIS B

Glycyrrhizin, a major component of a herb (licorice), has been intravenously used for the treatment of chronic hepatitis B in Japan and improves liver function with occasional complete recovery from hepatitis. This substance modifies the intracellular transport and suppresses sialylation of hepatitis B virus (HBV) surface antigen (HBsAg) in vitro. This study was designed to clarify the pharmacological basis for its effectiveness. The structure-bioactivity relationship of glycyrrhizin, glycyrrhetic acid 3-O-monoglucuronide and glycyrrhetic acid was determined, and glycyrrhetic acid was found to be the most active of them. The amounts of three substances bound to the liver were evaluated in guinea pigs after intravenous administration of glycyrrhizin. Glycyrrhizin and glycyrrhetic acid 3-O-monoglucuronide were detected at concentrations of 31.8-1.3 micrograms/g of liver, but glycyrrhetic acid was not detected. When glycyrrhizin attained these concentrations in the cellular fraction of the PLC/PRF/5 cell culture, it suppressed the secretion of HBsAg as reported previously. These results indicated that glycyrrhizin administered intravenously might bind to hepatocytes at the concentration at which glycyrrhizin could modify the expression of HBV-related antigens on the hepatocytes and suppress sialylation of HBsAg.

Antiepileptic effects of tiagabine, a selective GABA uptake inhibitor, in the rat kindling model of temporal lobe epilepsy

Summary: Purpose: We determined the antiepileptic profile of tiagabine (TGB), a selective γ‐aminobutyric acid (GABA) uptake inhibitor, in the rat kindling model of temporal lobe epilepsy (TLE).

Varicella-zoster virus open reading frame 2 encodes a membrane phosphoprotein that is dispensable for viral replication and for establishment of latency

ABSTRACT Varicella-zoster virus (VZV) encodes six genes that do not have homologs in herpes simplex virus. One of these genes, VZV open reading frame 2 (ORF2), was expressed as a 31-kDa phosphoprotein in the membranes of infected cells. Unlike equine and bovine herpesvirus type 1 ORF2 homologs that are associated with virions, VZV virions contained no detectable ORF2 protein. The ORF2 deletion mutant established a latent infection in cotton rats at a frequency and with a number of VZV genomes similar to that of the parental virus. ORF63 transcripts, a hallmark of latent infection, were present in ganglia latently infected with both the ORF2 deletion mutant and parental VZV. Thus, ORF2 is the first VZV gene shown to be dispensable for establishment of latent infection in an animal model.

Time-dependent and regional expression of GABA transporter mRNAs following amygdala-kindled seizures in rats

To investigate the role played by GABA transporters in epileptic seizures, we examined time-dependent and regional changes in expression of GAT-1 and GAT-3 GABA transporter mRNA in amygdala-kindled rat brain using an in situ hybridization method. GAT-1 mRNA was significantly increased bilaterally in the hippocampal dentate gyrus (111-116%) at 1 h after kindled generalized seizures. GAT-1 mRNA was also significantly increased bilaterally in the hippocampal subfields (CA1-4 and dentate gyrus [110-117%]) at 4 h after kindled seizures. There were no significant changes in GAT-1 mRNA level in the amygdalar nuclei, pyriform cortex or cerebral cortex either ipsilaterally or contralaterally at any time after kindled seizures. In contrast, GAT-3 mRNA was significantly increased bilaterally in the amygdalar nuclei and in the contralateral pyriform cortex and cerebral cortex 1 h after seizures. Since all these changes returned to control levels by 8 or 24 h after kindled seizures, the increases in GABA transporter mRNA appeared to be transient responses to seizure activity. These findings indicate that GAT-1 subtype transporter is specifically involved in seizure activity in the hippocampus, while GAT-3 subtype transporter is mainly involved in seizure activity in the amygdalar nuclei and pyriform cortex following amygdala-kindled generalized seizures.

Varicella-Zoster Virus (VZV) ORF17 Protein Induces RNA Cleavage and Is Critical for Replication of VZV at 37oC but Not 33oC

ABSTRACT Varicella-zoster virus (VZV) open reading frame 17 (ORF17) is homologous to herpes simplex virus (HSV) UL41, which encodes the viral host shutoff protein (vhs). HSV vhs induces degradation of mRNA and rapid shutoff of host protein synthesis. An antibody to ORF17 protein detected a 46-kDa protein in VZV-infected cells. While HSV vhs is located in virions, VZV ORF17 protein was not detectable in virions. ORF17 protein induced RNA cleavage, but to a substantially lesser extent than HSV-1 vhs. Expression of ORF17 protein did not inhibit expression from a β-galactosidase reporter plasmid, while HSV type 1 vhs abolished reporter expression. Two VZV ORF17 deletion mutants were constructed to examine the role of ORF17 in virus replication. While the ORF17 VZV mutants grew to peak titers that were similar to those of the parental virus at 33°C, the ORF17 mutants grew to 20- to 35-fold-lower titers than parental virus at 37°C. ORF62 protein was distributed in a different pattern in the nuclei and cytoplasm of cells infected with an ORF17 deletion mutant at 37°C compared to 33°C. Inoculation of cotton rats with the ORF17 deletion mutant resulted in a level of latent infection similar to that produced by inoculation with the parental virus. The importance of ORF17 protein for viral replication at 37°C but not at 33°C suggests that this protein may facilitate the growth of virus in certain tissues in vivo.

Variation in GABA-A subunit gene copy number in an autistic patient with mosaic 4 p duplication (p12p16)†

Autism has been associated with chromosomal aberrations, including duplications at chromosome 4, and the identification of genetic factors contributing to the etiology of this disease is the focus of much research. Here we report a Japanese girl with mosaic of chromosome 4p duplication, mos 46,XX,dup(4)(p12p16)[54]/46,XX[6], who was diagnosed with autism at 3 years of age. Fluorescence in situ hybridization (FISH) with probes covering the region spanning a cluster of the gamma aminobutyric acid A (GABA‐A) receptor subunit genes in the proximal short arm of chromosome 4 demonstrated total three signals for the GABRG1, GABRA4, and GABRA2 genes, but only two signals for GABRB1. This suggests that aberrant copy number of the GABA‐A receptor subunit genes may contribute to the etiology of autism in this patient.

Varicella-Zoster Virus Open Reading Frame 21, Which Is Expressed during Latency, Is Essential for Virus Replication but Dispensable for Establishment of Latency

ABSTRACT Varicella-zoster virus (VZV) open reading frame 21 (ORF21) is one of at least five VZV genes expressed in latently infected human and rodent ganglia. To determine whether ORF21 is required for latent and lytic infection, we deleted 99% of ORF21 from the viral genome. The ORF21 deletion mutant virus could be propagated only in a cell line expressing the ORF21 protein. Insertion of the herpes simplex virus type 1 (HSV-1) homolog of VZV ORF21, HSV-1 UL37, into the ORF21 deletion mutant failed to complement the mutant for growth in cell culture. Inoculation of cotton rats with the ORF21 deletion virus resulted in latent infection in numbers of animals similar to those infected after inoculation with the parental virus. The mean numbers of latent VZV genomes were similar in animals infected with parental and ORF21 deletion viruses. Transcription of ORF63, another latency-associated gene, was detected in ganglia from similar numbers of animals infected with the mutant and parental viruses. Thus, ORF21 is the first VZV gene expressed during latency that has been shown to be dispensable for the establishment of latent infection.

Acute encephalopathy in children with Dravet syndrome

Purpose:  The occurrence of acute encephalopathy in children with Dravet syndrome has been reported sporadically. This study clarified the features of acute encephalopathy in children with Dravet syndrome.

Varicella-Zoster Virus ORF47 Protein Kinase, Which Is Required for Replication in Human T Cells, and ORF66 Protein Kinase, Which Is Expressed during Latency, Are Dispensable for Establishment of Latency

ABSTRACT Varicella-zoster virus (VZV) results in a lifelong latent infection in human sensory and cranial nerve ganglia after primary infection. VZV open reading frame 47 (ORF47) and ORF66 encode protein kinases that phosphorylate several viral proteins, including VZV glycoprotein gE and ORF32, ORF62, and ORF63 proteins. Here we show that the ORF47 protein kinase also phosphorylates gI. While ORF47 is essential for virus replication in human T cells and skin, we found the gene to be dispensable for establishment of latent infection in dorsal root ganglia of rodents. ORF66 protein is expressed during latency. Rodents infected with VZV unable to express ORF66 developed latent infection at a rate similar to that for the parental virus. ORF63 transcripts, a hallmark of VZV latency, were also detected in similar numbers of animals infected with the ORF47 and ORF66 mutants and with the parental virus. VZV mutants unable to express four of the six genes that do not have herpes simplex virus (HSV) homologs (ORFs 1, 13, 32, 57) were also unimpaired for establishment of latency. While a truncated HSV VP16 mutant was previously reported to be unable to establish latency in a mouse model, we found that VZV with a deletion of ORF10, the homolog of HSV VP16, was dispensable for establishment of latency. Thus, seven genes, including one expressed during latency, are dispensable for establishing latent VZV infection.

BW1003C87, phenytoin and carbamazepine elevate seizure threshold in the rat amygdala-kindling model of epilepsy

We examined the anticonvulsant effects of BW1003C87 (5-(2,3,5-trichlorophenyl)-2,4-diaminopyrimidine ethane sulphonic acid), which is structurally related to the new antiepileptic drug, lamotrigine, and compared its effects to those of the conventional antiepileptic drugs, phenytoin and carbamazopine, using the rat amygdala-kindling model of epilepsy. BW1003C87 (2.5-10 mg/kg, i.p.) had potent and long-lasting (48 h after single administration) effects on amygdala-kindled seizures. The effects of BW1003C87 were completely reversed when the stimulus intensity was increased to 2 or 3 times the threshold determined. Since the same effects on seizure threshold were obtained for phenytoin and carbamazepine in the present study and for lamotrigine in our previous study, we propose that the principal mechanism of these antiepileptic drugs, which act primarily on voltage-sensitive Na+ channels, is significant elevation of the seizure threshold in epileptogenic foci and that BW1003C87 has a profile similar to that of these drugs.