STAT3 couples with 14-3-3σ to regulate BCR signaling,B-cell differentiation,and IgE production

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Novel and recurrent EBP mutations in X-linked dominant chondrodysplasia punctata

Chondrodysplasia punctata (CDP) is a heterogeneous group of skeletal dysplasias characterized by stippled epiphyses. A subtype of CDP, X-linked dominant chondrodysplasia punctata (CDPX2), known also as Conradi-Hünermann-Happle syndrome, is a rare skeletal dysplasia characterized by short stature, craniofacial defects, cataracts, ichthyosis, coarse hair, and alopecia. The cause of CDPX2 was unknown until recent identification of mutations in the gene encoding Delta(8),Delta(7) sterol isomerase emopamil-binding protein (EBP). Twelve different EBP mutations have been reported in 14 patients with CDPX2 or unclassified CDP, but with no evidence of correlation between phenotype and nature of the mutation. To characterize additional mutations and investigate possible phenotype-genotype correlation, we sequenced the entire EBP gene in 8 Japanese individuals with CDP; 5 of them presented with a CDPX2 phenotypes. We found EBP mutations in all 5 CDPX2 individuals, but none in non-CDPX2 individuals. Three of these CDPX2 individuals carried novel nonsense mutations in EBPand the other two, separate missense mutations that had been reported also in different ethnic groups. Our results, combined with previous information, suggest all EBP mutations that produce truncated proteins result in typical CDPX2, whereas the phenotypes resulted from missense mutations are not always typical for CDPX2. Patients with nonsense mutations showed abnormal sterol profiles consistent with a defect in Delta(8), Delta(7) sterol isomerase. X-inactivation patterns of the patients showed no skewing, an observation that supports the assumption that inactivation of the EBP gene occurs at random in affected individuals.     

CD69-null mice protected from arthritis induced with anti-type II collagen antibodies

CD69, known as an early activation marker antigen on T and B cells, is also expressed on platelets and activated neutrophils, suggesting certain roles in inflammatory diseases. In order to address the role of CD69 in the pathogenesis of arthritis, we established CD69-null mice. CD69-null mice displayed a markedly attenuated arthritic inflammatory response when injected with anti-type II collagen antibodies. Cell transfer experiments with neutrophils, but not T cells or spleen cells, from wild-type mice into CD69-null mice restored the induction of arthritis. These results indicate a critical role for CD69 in neutrophil function in arthritis induction during the effector phase. Thus, CD69 would be a possible therapeutic target for arthritis in human patients.     

Characterization and identification of Oya virus,a Simbu serogroup virus of the genus Bunyavirus,isolated from a pig suspected of Nipah virus infection

Summary.  A virus, named Oya virus, was isolated in Vero cell cultures from the lungs of a pig suspected of Nipah virus infection. The virus was revealed as a spherical enveloped RNA virus with a diameter of 79 nm. For identification of Oya virus, RT-PCR was performed. A common primer set for S-RNA of the Simbu serogroup of the genus Bunyavirus was able to amplify a cDNA from Oya virus RNA. The sequence data of the product revealed that the partial gene of Oya virus S-RNA segment had 65–70% homology with published cDNA sequences of Simbu serogroup viruses. The phylogenetic analysis of the data showed that the Oya virus is grouped in Simbu serogroup, but is genetically distinct from the serogroup viruses that have been analyzed molecularly. Serological surveys revealed that the virus distributed widely and densely in Malaysia. Received January 5, 2002; accepted April 16, 2002 Published online July 19, 2002     

Births of normal daughters after MicroSort sperm separation and intrauterine insemination, in-vitro fertilization, or intracytoplasmic sperm injection

The world's first deliveries of normal babies after use of flow cytometric separated human sperm cells (MicroSort) for preconception gender selection are reported. Offspring were of the desired female gender in 92.9% of the pregnancies. Most of these pregnancies and births were achieved after simple intrauterine insemination.      

Three-dimensional two-layer collagen matrix gel culture model for evaluating complex biological functions of monocyte-derived dendritic cells

Dendritic cell-like cells (Mo-DCs) generated from peripheral blood monocytes with interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been used as tools to treat cancer patients (DC-vaccines). Because Mo-DCs have multiple antigen presentation-related functions, including phagocytosis, migration, cytokine production, and T cell stimulation, establishment of a method for simultaneously evaluating the various functions of Mo-DCs is important. We developed a new in vitro three-dimensional two-layer collagen matrix culture model that consists of a collagen gel containing Mo-DCs as the lower layer and a collagen gel containing necrotic GCTM-1 tumor cells and/or T cells as the upper layer. We used this system to observe simultaneously multiple functions of Mo-DCs by phase-contrast or fluorescence microscopy and to assess IL-12 secretion during more than 2 weeks of culture. We also observed interactions between Mo-DCs and necrotic GCTM-1 or T cells on an individual cell basis by time-lapse videomicroscopy. In addition, we collected Mo-DCs from the collagen gels by collagenase treatment and analyzed the expression of antigen presentation-related molecules such as HLA-DR, CD80, CD83, and CD86 on Mo-DCs. This model may be a useful tool for evaluation of the various functions of Mo-DCs used as DC vaccines and for studies of the complex behaviors of Mo-DCs in vivo.     

Secreted Reelin molecules form homodimers

During mammalian brain development, neurons are generated along the ventricle, migrate radially, and become aligned in defined patterns. These precise patterns of neuronal alignment are regulated by an extracellular matrix protein Reelin, and binding of Reelin to its receptors induces tyrosine phosphorylation of the intracellular adaptor protein disabled 1 (Dab1). We recently reported that Reelin molecules assemble to form a homomeric protein complex. Although the number of molecules in the full-length complex is unknown, recombinant N-terminal fragments, which contain the epitope for the function-blocking CR-50 antibody, assembled to form a complex of more than 40 monomers. When the N-terminus was deleted from Reelin, the truncated protein did not form a stable complex. To further characterize the Reelin assembly, we performed biochemical analysis of the full-length Reelin assembly in this study. Here, we report that a full-length Reelin forms a disulfide-linked homodimer. A chemical crosslinking experiment on secreted Reelin confirmed that only dimers are formed by the full-length protein. However, interestingly, chemical crosslinking of the N-terminus-truncated Reelin resulted in the formation of larger complexes, in addition to dimers, suggesting that the tertiary structure required for the proper and stable assembly/dimerization was altered by the truncation. The truncated protein did not induce efficient tyrosine phosphorylation of Dab1, although it bound well to the receptors. These findings demonstrate the functional importance of the N-terminal region of Reelin for proper dimerization and signaling. Proper but not simple extracellular crosslinking of the receptors by these dimers may be important for Reelin signaling to occur.     

Selection of appropriate antimicrobial agents to test and report by clinical microbiology laboratories

Clinical microbiology laboratories in Japan have not yet established standards for selecting the most appropriate antimicrobial agents for testing and reporting antimicrobial susceptibility that are comparable to the performance standards of the National Committee for Clinical Laboratory Standards(NCCLS) in the United States of America. Selection of the most appropriate antimicrobial agents for testing and reporting was discussed by a working group(WG) consisting of medical physicians, surgeons, pharmacists, medical technologists and medical microbiologists. The WG agreed on the following basic criteria for the selection of antimicrobial agents: 1) the agent should be useful when screening various resistant bacteria, 2) the agent should serve as a useful guide for physicians and residents when selecting antimicrobial agents, and 3) the agent should be useful for controlling nosocomial infections and resistant bacteria. Clinically isolated microorganisms were classified into 7 groups based on susceptibility to antimicrobial agents. These groups were Staphylococcus spp. or Enterococcus spp., Streptococcus spp. or Haemophilus spp., enterobacteriae, glucose non-fermenting gram positive rods(NFRs), anaerobic bacteria, fungi and mycobacterium. After considering clinical and bacteriological evidence, the WG decided on several antimicrobial agents for testing in clinical microbiology laboratories in Jichi Medical School Hospital. For the NFR group, these were Piperacillin(PIPC), ceftazidime(CAZ), cefepime, imipenem, amikacin and levofloxacin(LVFX). For the enterobacteriae group, these were Amplicillin(ABPC), PIPC, aztreonam, CAZ and LVFX. For the Staphylococcus spp. or Enterococcus spp. group, these were oxacillin, ABPC, vancomycin and gentamicin. We concluded that the most appropriate antimicrobial agent for testing and reporting must be economical and agreed upon at the hospital level, although the ultimate selection must be based on the available clinical and bacteriological evidence.     

Dietary nucleic acid and intestinal microbiota synergistically promote a shift in the Th1/Th2 balance toward Th1-skewed immunity

BACKGROUND: Intestinal microbiota are known to play an important role in the establishment of oral tolerance, thereby protecting the organism from food allergies. Dietary intake of nucleic acid (NA) is also reported to have such an anti-allergic effect; however, one unsolved question is whether or not dietary NA would act through a process of toll-like receptor 9 signaling activated by DNA containing a CpG motif, a well-known sequence leading to immunostimulatory activity. In this study, we focused on the question of whether the addition of dietary NA lacking CpG motifs would allow continued modulation of the Th1/Th2 balance. METHODS: Germ free (GF) and Bifidobacterium-infantis-monoassociated BALB/c mice were maintained on either an NA-free casein diet or on an NA-supplemented casein diet for 4 weeks. Thereafter, both the in vivo anti-casein antibody levels and in vitro splenocyte cytokine secretion pattern were evaluated. RESULTS: Feeding with a casein diet elicited a substantial increase in the serum anti-casein-specific IgG1, IgG2a, and IgE levels of GF mice fed the NA free-diet. The in vitro cytokine production profile showed that enhanced IL-4 production in the GF mice fed the NA free-diet was markedly reduced by the supplementation with dietary NA in both the GF and B.-infantis-monoassociated mice. In addition, IFN-gamma secretion increased in the B.-infantis-reconstituted mice fed the diet containing NA. CONCLUSIONS: These results suggest that dietary intake of NA devoid of CpG motifs may prevent the development of allergies via acceleration of Th1-dominant immunity.     

Histamine downregulates CD14 expression via H2 receptors on human monocytes

Lipopolysaccharide (LPS) binds to LPS-binding protein (LBP) in plasma and is delivered to the cell surface receptor CD14 on human monocyte. LPS is transferred to the transmembrane signaling receptor toll-like receptor (TLR) 4. In the present study, the effect of histamine on the expression of CD14 on human monocytes was investigated. Histamine concentration- and time-dependently decreased the expression of cell surface CD14, whereas histamine did not decrease mRNA for CD14 nor increase soluble CD14 (sCD14). The inhibitory effects of histamine on CD14 expression were antagonized by H2-receptor antagonist, but not by H1 and H3/H4 antagonist. The effects of selective H2-receptor agonists, 4-methylhistamine and dimaprit, on CD14 expression mimicked that of histamine indicating that histamine regulated CD14 expression through the stimulation of H2-receptors. The pretreatment with histamine partially inhibited the LPS-induced TNF-alpha production in human peripheral blood mononuclear cells (PBMC). Such inhibition might be due to the down-regulation of CD14 expression on monocytes by histamine.