Formation of mu-oxo dimer by the reaction of (meso-tetraphenylporphyrinato)iron(III) with various imidazoles.

Reactions between (meso-tetraphenylporphyrinato)iron(III) perchlorate [Fe(tpp)]ClO4 and various imidazoles have been examined in CD2Cl2 solutions. 1H NMR analysis revealed the formation of three kinds of complex; mu-oxo dimer, mono-imidazole adduct, and bis-imidazole adduct. The product ratios changed to a great extent depending on the amount and nature of imidazoles. In general, addition of less than 1.0 equiv of imidazole relative to [Fe(tpp)]ClO4 led to the formation of both mu-oxo dimer and mono-imidazole adduct. However, by the addition of excess amount of imidazole, either the mu-oxo dimer or bis-imidazole adduct was formed exclusively depending on the bulkiness of the imidazole used. In the case of bulky imidazole such as 2-methylbenzimidazole or 2-isopropyl-1-methylimidazole, the mu-oxo dimer was formed quantitatively. In the case of less bulky imidazole such as parent imidazole or 1-methylimidazole, bis-imidazole adduct became the sole product. The results have been explained in terms of the difference in steric interactions between the axial ligands and porphyrin core; the severe steric repulsion prohibits the formation of bis-adduct in the case of bulky imidazoles. As a result, bulky imidazoles prefer to behave as a base; they abstract a proton from coordinated water, and lead to the formation of mu-oxo dimer. Thus, the role of bulky imidazoles in these reactions has some relevance to that of distal histidine in hemoglobin and peroxidase.

Cytotoxic T lymphocyte antigen 4 gene polymorphism confers susceptibility to type 1 diabetes in Japanese children: analysis of association with HLA genotypes and autoantibodies.

OBJECTIVE: Although the polymorphisms of the cytotoxic T lymphocyte antigen 4 (CTLA4) gene have been shown to be associated with Type 1 diabetes in Caucasians, some conflicting results have been reported among subjects of different ethnic backgrounds. We examined a CTLA4 polymorphism and its relationship to human leucocyte antigen (HLA) genotypes and autoantibodies for glutamic acid decarboxylase 65 (GAD65) and IA-2 in Japanese children with Type 1 diabetes. SUBJECTS AND MEASUREMENTS: The study group consisted of 125 childhood-onset Japanese subjects (50 males, 75 females) with Type 1 diabetes. The CTLA4 A/G polymorphism at position 49 was analysed using a PCR-restriction fragment length polymorphism (PCR-RFLP) method. The HLA-DRB1 and DQB1 genotypes were defined by DNA analysis using PCR-sequence-specific oligonucleotide (PCR-SSO) probes. The GAD65 autoantibody (GAD65Ab) and IA-2 autoantibody (IA-2Ab) titres were measured using radioimmunoassay. RESULTS: The distribution of genotype frequencies differs between subjects with Type 1 diabetes (GG: 46%, AG: 50%, AA: 5%) and controls (GG: 39%, AG: 44%, AA: 17%) (P < 0.01). The frequency of the G allele is higher in the diabetes group than in the controls (P < 0.05). When the subjects were subdivided according to HLA genotype, the two major HLA high-risk groups, with DR9-DQ9 and DR4-DQ4, that are unique to Japanese populations showed no difference in their CTLA4 polymorphism frequencies. Although no association between the CTLA4 polymorphism and the prevalence of GAD65Ab was found, CTLA4 GG subjects that had been newly diagnosed (< 9 months) had significantly higher levels of autoantibodies than AG subjects (P < 0.01). The prevalence and titres of IA-2Ab were not associated with the CTLA4 polymorphism. CONCLUSIONS: The CTLA4 gene might confer a susceptibility to childhood-onset Type 1 diabetes in the Japanese population. The association between this CTLA4 polymorphism and the HLA genotype was similar for both major groups with HLA high-risk alleles. CTLA4 might contribute to the humoral immune response to GAD in newly diagnosed subjects.

Structure and function of 6,7-dicarboxyheme-substituted myoglobin.

Myoglobin was reconstituted with 6,7-dicarboxy-1,2,3,4,5, 8-hexamethylheme, a compact synthetic heme with the shortest acid side chains, to pursue the structural and functional consequences after intensive disruption of the heme propionate-apoglobin linkages in the native protein. The electron-withdrawing carboxylate groups directly attached to the porphyrin ring lowered the oxygen affinity by 3-fold as compared with native myoglobin. Autoxidation of the oxy derivative to the ferric protein proceeded with 1.6 x 10(-)2 min-1 at pH 7.0 and 30 degrees C. The crystallographic structure of the cyanomet myoglobin with 1.9 A resolution shows that the heme adopts a unique orientation in the protein pocket to extend the two carboxylates toward solvent sphere. The native globin fold is conserved, and the conformations of globin side chains are almost intact except for those located nearby the heme 6,7-carboxylates. The 7-carboxylate only weakly interacts with Ser92 and His97 through two mediating water molecules. The 6-carboxylate, on the other hand, forms a novel salt bridge with Arg45 owing to conformational flexibility of the guanidinium side chain. The proton NMR shows that the small heme does not fluctuate about the iron-histidine bond even at 55 degreesC, suggesting that the salt bridge between Arg45 and heme 6-carboxylate is of critical importance to recognize and fix the heme in myoglobin.

Ganglioneuroma of left adrenal gland in a patient with Turner syndrome during growth hormone therapy.

We report on a Japanese girl with Turner syndrome (45,XO) who developed ganglioneuroma of the left adrenal gland during growth hormone (GH) therapy. She had received GH replacement therapy from the age of 6.8 years. At the age of 10.3 years, abdominal ultrasonography revealed a mass which occupied the upper area of her left kidney. Computed tomography and magnetic resonance imaging of the abdomen showed a low density mass with a smooth surface located between the upper portion of the left renal vein and the pancreas. Microscopic examination resulted in a diagnosis of ganglioneuroma of the left adrenal gland. At present we cannot conclude that patients who have received GH replacement therapy are at higher risk for developing tumors compared to those without GH replacement therapy.

[A quantitative analysis of the cells infected with Epstein-Barr virus in the peripheral blood mononuclear cells derived from the patients with infectious mononucleosis].

The quantitative analysis of the cells infected with Epstein-Barr virus was performed on the peripheral blood mononuclear cells from the patients with infectious mononucleosis, by using in situ hybridization with Epstein-Barr virus encoded small nuclear RNA1 (EBER1). An alkaline-phosphatase conjugated oligonucleotide probe complementary to EBER1 was used as an antisense probe, while oligonucleotide DNA probe compatible with the sequence of EBER1 was used as a sense probe, control probe. The EBER1 positive cells on the slide-glass were enumerated microscopically. In situ hybridization revealed that 50,000 peripheral blood mononuclear cells from the patients in the acute phase of infectious mononucleosis contained 35 +/- 36 cells infected with Epstein-Barr virus (n = 11). The cells infected with Epstein-Barr virus apparently decreased in the convalescence of all the patients with infectious mononucleosis and the mean of the cells infected with Epstein-Barr virus was 3 +/- 4 in the convalescence (n = 6) (p < 0.02). On the other hand, no positive cells were detected in healthy individuals with past-infection of Epstein-Barr virus (n = 10) or without any previous Epstein-Barr virus infection (n = 11). The striking increase of the cells with Epstein-Barr virus genome was clearly demonstrated in the peripheral blood mononuclear cells from the patients with infectious mononucleosis.

Prediction of efficacy of antidepressant by 1-week test therapy in depression.

Forty patients suffering from depression were treated with fixed doses of clomipramine for 4 weeks. The most effective predictor of efficacy during the 4 weeks was the percentage improvement in the Hamilton rating scale for depression during the first week. Baseline severity did not show any predictive ability. The Beck depression rating scale had only low predictive ability, and the dexamethasone suppression test did not predict the outcome at all. These results suggest the benefit of 1-week test therapy for predicting the efficacy of antidepressants.

Double-blind study of the chronopharmacotherapy of depression.

To investigate the possibility of chronotherapy with antidepressants for patients with depression, we gave single daily doses of clomipramine 150 mg/day to 30 patients with depression at three different times of day, i.e., morning, noon, or before bedtime, using a double-blind method over a 4-week period. Beneficial effects differed according to the administration time of day, with the most effective result being found for the administration at noon. Time-dependent differences in side effects were observed in tremors and dryness of mouth. An additional 10 patients were administered their medications three times a day by traditional, equally divided doses, and the efficacy was inferior than daily single doses at noon. The study results showed the significance of the administration time of day for the benefits and side effects of antidepressant therapy for depression.