Paternal mosaicism of an STXBP1 mutation in OS.

Ohtahara syndrome (OS) is one of the most severe and earliest forms of epilepsy. We have recently identified that the de novo mutations of STXBP1 are important causes for OS. Here we report a paternal somatic mosaicism of an STXBP1 mutation. The affected daughter had onset of spasms at 1 month of age, and interictal electroencephalogram showed suppression-burst pattern, leading to the diagnosis of OS. She had a heterozygous c.902+5G>A mutation of STXBP1, which affects donor splicing of exon 10, resulting in 138-bp insertion of intron 10 sequences in the transcript. The mutant transcript had a premature stop codon, and was degraded by nonsense-mediated mRNA decay in lymphoblastoid cells derived from the patient. High-resolution melting analysis of clinically unaffected parental DNAs suggested that the father was somatic mosaic for the mutation, which was also suggested by sequencing. Cloning of PCR products amplified with the paternal DNA samples extracted from blood, saliva, buccal cells, and nails suggested that 5.3%, 8.7%, 11.9%, and 16.9% of alleles harbored the mutation, respectively. This is a first report of somatic mosaicism of an STXBP1 mutation, which has implications in genetic counseling of OS.

Effect of voluntary participation on an alternating and a simultaneous prisoner's dilemma.

We studied the evolution of cooperation in the framework of evolutionary game theory, implementing voluntary participation in the prisoner's dilemma. Although previous studies have tried to overcome the dilemma by introducing voluntary participation called a "loner," the question of which strategies among various strategies including voluntary participation are adaptive under competitive circumstances is still an unsolved puzzle. Here we have developed a model that consists of all possible strategies using a one-period memory of past actions. This model enables us to analyze a "melting pot" of strategies, wherein several strategies interact and compete with each other. Our results revealed that one strategy, in which one escapes if a partner defects or cooperates if a partner becomes a loner, dominates and maintains cooperation in an alternating prisoner's dilemma game. However, the so-called "win-stay, lose-shift" strategy dominates in a simultaneous prisoner's dilemma game. Our simulations clearly show that voluntary participation in the prisoner's dilemma game works in the alternating situation rather than the simultaneous one.

Inotropic agents differentially inhibit the induction of nitric oxide synthase by endotoxin in cultured macrophages.

We investigated the effects of inotropic agents with phosphodiesterase III inhibitory properties, amrinone, pimobendan and vesnarinone, and cell permeable cyclic nucleotide analogue, 8-bromo adenosine 3'5'-cyclic monophosphate (8 Br-cAMP) on the induction of nitric oxide synthase (NOS) by lipopolysaccharide in J774A.1 macrophages in vitro. Although all three inotropic agents inhibited nitrite accumulation, the degree of inhibition was different, with pimobendan being the most potent inhibitor and amrinone the least. Vesnarinone inhibited nitrite formation biphasically. 8 Br-cAMP increased nitrite production at high concentrations, suggesting that the inhibitory effects of inotropic agents could not be explained by an increase in cAMP. Although differential inhibition of inducible NOS by inotropic agents may explain the different effects of these drugs in patients with heart failure, further study is necessary to reach this conclusion.

One amino Acid change in cucumber mosaic virus RNA polymerase determines virulent/avirulent phenotypes on cowpea.

ABSTRACT The elicitation of the hypersensitive response (HR) is known to depend on the interaction between a resistance gene of a host plant and a corresponding avirulence gene of a pathogen. The cv. Kurodane-Sanjaku of cowpea (Vigna unguiculata) has the Cry locus that confers resistance against cucumber mosaic virus strain Y (CMV-Y). The resistance is overcome by infection with a legume strain of CMV (CMV-L). RNA 2, which codes for the 2a protein, a subunit of the viral replicase components, has been known to control virulent/avirulent phenotypes. We generated chimeric constructs of full-length cDNA clones of RNA 2 of both strains and inoculated infectious transcripts to delimit the domain controlling symptoms. A 243-base pair fragment containing a coding region for the GDD RNA-dependent RNA polymerase core sequence was shown to be responsible for the phenotypic differences. From sequence alignment analysis, two amino acids (Phe631 and Ala641) of the HR-type 2a protein encoded in this fragment were specifically exchanged to Tyr and Ser, respectively, in the 2a proteins of resistance-breaking strains. Point mutations introduced into RNA 2 backgrounds of both strains that were designed to change the amino acid at position 631 resulted in a change of symptoms, indicating that a single nucleotide change determines the reactions elicited by both strains. Analysis for one additional mutant RNA 2 showed that symptom determination may be correlated with the nature of the lateral chain of amino acid 631.

Inhibition of turkey herpesvirus replication by anti-cellular serum.

Quail embryo fibroblasts were used to investigate how rabbit and chicken antisera against chicken erythrocytes carrying different B alleles of the major histocompatibility antigens affect the neutralization of herpesvirus of turkeys (HVT). Although the neutralizing activities of these antisera were rather low, the HVT propagated in chicken embryo fibroblasts (CEF) from certain genotypic embryos was neutralized more by the antisera to the corresponding erythrocytes. After absorption of these antisera with homologous erythrocytes, the neutralizing activity of the absorbed sera was reduced slightly. These results reveal that the virion antigens of HVT might be partially associated with the host cell antigens of the fibroblast infected with the virus. The virus grown in these cells might incorporate the host cell antigens, including histocompatibility antigens, into the virion envelope.

Immunocompetences and Marek's disease resistance in three pairs of chicken lines selected for different immunological characters.

Three pairs of chicken lines selected for high (H) and low (L) graft vs. host reaction (GVHR) competences, serum immunoglobulin G (IgG) levels, and antibody responses to Leucocytozoon caulleryi were examined for their immunocompetences and Marek's disease (MD) resistance. The GVHR-H and GVHR-L lines were further divided into two sublines according to their major histocompatibility B genotype. Immune responses to sheep erythrocytes (SRBC), bovine serum albumin (BSA), and a lipopolysaccharide (LPS) were compared between the high and low lines of each pair of selected lines. Significant differences were found in responses to SRBC and LPS in IgG-selected lines and in response to BSA in Leucocytozoon-selected lines. In all three instances antibody titers of the H line were higher than those of the L line. The GVHR competence expressed by the splenomegaly index (SI) was also significantly different between the H and L lines of all three selected-line pairs. The SI values in the GVHR-selected and IgG-selected lines were higher in the H line than in the L line, whereas those in the Leucocytozoon-selected lines were lower in the H line. Differences in MD incidence and in MD mortality were found between the GVHR-selected B11B11 subline and the IgG-selected lines. In both instances the L line was more resistant to MD than the H line.

Parabiosis between avian embryos selected for high and low competences of the graft-versus-host reaction.

Parabiosis of chicken embryos was conducted to determine if changes in immunological competences could occur by parabiosis between lines of chickens selected for high (H) and low (L) graft-versus-host reaction (GVHR). Eggs of the B9B9 genotype from the H line (H-B9B9) were parabiosed to eggs of the B11B11 from the L line (L-B11B11) and also the H-B11B11 were parabiosed to the L-B9B9. There were significant differences in hatchability between the line-genotype groups. The percentage of blood chimerism of parabionts showed also significant genetic differences. Chimerism was the highest in the H-B11B11 birds and the lowest in the L-B11B11. All chimeric parabionts showed immunological tolerance and almost all nonchimeric parabionts did not. Immunological competence was measured by splenomegaly index (SI) based on GVHR. The SI value of parabionts changed in the direction of the SI value of the partner. The SI value of chimeric parabionts differed significantly from those of nonchimeric parabionts and controls. Antibody response to bovine serum albumin was tested only for the H-B11B11. Antibody titers of parabionts were significantly lower than those of controls.

Comparison of the major histocompatibility antigen on Marek's disease-derived cell lines detected by membrane fluorescent antibody, cytotoxicity, and lymphoagglutination tests.

The reactivity of 13 antisera specific for the major histocompatibility antigen (MHA) of the B complex against cells in 6 cell lines (MDCC-JP1, JP2, HP1, HP2, RP1, and MSB1), derived from Marek's disease (MD) lymphoma, was compared by using membrane fluorescent antibody, cytotoxicity, and lymphoagglutination tests. The membrane fluorescent antibody test was found to be the most sensitive method for detecting the MHA of the B complex on the MD cell lines. After absorption of these antisera with the homologous erythrocytes, the reactivities of the absorbed sera to the homologous erythrocytes and the MD line cells were reduced or lost altogether. However, after absorbing the antisera reactive for MD cell lines with the respective cell line, the reactivities of the absorbed sera to the line cells were not detected at all when examined by the above tests. However, the reactivities of these absorbed sera to the homologous erythrocytes could still be observed when examined by a hemagglutination test. These results indicate that the antisera contain at least two kinds of antibody (against Class I and Class IV antigens), and the three different tests used here are detecting the Class I antigens of MHA on the MD line cells.

Genetic resistance to a Marek's disease transplantable tumor cell line in chicken lines selected for different immunological characters.

The tumor incidence and mortality of Marek's disease (MD) were determined for 471 progeny from four pairs of lines two-way selected for different immunological characters: graft versus host reaction (GVHR) competence, IgG levels, antibody response to rabbit serum albumin (RSA), and anaphylactic shock to BSA. All chicks were inoculated at 1 wk of age with 1 to 2 x 10(6) viable cells of a MD lymphoblastoid cell line, MDCC-MSB1-41C (41 C). Tumor incidence differed significantly between the high (H) and low (L) lines of each GVHR-, IgG-, RSA-, or BSA-selected group. There were also significant differences in mortality rates between H and L lines of all selected groups. The IgG-H, BSA-H, and RSA-H lines were more resistant than their respective L lines. The H line of the GVHR-selected lines was more susceptible than the L line. Tumor regression could be detected only in the IgG- and RSA-selected lines. No definite correlation could be found between genetic resistance to 41C and genetic resistance to MD virus.

[Evaluation of lead and cadmium levels in milk in environmental contamination in the Vale do Paraíba Region, southeastern Brazil]. / Avaliação dos níveis de chumbo e cádmio em leite em decorrência de contaminação ambiental na região do vale do Paraíba, Sudeste do Brasil.

INTRODUCTION: A factory producing lead ingots, located in Caçapava, caused lead and cadmium contamination of the environment, in the Paraiba Valley region of Southeastern, Brazil, through the discharge of industrial waste and the recycling of batteries. The factory, set in a rural, dairy cattle breeding area, worried sanitary authorities who envisaged the possibility of these metals' having entered the food chain. For the purpose of assessing the levels of contamination of the milk produced in the region, due to the cattle's possible consumption of contaminated grass and water, the amounts of cadmium and lead present in the milk were verified. MATERIAL AND METHOD: Major producers, covering an area of up to 20 km from the contaminated source, authorized collection of 218 samples of both pasteurized and non-pasteurized milk, which were analysed. Lead and cadmium levels were determined by flame atomic absorption spectrophotometry, the lead being pre-concentrated by complexation with APDC (ammonium 1-pyrrolidinecarbodithioate) and further extraction with isobutyl methylketone. RESULTS AND CONCLUSIONS: Of the total number of samples, 43 presented lead levels over the maximum limit of 0.05 mg/kg established by Brazilian legislation. The median value found for lead was 0.04 mg/L. The variance analysis, with 95% confidence level, found no significant difference among the types of milk studied with regard to lead levels. As for cadmium, all samples showed levels below the 0.02 mg/L quantification limit of the method. In spite of the environmental contamination, the levels of cadmium found in the milk were below the 1.0 mg/kg limit established by Brazilian legislation.

[Clinical trial of 111In-antimyosin antibody imaging: (2). Imaging of myocardial infarction and myocarditis].

A new scintigraphic method to detect myocardial necrosis has been developed using antimyosin monoclonal antibody F ab labeled with indium-111 (111In-antimyosin). We investigated 111In-antimyosin scintigraphy in 35 patients with myocardial infarction, 5 patients with myocarditis and 3 patients with angina pectoris. 111In-antimyosin F ab was administered iv and antimyosin images were recorded by planar and single photon emission computed tomography (SPECT) 48-72 hrs after injection. Planar images showed discrete localization of 111In-antimyosin in 26 of 27 patients within 16 days after the onset of acute myocardial infarction in 14 of whom creatine kinase, glutamic oxaloacetic transaminase and lactic dehydrogenase had already normalized. In addition, positive scans were also obtained in 4 of 8 patients 1 to 9 months after the onset of the disease. Three patients with acute myocarditis (two of whom were biopsy-proven) had positive scans 2 and 4 weeks after the onset of the disease. Although mechanism of persistent positive anti-myosin images in the chronic stage remains to be clarified, 111In-antimyosin scintigraphy holds potential promise as a noninvasive method for the detection of myocardial injury.

Homozygous c.14576G>A variant of RNF213 predicts early-onset and severe form of moyamoya disease.

OBJECTIVE: RNF213 was recently reported as a susceptibility gene for moyamoya disease (MMD). Our aim was to clarify the correlation between the RNF213 genotype and MMD phenotype. METHODS: The entire coding region of the RNF213 gene was sequenced in 204 patients with MMD, and corresponding variants were checked in 62 pairs of parents, 13 mothers and 4 fathers of the patients, and 283 normal controls. Clinical information was collected. Genotype-phenotype correlations were statistically analyzed. RESULTS: The c.14576G>A variant was identified in 95.1% of patients with familial MMD, 79.2% of patients with sporadic MMD, and 1.8% of controls, thus confirming its association with MMD, with an odds ratio of 259 and p < 0.001 for either heterozygotes or homozygotes. Homozygous c.14576G>A was observed in 15 patients but not in the controls and unaffected parents. The incidence rate for homozygotes was calculated to be >78%. Homozygotes had a significantly earlier age at onset compared with heterozygotes or wild types (median age at onset 3, 7, and 8 years, respectively). Of homozygotes, 60% were diagnosed with MMD before age 4, and all had infarctions as the first symptom. Infarctions at initial presentation and involvement of posterior cerebral arteries, both known as poor prognostic factors for MMD, were of significantly higher frequency in homozygotes than in heterozygotes and wild types. Variants other than c.14576G>A were not associated with clinical phenotypes. CONCLUSIONS: The homozygous c.14576G>A variant in RNF213 could be a good DNA biomarker for predicting the severe type of MMD, for which early medical/surgical intervention is recommended, and may provide a better monitoring and prevention strategy.