A mouse model for cerebral babesiosis

Clinical symptoms and pathology observed in the cattle infected with Babesia bovis are quite similar to those of human cerebral malaria. Mechanisms involved in the pathogenesis of cerebral babesiosis, however, are still poorly understood because of the lack of a suitable experimental animal model. In this report, Masayoshi Tsuji and his colleagues describe B. bovis infection in severe combined immunodeficiency (SCID) mice, whose circulating red blood cells (RBCs) have been substituted with bovine RBCs (Bo-RBC-SCID mice). The infected mice not only develop a substantial level of parasitemia, but also show nerve symptoms and pathology similar to those observed in infected cattle.     

Accumulation of Japanese cedar pollen allergen,Cry j 1, in the protein body I of transgenic rice seeds using the promoter and signal sequence of glutelin GluB-1 gene

Recombinant Cry j 1, a Japanese cedar pollen allergen, was produced in rice seeds for potential use for oral immunotherapy. Cry j 1 cDNA was divided into two parts, an N-terminal half and a C-terminal half, and each was fused downstream to glutelin GluB-1 gene containing sequences of the promoter, 5 untranslated region and signal peptide. A gene for green fluorescent protein was also fused to the 3 end of the Cry j 1 fragment. Recombinant Cry j 1 of up to 16.6 g per mg total protein of the seeds was expressed in transgenic rice seeds. Although the recombinant Cry j 1 was expected to be accumulated in protein body II because of the employment of glutelin signal peptide, it was demonstrated to be accumulated exclusively in protein body I. The recombinant Cry j 1 was not shown to react with IgE of allergic patients, indicating the reduction of the risk of anaphylactic reaction. These results demonstrate that the transgenic rice seeds with the recombinant Cry j 1 would be useful for the study of oral immunotherapy.     

Changes in Chlorophyll a and b Content in Dark-Incubated Cotyledons Excised from Illuminated Seedlings: THE EFFECT OF CALCIUM

Cucumber seedlings were illuminated for various time periods, cotyledons excised, placed in the dark, and changes in chlorophyll a and b content monitored. During the dark periods, chlorophyll b content decreased while chlorophyll a did not. When the illumination time was lengthened, the percentage of chlorophyll b decomposition from initial levels decreased. Ca²⁺ at 50 millimolar prevented the decrease in chlorophyll b and caused a decrease in chlorophyll a. The effect of Ca²⁺ decreased with increased illumination time. Cycloheximide and chloramphenicol inhibited chlorophyll b decrease, but did not induce chlorophyll a decrease.     

Th1 cytokine-conditioned bone marrow-derived dendritic cells can bypass the requirement for Th functions during the generation of CD8+ CTL

Bone marrow-derived dendritic cell (BMDC) subsets have distinct immunoregulatory functions. Th1 cytokine-induced BMDC (BMDC1), compared with Th2 cytokine-induced BMDC2, have superior activities for the differentiation and expansion of CTL. To evaluate the cellular interactions between dendritic cells and CD8+ T cells for the induction of CTL, BALB/c-derived BMDC subsets were cocultured with purified CD8+ T cells from C57BL/6 mice. Our results demonstrate that BMDC1 support the generation of allogeneic CD8+ CTL in the absence of CD4+ Th cells. In contrast, BMDC0 (GM-CSF- plus IL-3-induced BMDC) and BMDC2 failed to promote the differentiation of CD8+ CTL. Using Ab-blocking experiments and studies with gene knockout mice, IL-2 and LFA-1 are demonstrated to be critical for BMDC1-induced CTL differentiation. Unexpectedly, BMDC1 were able to induce CTL from CD8+ T cells isolated from IFN-gamma-/- and IFN-gamma receptor-/- mice. However, BMDC1 produced higher levels of IFN-beta than other BMDC subsets, and anti-IFN-beta mAb blocked BMDC1-dependent CTL generation. These results indicated an indispensable role of IFN-beta, but not IFN-gamma, during BMDC1-induced CTL differentiation. We conclude that Th1-cytokine-conditioned BMDC1 can bypass Th cell function for the differentiation of naive CD8+ T cells into CTL.     

Galactosialidosis: molecular heterogeneity in biosynthesis and processing of protective protein for β-galactosidase

Summary Biosynthesis and processing of the protective protein for -galactosidase in normal and galactosialidosis fibroblasts were investigated using specific antiserum preparations. A 45-kd precursor was processed to a mature 30-kd protein in normal fibroblasts. The mature protective protein was not detected in any of the twelve galactosialidosis fibroblast strains examined in this study. The precursor was not detected in two cases and in the others was of heterogeneous molecular weight, i.e., normal, abnormally low, or abnormally high. These molecular abnormalities were not correlated with clinical manifestations of the patients.     

Metabolomics for metabolically manipulated plants: effects of tryptophan overproduction

Advances in molecular breeding technologies have enabled manipulation of the concentrations of specific plant components by modifying the genes that play a key role in their production. This has provided new opportunities to enhance the nutritional quality of major crops. However, given that metabolic pathways form a highly integrated network, any alteration in a given biosynthetic pathway is most likely to effect secondary and unpredicted changes in the metabolite profile of other pathways. Metabolomics technologies can contribute to the efficient detection of such unexpected effects caused by genetic modification. This has relevance not only from the perspective of safety evaluations of newly developed crops, but to basic science focused on uncovering hitherto unknown regulatory mechanisms associated with the biosynthesis and catabolism of primary and secondary metabolites in plants. In this review, recent advances in plant metabolic engineering for the overproduction of tryptophan (Trp), one of the essential amino acids, are described. In particular, the efficacy of a transgene OASA1D that encodes a mutant anthranilate synthase (AS) α subunit of rice in specifically elevating levels of Trp without marked secondary effects on the metabolite profile of rice is demonstrated. Related topics, such as regulation of Trp biosynthesis, possible interactions between the biosyntheses of Trp and other aromatic amino acids, and translocation of Trp in are discussed based on findings derived from metabolomic analyses of Trp-overproducing transgenic plants.     

Ornithine decarboxylase antizyme upregulates DNA-dependent protein kinase and enhances the nonhomologous end-joining repair of DNA double-strand breaks in human oral cancer cells

Ornithine decarboxylase (ODC) antizyme targets ODC for ubiquitin-independent proteosome degradation, thereby inhibiting polyamine synthesis. It has been shown to regulate DNA methylation and has tumor suppressor activity. Increasing evidence suggested that antizyme may also have ODC-independent functions. Here, we report that antizyme plays a role in DNA double-strand break repairs. A zinc-inducible human antizyme gene expression vector was transfected into UM1 human oral squamous cancer cells that do not express endogenous antizyme. The resultant upregulated genes were screened by cDNA arrays and confirmed by quantitative real-time polymerase chain reaction. DNA-dependent protein kinase including its catalytic subunit DNA-PKcs and regulatory subunit Ku70, two key proteins of the DNA damage repair machinery, was significantly upregulated after ectopic expression of antizyme. Consistently, we found that UM1 cells are sensitive to gamma irradiation and deficient in DNA damage repairs, as shown by radio-sensitivity and Comet assays. Ectopic expression of antizyme increased radio-resistance of UM1 cells and restored their capacity of DNA damage repairs to the level of UM2 cells that have an identical genetic background but express endogenous antizyme. Plasmid end-joining assays confirmed that antizyme enhances the ability of UM1 cells to repair DNA double-strand breaks by the nonhomologous end-joining pathway.     

Protective Effect of Oren-gedoku-to Against Induction of Neuronal Death by Transient Cerebral Ischemia in the C57BL/6 Mouse

We examined the neuroprotective effects of oren-gedoku-to (TJ15), a herbal medicine, after transient forebrain ischemia. Transient forebrain ischemia was induced by occlusion of both common carotid arteries for 15 min in C57BL/6 mice treated with TJ15. In the control ischemic group without TJ15 treatment, histologic examination of brain tissue collected seven days after reperfusion showed death of pyramidal cells in CA2-3 area of the hippocampus, unilaterally or bilaterally. In mice treated with oral TJ15 (845 mg/kg/day) for five weeks, the frequency of ischemic neuronal death was significantly lower. Immunohistochemistry for Cu/Zn-superoxide dismutase (Cu/Zn-SOD) showed strongly reactive astrocytes in the hippocampus of ischemic mice treated with TJ15. Damage to nerve cells by free radicals plays an important role in the induction of neuronal death by ischemia-reperfusion injury. Our results suggest that TJ15 protects against ischemic neuronal death by increasing the expression of Cu/Zn-SOD and suggest that oren-gedoku-to reduces the exposure of hippocampal neurons to oxidative stress.     

Molecular Cloning and Chromosomal Localization of Human Salt-Tolerant Protein

We have isolated a novel human cDNA coding for human salt-tolerant protein (HSTP), that is a homologue of the rat salt-tolerant protein (STP) and may contribute to salt-induced hypertension by modulating renal cation transport. The nucleotide sequence (1988 bp) of the HSTP cDNA contains an open reading frame encoding a polypeptide comprising 545 amino acids, two residues fewer than the rat STP cDNA. The predicted amino acid sequence exhibits 92% identity to that of the rat protein. HSTP contains predicted coiled-coil domains and Src Homology 3 domain, and shows a high degree of identity to CIP4 (Cdc42 target protein) and human Trip 10 (thyroid-hormone receptor interacting protein). We have mapped the HSTPgene to human chromosome 19 by fluorescence in situhybridization. This revised version was published online in July 2006 with corrections to the Cover Date.     

Plasma free amino acid profiling of five types of cancer patients and its application for early detection

Background

Recently, rapid advances have been made in metabolomics-based, easy-to-use early cancer detection methods using blood samples. Among metabolites, profiling of plasma free amino acids (PFAAs) is a promising approach because PFAAs link all organ systems and have important roles in metabolism. Furthermore, PFAA profiles are known to be influenced by specific diseases, including cancers. Therefore, the purpose of the present study was to determine the characteristics of the PFAA profiles in cancer patients and the possibility of using this information for early detection.

Methods and Findings

Plasma samples were collected from approximately 200 patients from multiple institutes, each diagnosed with one of the following five types of cancer: lung, gastric, colorectal, breast, or prostate cancer. Patients were compared to gender- and age- matched controls also used in this study. The PFAA levels were measured using high-performance liquid chromatography (HPLC)–electrospray ionization (ESI)–mass spectrometry (MS). Univariate analysis revealed significant differences in the PFAA profiles between the controls and the patients with any of the five types of cancer listed above, even those with asymptomatic early-stage disease. Furthermore, multivariate analysis clearly discriminated the cancer patients from the controls in terms of the area under the receiver-operator characteristics curve (AUC of ROC >0.75 for each cancer), regardless of cancer stage. Because this study was designed as case-control study, further investigations, including model construction and validation using cohorts with larger sample sizes, are necessary to determine the usefulness of PFAA profiling.

Conclusions

These findings suggest that PFAA profiling has great potential for improving cancer screening and diagnosis and understanding disease pathogenesis. PFAA profiles can also be used to determine various disease diagnoses from a single blood sample, which involves a relatively simple plasma assay and imposes a lower physical burden on subjects when compared to existing screening methods.