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Soy isoflavones have been shown to have anti-inflammatory properties; however, the anti-inflammatory effects of isoflavone metabolites produced during soybean germination remain unclear. We found that the daidzein and genistein derivatives, 8-prenyl daidzein (8-PD) and 8-prenyl genistein (8-PG), demonstrated a more potent effect than daidzein and genistein on repressing inflammatory responses in macrophages. Although IkB protein levels were unaltered, 8-PD and 8-PG repressed nuclear factor kappa B (NF-κB) activation, which was associated with reduced ERK1/2, JNK, and p38 MAPK activation and suppressed mitogen- and stress-activated kinase 1 phosphorylation. Inflammatory responses induced by the medium containing hypertrophic adipocyte secretions were successfully suppressed by 8-PD and 8-PG treatment. In the ex vivo study, 8-PD and 8-PG significantly inhibited proinflammatory C-C motif chemokine ligand 2 (CCL2) secretion from the adipose tissues of mice fed a long-term high-fat diet. The data suggest that 8-PD and 8-PG could regulate macrophage activation under obesity conditions., (© The Author(s) 2023. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)

Protein intake potently increases body temperature and energy expenditure, but the underlying mechanism thereof remains incompletely understood. Simultaneously, protein intake potently stimulates glucagon-like peptide-1 (GLP-1) secretion. Here, we examined the involvement of GLP-1 in the thermic effects of dietary proteins in rodents by measuring rectal temperature and energy expenditure and modulating GLP-1 signaling. Rectal temperature of rats or mice fasted for 4 or 5 hours were measured using a thermocouple thermometer before and after an oral administration of nutrients. Oxygen consumption after oral protein administration was also measured in rats. Rectal temperature measurements in rats confirmed an increase in core body temperature after refeeding, and the thermic effect of the oral administration of protein was greater than that of a representative carbohydrate or lipid. Among the five dietary proteins examined (casein, whey, rice, egg, and soy), soy protein had the highest thermic effect. The thermic effect of soy protein was also demonstrated by increased oxygen consumption. Studies using a nonselective β-adrenergic receptor antagonist and thermal camera suggested that brown adipose tissue did not contribute to soy protein-induced increase in rectal temperature. Furthermore, the thermic effect of soy protein was completely abolished by antagonism and knockout of the GLP-1 receptor, yet potentiated via augmentation of intact GLP-1 levels through inhibition of dipeptidyl peptidase-4 activity. These results indicate that GLP-1 signaling is essential for the thermic effects of dietary proteins in rats and mice, and extend the metabolic actions of GLP-1 ensuing from nutrient ingestion to encompass the thermic response to ingested protein., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Oryza sativa L. ssp. japonica cv. Nipponbare produces a nonproteinogenic amino acid (3R)-β-tyrosine from l-tyrosine by tyrosine aminomutase (OsTAM1). However, physiological and ecological function(s) of β-tyrosine have remained obscure. Often an improved understanding of metabolite localization and transport can aid in design of experiments to test physiological functions. In the current study, we investigated the distribution pattern of β-tyrosine in rice seedlings and found that β-tyrosine is most abundant in the youngest leaves. Based upon observations of high TAM1 activity in mature leaves, we hypothesized that β-tyrosine is transported from mature leaves to young leaves. Patterns of predominant mature synthesis and young leaf accumulation were supported by stable isotope studies using labeled β-tyrosine and the removal of mature leaves. Stem exudate analyses was also consistent with β-tyrosine transport through phloem. Thus, we identify young leaves as a key target in efforts to understand the biological function(s) of β-tyrosine in rice., (© The Author(s) 2022. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)

Isoflavonoids are commonly found in leguminous plants. Glycitein is one of the isoflavones produced by soybean. The genes encoding the enzymes in the isoflavone biosynthetic pathway have mostly been identified and characterized. However, the gene(s) for isoflavone O-methyltransferase (IOMT), which catalyzes the last step of glycitein biosynthesis, has not yet been identified. In this study, we conducted multi-omics analyses of fungal-inoculated soybean and indicated that glycitein biosynthesis was induced in response to biotic stress. Moreover, we identified a unique type of IOMT, which participates in glycitein biosynthesis. Soybean seedlings were inoculated with Aspergillus oryzae or Rhizopus oligosporus and sampled daily for 8 d. Multi-omics analyses were conducted using liquid chromatography-tandem mass spectrometry and RNA sequencing. Metabolome analysis revealed that glycitein derivatives increased following fungal inoculation. Transcriptome co-expression analysis identified two candidate IOMTs that were co-expressed with the gene encoding flavonoid 6-hydroxylase (F6H), the key enzyme in glycitein biosynthesis. The enzymatic assay of the two IOMTs using respective recombinant proteins showed that one IOMT, named as GmIOMT1, produced glycitein. Unlike other IOMTs, GmIOMT1 belongs to the cation-dependent OMT family and exhibited the highest activity with Zn2+ among cations tested. Moreover, we demonstrated that GmIOMT1 overexpression increased the levels of glycitein derivatives in soybean hairy roots when F6H was co-expressed. These results strongly suggest that GmIOMT1 participates in inducing glycitein biosynthesis in response to biotic stress., (© The Author(s) 2020. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Objective: Patients with adeno/adenosquamous carcinoma may have a poorer prognosis than patients with squamous cell carcinoma. Radiotherapy and concurrent chemoradiotherapy are used as adjuvant therapies for cervical cancer, regardless of the histological subtype. The aim of this study was to investigate the prognostic outcome of adjuvant therapy for patients with adeno/adenosquamous carcinoma with pathological risk factors., Methods: The medical records of 135 patients with stage IB-IIB cervical cancer with squamous cell carcinoma or adeno/adenosquamous carcinoma who underwent primary surgery followed by adjuvant therapy were retrospectively reviewed. Patients with a pathologically confirmed bulky tumor (≥4 cm), nodal metastasis and/or parametrium invasion were included in the study., Results: The median follow-up period was 48 (1-132) months. Of the 135 patients, 90 with squamous cell carcinoma and 23 with adeno/adenosquamous carcinoma were treated with adjuvant radiotherapy and concurrent chemoradiotherapy (SCC-RT/CCRT and AC-RT/CCRT groups), and 22 with adeno/adenosquamous carcinoma were treated with adjuvant systemic chemotherapy (AC-CT group). There were no significant differences in clinicopathological factors between the SCC-RT/CCRT and AC-RT/CCRT groups and between the AC-RT/CCRT and AC-CT groups. Progression-free survival was significantly shorter in the AC-RT/CCRT group compared to the SCC-RT/CCRT group (P = 0.002). Adeno/adenosquamous carcinoma histology and multiple lymph node metastasis were independent prognostic factors for shorter progression-free survival in patients treated with adjuvant radiotherapy and concurrent chemoradiotherapy. Progression-free survival was also significantly shorter in the AC-RT/CCRT group compared to the AC-CT group (P = 0.026)., Conclusions: Adjuvant radiotherapy and concurrent chemoradiotherapy may be less effective for patients with adeno/adenosquamous carcinoma than for those with squamous cell carcinoma. Adjuvant systemic chemotherapy may be beneficial for adeno/adenosquamous carcinoma and further studies are warranted., (© The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: This Phase II study was carried out to investigate the efficacy, safety and pharmacokinetics of topotecan in Japanese patients with relapsed ovarian carcinoma., Methods: Patients with relapsed ovarian carcinoma after having received one regimen containing platinum-based chemotherapy were eligible for this study. Topotecan was administered at 1.2 mg/m(2)/day for five consecutive days, repeated every 3 weeks., Results: Seventy-two patients were enrolled in the study. The response rate was 28.2% (95% confidence interval, 18.1-40.1%). Signs of myelosuppression, such as neutropenia (Grade 3, 12.5%; Grade 4, 83.3%), thrombocytopenia (Grade 3, 36.2%; Grade 4, 4.2%) and decreased hemoglobin (Grade 3, 36.1%; Grade 4, 11.1%), were the most common hematological toxicities. Grade 3 febrile neutropenia occurred in 5 (6.9%) patients. There was little intraindividual or interindividual variability in the pharmacokinetics of topotecan., Conclusions: Topotecan at 1.2 mg/m(2)/day is an effective and tolerable therapeutic option for Japanese patients with relapsed ovarian carcinoma.

In this study, we examined the role of nitric oxide (NO) in controlling vascular integrity mediated by vascular endothelial (VE)-cadherin in chronic inflammation. Periapical granulomas were analysed for the expression of inducible NO synthase (iNOS) and VE-cadherin, and more iNOS expression than VE-cadherin was shown. Human umbilical vein endothelial cells (HUVECs) were stimulated with proinflammatory cytokines and lipopolysaccharide extracted from Porphyromonas gingivalis and it induced iNOS expression, whereas it reduced VE-cadherin expression, compared with negative controls. On the other hand, pre-incubation with 1400W, an iNOS-specific inhibitor, markedly reduced iNOS expression in stimulated HUVECs and restored VE-cadherin expression to its control level, suggesting that vascular integrity was modulated in conjunction with the reduction of NO. Immunocytochemistry confirmed the functional role of NO in cultured HUVEC monolayers with or without 1400W. These data are consistent with a hypothesis suggesting that NO could attenuate VE-cadherin-mediated vascular integrity in human chronic inflammation.

Objective: This study was conducted to evaluate the efficacy and safety of pegylated liposomal doxorubicin (PLD) in Japanese patients with Müllerian carcinoma having a therapeutic history of platinum-based chemotherapy., Methods: Patients who were diagnosed with Müllerian carcinoma (epithelial ovarian carcinoma, primary carcinoma of fallopian tube and peritoneal carcinoma) by histological examination and had received the initial platinum-based chemotherapy were included in the study. The study drug was administered to the patients at 50 mg/m(2) every 4 weeks., Results: Seventy-four patients were enrolled in the study. All patients had received platinum-based chemotherapy as first-line regimen and more than 90% of patients had also received taxanes. The overall response rate was 21.9% (95% confidence interval, 13.1-33.1%) and 38.4% of patients had stable disease. The median time to progression was 166 days. The major non-haematological toxicities were hand-foot syndrome (Grade 3; 16.2%) and stomatitis (Grade 3; 8.1%). Myelosuppression such as leukopenia (Grade 3; 52.7%, Grade 4; 6.8%), neutropenia (Grade 3; 31.1%, Grade 4; 36.5%) and decreased haemoglobin (Grade 3; 14.9%, Grade 4; 2.7%) were the most common haematological toxicities., Conclusion: We confirmed that a 50 mg/m(2) every 4 weeks regimen of PLD was active in Japanese patients with Müllerian carcinoma having a therapeutic history of platinum-based chemotherapy and toxicity was manageable by dose modification of PLD or supportive care.

Upon antigen stimulation, B lymphoid cells undergo terminal differentiation into antibody-secreting plasma cells. This process accompanies drastic changes in cell functions such as a loss of B-cell identity, induction of secretory apparatus, and an extremely increased transcription of antibody genes. These changes are the result of re-wiring of a transcription factor network in B and plasma cells. While the transcription repressor Blimp-1 induces plasma cell differentiation, another repressor Bach2 has emerged as a negative regulator of Blimp-1 in B cells. These two transcription factors, together with other several factors, appear to constitute a main transcriptional regulatory network for the terminal differentiation process of plasma cells from B cells.

There are several vacant channels for diagnosis, RF heating and so on through the shielding structure in fusion reactors. Some of them consist of dogleg ducts, through which neutrons stream in a complex manner. An experiment was conducted with the Fusion Neutronics Source (FNS) facility at JAERI to study the behaviour of neutrons in the duct and assess the reliability of calculation methods for the design of fusion reactors such as ITER. The assembly was an iron slab 180 cm in thickness with a doubly bent duct 30 x 30 cm2 in cross section. The experiment was analysed using a simple design code for radiation streaming, DUCT-III, and the Monte Carlo code MCNP. The results indicate that the simple design code is reliable enough to be used for shielding design analyses as well as the Monte Carlo method, which showed excellent agreement between calculated and measured values.

In this study spatial and time distribution of neutrons leaking from Hokkaido University 45 MeV Electron Linac facility have measured and compared with the Monte Carlo simulations. The neutron transport processes inside and outside the facility building has been simulated using MCNP. The neutrons have measured by BF3 counters and 3He counters with polyethylene moderators up to the distance of 330 m from the facility. The spatial distribution of ambient dose equivalent converted from the counts has been compared with the simulation. The distribution estimated from the counts by the BF3 counter has been shown fairly good agreement with the calculation. The spatial distribution of counts obtained at the 45 MeV Electron Linac facility has been compared with that obtained at the Fusion Neutronics Source (FNS) facility of JAERI. The difference between the propagation characteristics of neutrons leaking from those facilities has been discussed.

Background: Remnant lipoprotein-cholesterol (RLP-C) concentrations in sera of healthy individuals are very low (0.080-0.437 mmol/L), making conventional cholesterol methods poorly suited to this purpose. We have developed a highly sensitive cholesterol assay (CD method) and applied it to the RLP-C assay., Methods: The CD shuttled cholesterol reversibly between reduced and oxidized forms in the presence of thio-NAD and NADH. The production rate of thio-NADH correlated with the cholesterol concentration and was measured by the absorbance at 404/500 nm. This CD method was combined with an immunoaffinity separation procedure with specific monoclonal antibodies to apolipoprotein (apo) A1 and apo B-100 and used for RLP-C assay. Results were compared with a RLP-C method that uses cholesterol oxidase, peroxidase, and chromogenic substrate., Results: The CD method could detect 0.10 x 10(-3) mmol/L cholesterol and was at least 5 times more sensitive than the conventional enzymatic method. Within- and between-day imprecision (as CVs) of the RLP-C assay with the CD method was <4%. Regression analysis of RLP-C assays with the new (y) and conventional (x) cholesterol methods yielded: y = 1.02x - 0.008 mmol/L (S(y/x) = 0.0065 mmol/L; r = 0.997; n = 297)., Conclusions: Serum RLP-C can be measured by the CD method. The CD method may be useful for other assays that require sensitive cholesterol measurements in biological materials.

An ATPase called EA4 seems to measure time as a diapause-duration timer in the seasonal cycle of the silkworm, Bombyx mori. A peptide named PIN seems to regulate the time measurement of EA4. We characterize the EA4 as the first step to analyse its interaction with PIN. Matrix-assisted laser desorption/ionization-time of flight-mass spectrometry shows EA4 forms an equimolar complex with PIN. The binding affinity of EA4 for PIN is about 460 nM, as measured by surface plasmon resonance. Western blot analysis of EA4 with a variety of biotinylated lectins suggests that EA4 is a glycoprotein containing N-linked oligosaccharide. On enzymatic cleavage of the glycosyl chain, the carbohydrate is revealed to be essential for the regulation of EA4-time measurement through the interaction with PIN. PIN holds the timer by binding to EA4, and the dissociation of the complex could constitute the cue for the time measurement.

The purpose of this study was to determine whether the JAK pathway is involved in eosinophil activation and survival through IFN-gamma receptor signalling in human peripheral eosinophils. Eosinophils were purified from the blood of six atopic disease patients by anti-CD16 magnetic bead-negative selection. IFN-gamma significantly up-regulated survival and CD69 expression in 24-48 h cultured eosinophils. Further, IFN-gamma induced tyrosine phosphorylation of JAK2 in eosinophils, as indicated by Western blot analysis. Finally, the specific JAK2 inhibitor AG-490 inhibited the tyrosine phosphorylation of JAK2, IFN-gamma-induced survival and CD69 expression in eosinophils. In conclusion, these results indicate that IFN-gamma induces eosinophil survival and CD69 expression through the activation of JAK2 in peripheral eosinophils, suggesting that JAK2 may play a significant role in eosinophil regulation by IFN-gamma-IFN-gammaR interaction.

To search for compounds that reverse the drug resistance induced by glutathione (GSH), an original screening system to detect intracellular GSH depleters was established. Among 8843 microbes derived from the soil samples tested, the extracts of two Streptomyces species named KS6701 and KS8846, lowered the intracellular GSH level of Saccharomyces cerevisiae 5 x 47. From both the microbes, 5-hydroxy-4-oxo-L-norvaline (HON) was isolated as the active compound. At a concentration of 50-100 micrograms/ml, HON also decreased the GSH/protein level of the human ovarian tumor cell line, 2008/C13*5.25 and reversed its resistance to cisplatin. We also investigated the mechanism of the depletion. HON had little effect on gamma-glutamylcysteine synthetase (gamma-GCS) or glutathione synthetase, but HON decreased the quantity of thiol substances when it was spontaneously reacted with them. This suggested that the GSH depletion by HON occurred through a mechanism different from that of buthionine sulfoximine, a selective gamma-GCS inhibitor.

In order to determine whether or not IFN-gammaR is associated with regulatory mechanisms on human eosinophil function, we examined the expression of functional IFN-gammaR on human peripheral eosinophils. In this study, peripheral blood eosinophils were obtained from seven normal controls and 12 patients (bronchial asthma, n = 9, and hypereosinophilic syndrome (HES), n = 3), and the purity of eosinophils was 97.11 +/- 2.31%, n = 19. We first showed that anti-IFN-gammaR alpha-chain MoAb reacted with all tested eosinophils of both normal controls and patients by flow cytometry analysis. We also showed expression of mRNA for the alpha-chain of IFN-gammaR in all purified eosinophils of six individuals. Further, to characterize IFN-gammaR on eosinophils, we did binding experiments with 125I-IFN-gamma on purified peripheral eosinophils. The linear Scatchard plot indicated a single type of high-affinity binding sites (dissociation constant (Kd) = 3.89-4.95 x 10(-10) M, numbers of binding sites = 183-233/cell, n = 3). To determine whether IFN-gammaR on eosinophils is functional, we examined surface eosinophilic cationic protein (ECP) and CD69 induction after IFN-gammaR ligation with recombinant human IFN-gamma (rhIFN-gamma) on eosinophils by flow cytometry. rhIFN-gamma stimulation significantly induced both ECP and CD69 expression on the 2-18 h-cultured eosinophils in a dose-dependent manner. Further, the effects of rhIFN-gamma stimulation were significantly blocked by both a neutralizing anti-IFN-gamma MoAb and a blocking anti-IFN-gammaR MoAb. These results suggest that human peripheral eosinophils express functional IFN-gammaR.

We isolated a bacterial strain with an enzyme which releases dipeptide from Gly-Arg-p-nitroanilide. The bacterium was tentatively identified as Aureobacterium sp. The enzyme, named AuDAP, was purified and characterized. It was homogenous by SDS-PAGE and IEF, and had a molecular mass of 90,000 Da by SDS-PAGE and 88,000 Da by gel filtration, so it may be a monomer. The isoelectric point was 3.8 and the optimum pH was 10.0. The purified enzyme hydrolyzed Gly-Arg-pNA, a model substrate for DAP I, and Arg-Arg-MNA, a model substrate for DAP III. However, this enzyme did not hydrolyze Gly-Phe pNA, also a model substrate for DAP I. These results suggested that this enzyme did not fall under the classification of mammalian DAPs and was similar to DAP BI from Pseudomonas sp. WO24 and dDAP from Dictyostelium discoideum, although several differences were observed between them. The N-terminal amino acid sequence of this enzyme showed no significant homology to any enzyme and protein, except only for DAP BI.

In order to determine regulatory mechanisms of eosinophil apoptosis, we examined the effect of recombinant IL-5 and interferon-gamma (IFN-gamma) on eosinophil apoptosis and bcl-2 expression. rhIL-5 (2.5 ng/ml) significantly inhibited eosinophil apoptosis in 96 h in vitro culture compared with medium only-cultured eosinophils (89.4 +/- 3.6% versus 31.3 +/- 12.2% (mean +/- s.d.); n = 7, P < 0.05). Further, rhIL-5 significantly increased bcl-2 protein and mRNA expression on cultured eosinophils. A phosphorothioate antisense oligonucleotide targeted at the ATG translation initiation codon of bcl-2 (10(-5) M) could significantly block the supportive effect of rhIL-5 (0.25 ng/ml) for eosinophil survival compared with sense cDNA of bcl-2 on 96 h culture (inhibition rate 28.01 +/- 4.56% versus 0.07 +/- 1.73%; n = 4, P < 0.05). In contrast, rhIFN-gamma (100 U/ml) significantly inhibited eosinophil apoptosis on 96 h in vitro culture (72.7 +/- 10.5%; n = 7, P < 0.05), but did not significantly up-regulate bcl-2 protein and mRNA. These results indicate that IL-5 has inhibitory effects on eosinophil apoptosis by regulation of bcl-2 expression.

We previously established murine hybridomas producing a monoclonal antibody monospecific against three glucosyltransferases (I, SI and S) of Streptococcus mutans which contribute to dental caries formation. Here, we developed a new immunochemical technique (cross-dot system) with which individual levels of glucosyltransferases expressed by S. mutans can be evaluated. We also examined glucosyltransferase production and in vitro artificial plaque formation by a reference strain and several clinical isolates of S. mutans. The findings indicate that the levels of glucosyltransferases produced greatly vary with the cells and the culture medium, and that the cells producing high levels of both glucosyltransferase-SI and glucosyltransferase-I enzymes may possess high in vitro artificial plaque forming ability. We suggest that the cross-dot system will be useful for estimating the cariogenic potential of S. mutans isolates.

We investigated the usefulness of gadolinium-enhanced magnetic resonance (Gd-MR) imaging for diagnosis of right ventricular (RV) infarction in 14 patients with acute inferior infarction. Myocardial perfusion images with thallium 201 and technetium 99m pyrophosphate (dual single-photon emission computed tomography: dual SPECT) and haemodynamic data were obtained on admission. Patients were classified into three groups based on dual SPECT and Gd-MR findings: no accumulation in the right ventricle (negative, Type 1), posterior RV infarction (Type 2) and anterior and posterior RV infarction (Type 3). No patients exhibited the haemodynamic criteria for RV infarction (a mean right atrial pressure above 10 mmHg and a ratio of mean right atrial pressure to pulmonary artery wedge pressure above 0.8). Dual SPECT identified RV infarction in eight patients (three Type 2 and five Type 3, 57% of the total). Gd-MR imaging also identified eight patients (57%) as positive (five Type 2 and three Type 3). Our results showed that Gd-MR imaging was not only useful for diagnosis of RV infarction, but also equal to dual SPECT in sensitivity and specificity, and superior to it as regards acquisition time and assessing the spatial anatomy of heart. In conclusion, Gd-MR imaging is superior to dual SPECT for detection of RV infarction.

Ribosomal proteins from 22 strains of 15 different species belong to the genus Arthrobacter were analyzed by an improved two-dimensional gel electrophoresis. Electrophoretograms of ribosomal proteins from 15 type strains had species-specific patterns. Similarity coefficients (SAB values) of ribosomal proteins with mol. wt. of greater than about 20,000, among strains of the same species (DNA relatedness values of more than 61%) were greater than 0.85, but the SAB values among strains of different species were less than 0.60. The N-terminal amino acid sequences of the AL2 proteins, which migrated into similar positions in this method, from 5 type strains were shown to be highly homologous. Our results indicated that ribosomal proteins have been conserved within species during evolution and that the members of the genus Arthrobacter are phylogenetically homogeneous. Thus, ribosomal protein profiles by this method are a potential tool for strain identification.

IL-4 and IFN-gamma positively and negatively regulate allergic inflammation. To determine the regulatory mechanisms of eosinopoiesis by cytokines, we examined the effect of recombinant IL-4 and IFN-gamma and of anti-IL-4 and anti-IFN-gamma antibodies on IL-3- and IL-5-induced eosinophil differentiation from human umbilical cord blood mononuclear cells. rhIL-4 (10-300 U/ml) inhibited IL-3- and IL-5-induced eosinophil differentiation from cord blood mononuclear cells on day 28 of culture by 62-81% in a concentration-dependent manner. rhIFN-gamma (5-500 U/ml) also inhibited IL-3- and IL-5-induced eosinophil differentiation by 80-99% in a concentration-dependent manner. The inhibitory effect of rhIL-4 and rhIFN-gamma was observed only when rhIL-4 or rhIFN-gamma were present in the culture from day 0 to day 14, but not from day 15 to day 28. Addition of anti-IL-4 antibody to the culture enhanced IL-3- and IL-5-induced eosinophil differentiation on day 28 of culture by 30%, whereas anti-IL-2 MoAb and anti-IFN-gamma MoAb had no significant effect. These results indicate that IL-4 and IFN-gamma have inhibitory effects on IL-3- and IL-5-induced eosinophil differentiation from its progenitor cells.

Sugar chains were liberated from mouse tissues (liver, heart, spleen, kidney, thymus, cerebrum, cerebellum, and brain stem) by hydrazinolysis. After acetylation of the free amino groups, the sugar chains released were pyridylaminated. Pyridylamino (PA-) derivatives of the sugar chains from each tissue were fractionated by anion-exchange high-performance liquid chromatography (HPLC) according to their negative charges. Fractions containing PA-neutral sugar chains thus obtained were separated successively by size-fractionation HPLC and reversed-phase HPLC. Two sugar chains that were more abundant in neural tissues than in other tissues were purified. The structures of the two sugar chains were determined by sugar composition analysis, sequential exoglycosidase digestion, and methylation analysis. The proposed structures are shown below. A structure with n = 1 was found in the cerebrum, cerebellum, and brain stem. A structure with n = 0 was abundant in the cerebrum and brain stem, but less so in the cerebellum. [formula: see text]

To investigate cell surface antigens of activated human eosinophils using monoclonal antibodies, we established a murine anti-human eosinophil monoclonal antibody AE500 by immunizing with blood eosinophils from patients with idiopathic hypereosinophilic syndrome (HES) and characterized the reactivity to a variety of human leucocytes by a fluorescence-activated cell sorter. AE500 reacted with blood eosinophils and neutrophils in nine out of 11 patients with marked eosinophilia (greater than or equal to 2500/microliters) (seven with idiopathic eosinophilia including HES and two with asthma), but not with those in asthmatic patients with mild eosinophilia (n = 10) or in healthy subjects (n = 8). AE500 did not react with blood lymphocytes, monocytes or platelets. AE500 did not react with human myeloid or lymphoid cell lines, including eosinophilic leukemia cell lines EOL-1 and EOL-3. The reactivity of AE500 to blood eosinophils and neutrophils in patients with marked eosinophilia changed in relation to blood eosinophil counts and prednisolone therapy. In addition, the reactivity of AE500 to blood eosinophils was increased in three out of four AE500-positive eosinophils by the incubation of the cells with granulocyte-macrophage colony-stimulating factor (GM-CSF) at 37 degrees C for 30 min, but not with interleukin 3 or interleukin-5. These results suggest that the anti-eosinophil antibody AE500 detects a cell surface antigen expressed on blood granulocytes in a hypereosinophilic state. This anti-eosinophil antibody would be useful for analysing the mechanism of eosinophilia.

We investigated the distribution of neutral endopeptidase (NEP; EC 3.4.24.11) activity, a possible regulatory enzyme for neuropeptide-induced leukocyte activations, in each cell type of human blood leukocytes. The NEP activity assessed by an NEP inhibitor phosphoramidon-sensitive Met5-enkephalin degrading activity was present in neutrophils and the common acute lymphoblastic leukemia antigen (CALLA)-positive leukemic cells (59 pmol/min/10(6) cells and 62 pmol/min/10(6) cells, respectively); however, the NEP activity was virtually absent in lymphocytes, monocytes, eosinophils, basophils, CALLA-negative leukemic cells, or a promyelocytic cell line HL-60. The enzymatic activity was characterized as NEP on the basis of the values of kinetic parameters (Km = 61 microM, Kcat = 1,692 min-1, and Kcat/Km = 28 min-1 microM-1) and the values of IC50 of two NEP inhibitors phosphoramidon and thiorphan (7.4 nM and 8.4 nM, respectively). The distribution of NEP detected immunocytochemically using anti-NEP monoclonal antibodies was also found to be parallel with the distribution of NEP activity among peripheral blood leukocytes.

This assay allows simultaneous determination of the enantiomers of both disopyramide and its active metabolite, mono-N-dealkyldisopyramide, in 1 mL of plasma or 0.1 mL of urine within approximately 35 min by HPLC with a chiral cellulose-derivative column and ultraviolet detection. Recoveries for the analytes and the internal standard (racemic verapamil) with an extraction from alkalinized plasma or urine into diethyl ether were greater than 90%. Intra- and interassay CVs for disopyramide enantiomers were less than 5.5% at 2.5 mg/L in plasma and less than 6.5% at 25 mg/L in urine; for mono-N-dealkyldisopyramide enantiomers they were less than 6.3% and less than 8.9%, respectively. Intra- and interassay relative errors for determining these analytes in plasma and urine at 2.5 and 25 mg/L, respectively, ranged from -5.9% to +2.5%. The calibration curves for the respective analytes were linear (r = 0.995 or greater, P less than 0.01) from 0.025 to 5.0 mg/L in plasma and from 0.5 to 10 mg/L in urine. The lower detection limits (signal-to-noise ratio of 3) for S(+)-disopyramide and the other analytes were 0.010 and 0.025 mg/L, respectively. We evaluated clinical applicability of this method by determining steady-state plasma concentrations and urinary excretions of the respective analytes in a pediatric patient being treated with racemic disopyramide.

Adult rats were hypophysectomized and their pituitary glands autotransplanted on day 2 of an estrous cycle (day 1, ovulation); the rats were either hysterectomized or sham hysterectomized immediately before this operation. On day 21, a few days after the beginning of the 3+ month period of regression of progesterone secretion, they were injected sc with 500 micrograms prostaglandin F2 alpha (PGF2 alpha) either on day 21 alone, on days 21 and 22, on days 21, 22, and 23, or on days 21 and 23; controls were injected with saline on days 21, 22, and 23. In the latter the serum progesterone level slowly fell, as it does in untreated rats subjected to pituitary autotransplantation on day 2. PGF2 alpha treatment induced a rapid, drastic fall in the progesterone level, but in each group except the one injected on days 21 and 23, the level returned to, or close to, the control level in about 10 days. Even in the group injected on days 21 and 23, the pattern of serum progesterone resembled the others although the values were much lower. In a similar group of rats subjected to pituitary autotransplantation on day 2 (all of which were also hysterectomized on day 2), a single injection of 500 micrograms PGF2 alpha (which again induced the same changes in serum progesterone as in the first experiment) was combined with either a 6-h or 4-h withdrawal of PRL; this was achieved by injecting 2 bromo-alpha-ergocryptine (CB-154) at zero time to lower PRL secretion, and beef PRL, in a delaying vehicle, 6 or 4 h later, followed by a second dose of PRL 18 h after the first. When combined with such a brief period of PRL withdrawal, the PGF2 alpha treatment induced a very rapid, drastic and permanent fall in serum progesterone, equal to that induced by CB-154 alone, or by removing the pituitary transplant on day 21. The brief periods of PRL withdrawal, by themselves had very little (6 h) or no (4 h) effect on the progesterone level, and treatment with PRL alone was also without effect. Treatment with CB-154, PGF2 alpha, and PRL simultaneously (i.e. without a period of PRL withdrawal) had the same effect as treatment with PGF2 alpha alone.(ABSTRACT TRUNCATED AT 400 WORDS)

In rats bearing a full complement of conceptuses (FC-bearing rats) progesterone secretion increases about 1.5 times and the corpora lutea double in weight between days 12 and 16. Reducing the number of conceptuses to one (by aspirating all but the single conceptus) at any time between days 7 and 12 of pregnancy (day 1 = insemination) caused progesterone secretion and corpus luteum weight to remain at or below the day 12 level until term. When rats bearing one conceptus were hypophysectomized on day 12, however, serum progesterone increased and fell after day 12 in a pattern almost identical to that in FC-bearing rats, except that the peak and trough occurred about 2 days later; hypophysectomy on day 15 or 18 also increased serum progesterone to values approaching those in intact FC-bearing rats. In a series of rats bearing experimentally determined conceptus numbers between 0 and 5 or naturally occurring ones of 6-16, the serum level of placental lactogen on day 12 (the day of the first peak of secretion of this hormone) was directly proportional to conceptus number over the entire range. The pattern of rat placental lactogen secretion between days 7 and 23 in FC-bearing rats included two peaks, on day 12 and on days 18-21, but in one-conceptus-bearing rats, the level was only slightly higher than the PRL level, and it remained unchanged, even by hypophysectomy, until term. Serum PRL levels bore no relation to conceptus number, were very low, and were unaffected by hypophysectomy except in the FC-bearing rats, in which lypophysectomy prevented the rise that normally occurs after day 21. These results, especially in relation to others previously reported, show that by day 12 of pregnancy, the rat pituitary, instead of merely becoming luteotropically inactive, produces a substance that inhibits further development of the corpora lutea. The results also imply that the placentas maintain luteal growth and progesterone secretion as much by suppressing this luteolytic activity as by stimulating the corpus luteum directly.

Cyclic rats were hypophysectomized and their pituitaries transplanted beneath the kidney capsule on day 2 (day 1, ovulation); they were then either hysterectomized or sham hysterectomized (uterus intact rats) and injected sc daily with either 1.0 micrograms 17 beta-estradiol (E1) or with the sesame oil vehicle until autopsy on day 105. Blood samples were drawn by jugular venipuncture every 2 to 3 days until day 21 and once weekly thereafter. In these rats the serum progesterone level rose until about day 8 and tended to reach a plateau between days 8 and 14. The regression phase began, in general, after days 8 to 14 and continued, without change in rate, for at least 3 months. In both the uterus-intact and the hysterectomized rats, E1 seemed to induce an earlier onset of regression and a brief increase in its rate, but did not otherwise affect the rate of regression. In the hysterectomized rats regression began later than in the uterus-intact rats, but its rate was also not different from the latter. The serum progesterone pattern of rats subjected to pituitary autotransplantation on day 2 was compared with that of pseudopregnant rats which were either subjected to pituitary autotransplantation on day 2, 5, 7, or 9, or were given a pituitary homotransplant on day 2, and then hypophysectomized on day 2, 3, 5, or 7. A group of rats with sham operations served as additional controls. In these rats, the same pattern of serum progesterone already described was seen in those subjected to either pituitary autotransplantation on or before day 5, or to pituitary homotransplantation, and to hypophysectomy on or before day 5. However, when the pituitary was autotransplanted on day 7 or day 9, or when the pituitary homotransplant-bearing rat was hypophysectomized on day 7, regression began earlier and was more rapid than in rats operated on on or before day 5. In fact, in the rats operated on on day 9, regression was as rapid as it was in the intact (sham operation) controls. Another group of rats subjected to pituitary autotransplantation and hysterectomy on day 2 was divided into four subgroups on day 21. In two of these, the rats received indomethacin-containing Silastic capsules inserted into each ovarian bursa; in the other two groups blank capsules were similarly inserted.(ABSTRACT TRUNCATED AT 400 WORDS)