: Omenn syndrome is a variant form of severe combined immunodeficiency. It is fatal unless treated by allogeneic stem cell transplantation (SCT), which is the only curative approach. However, both ...treatment‐related complications and graft rejection are major obstacles to treatment success. This report describes a case with Omenn syndrome who developed mixed chimerism after unrelated cord blood transplantation (UCBT). This case was successfully treated by altering the patient's immunosuppression and donor lymphocyte infusion (DLI) with donor cord blood‐derived activated CD4+ T cells ex vivo expanded from the cord blood cell residues in an infused bag. This novel development to expand CD4+ T‐lymphocytes from the donor cord blood unit for the use of DLI would serve as a useful method to overcome the risk of graft rejection in SCT for primary immunodeficiency disorders with residual cell‐mediated immunity without compounding graft‐vs.‐host disease, especially in the UCBT setting.
Dictyostelium is a favored model for studying problems in cell and developmental biology. To comprehend the genetic potential and networks that direct growth and multicellular development, we are ...performing a large‐scale analysis of Dictyostelium cDNAs. Here, we newly determine 7720 nucleotide sequences of cDNAs from the multicellular, slug stage (S) and 10 439 from the unicellular, vegetative stage (V). The combined 26 954 redundant ESTs were computer assembled using the PHRAP program to yield 5381 independent sequences. These 5381 predicted genes represent about half of the estimated coding potential of the organism. One‐third of them were classified into 12 functional categories. Although the overall classification patterns of the V and S libraries were very similar, stage‐specific genes exist in every category. The majority of V‐specific genes function in some aspect of protein translation, while such genes are in a minority in the S‐specific and common populations. Instead, genes for signal transduction and multicellular organization are enriched in the population of S‐specific genes. Genes encoding the enzymes of basic metabolism are mainly found in the common gene population. These results therefore suggest major differences between growing and developing Dictyostelium cells in the nature of the genes transcribed.
Abstract
Standard reference ranges for all laboratory test values are mandatory. This study was designed to establish a reference range for blood vitamin B
1
levels, since the normal range has not ...been determined in the Japanese population. We founded the Japan Committee for Vitamin Laboratory Standards, which was incorporated with the Vitamin Society of Japan and the Japanese Society of Nutrition and Food Science. We standardized whole blood vitamin B
1
levels using three HPLC techniques (post‐column reverse‐phase HPLC, pre‐column reverse‐phase HPLC, and precolumn GP‐HPLC). The reference range was obtained in 54 volunteers administered a 1,800 kcal diet with 2 mg of vitamin B
1
(1.74 mg measured) daily to avoid marginal vitamin B
1
deficiency in the population. The range for each assay was 26—47, 28—51, and 28—56 ng/ml, respectively. Our data suggest that 26—28 ng/ml is the lower limit of normal for whole blood vitamin B
1
, but further studies in a larger population are needed in order to obtain more definitive results.
The macropinocytic pathway in Dictyostelium discoideum is organized linearly. After actin-driven internalization, fluid material passes sequentially from endosomes to lysosomes, where molecules are ...degraded and absorbed. Residual material is exocytosed via post-lysosomal compartments. Syntaxin 7 is a SNARE (soluble N -ethylmaleimide-sensitive fusion protein attachment protein receptor) protein that is present and active in D. discoideum endosomes Bogdanovic, Bruckert, Morio and Satre (2000) J. Biol. Chem. 275, 36691-36697. Here we report the identification of its main SNARE partners by co-immunoprecipitation and MS peptide sequencing. The syntaxin 7 complex contains two co-t-SNAREs Vti1 (Vps10p tail interactor 1) and syntaxin 8 and a v-SNARE VAMP7 (vesicle-associated membrane protein 7) (where t-SNAREs are SNAREs of the target compartment and v-SNAREs are SNAREs present in donor vesicles). In endosomes and in vitro, syntaxin 7, Vti1 and syntaxin 8 form a complex that is able to bind VAMP7. Antibodies to syntaxin 8 and a soluble recombinant VAMP7 fragment both inhibit in vitro reconstituted D. discoideum endosome fusion. The lysosomal content of syntaxin 7, Vti1, syntaxin 8 and VAMP7 is low compared with that in endosomes, implying a highly active recycling or retention mechanism. A likely model is that VAMP7 is a v-SNARE present on vesicles carrying lysosomal enzymes, and that the syntaxin 7-Vti1-syntaxin 8 t-SNARE complex is associated with incoming endocytic material.
The Dictyostelium stalk cell inducer differentiation-inducing factor (DIF) directs tyrosine phosphorylation and nuclear accumulation of the STAT (signal transducer and activator of transcription) ...protein Dd-STATc. We show that hyperosmotic stress, heat shock and oxidative stress also activate Dd-STATc. Hyperosmotic stress is known to elevate intracellular cGMP and cAMP levels, and the membrane-permeant analogue 8-bromo-cGMP rapidly activates Dd-STATc, whereas 8-bromo-cAMP is a much less effective inducer. Surprisingly, however, Dd-STATc remains stress activatable in null mutants for components of the known cGMP-mediated and cAMP-mediated stress-response pathways and in a double mutant affecting both pathways. Also, Dd-STATc null cells are not abnormally sensitive to hyperosmotic stress. Microarray analysis identified two genes, gapA and rtoA, that are induced by hyperosmotic stress. Osmotic stress induction of gapA and rtoA is entirely dependent on Dd-STATc. Neither gene is inducible by DIF but both are rapidly inducible with 8-bromo-cGMP. Again, 8-bromo-cAMP is a much less potent inducer than 8-bromo-cGMP. These data show that Dd-STATc functions as a transcriptional activator in a stress-response pathway and the pharmacological evidence, at least, is consistent with cGMP acting as a second messenger.
Trichomonas vaginalis, a flagellated protozoan parasite, is the causative organism of trichomoniasis. We have recently demonstrated that
T. vaginalis induces apoptotic cell death via a Bcl-x
...L-dependent pathway in RAW264.7 macrophages. In this study, we attempted to characterize in detail the signaling cascades resulting in
T. vaginalis-induced macrophage apoptosis, focusing particularly on mitochondrial changes and the role of p38 mitogen-activated protein kinase (p38 MAPK) activation. We found that
T. vaginalis induced mitochondrial changes including the release of cytochrome
c and the serial activation of caspases, leading to the activation of p38 MAPK in macrophages. These biochemical changes culminated in the apoptosis of the host cells. Caspase inhibitors induced a significant inhibition of
T. vaginalis-induced nuclear damage, as well as the activation of p38 MAPK. Treatment with the p38 MAPK inhibitor, SB203580, or the overexpression of kinase-inactive p38 MAPK, induced an attenuation of
T. vaginalis-induced apoptosis but not cytochrome
c release, the activation of caspase-9 and caspase-3, or PARP cleavage. Furthermore, SB203580 treatment to human macrophages consistently blocked
T. vaginalis-induced apoptosis. Collectively, our findings indicate that p38 MAPK signaling cascade is requisite to apoptosis of
T. vaginalis-infected macrophage, and this apoptotic process occurs via the phosphorylation of p38 MAPK, which is located downstream of mitochondria-dependent caspase activation, conferring insight into the plausible molecular mechanism of
T. vaginalis-immune evasion from macrophage attack.
Tautomycetin (TMC) was identified as an immunosuppressor of activated T cells. Inhibition of T cell proliferation with TMC was observed at concentrations 100-fold lower than those needed to achieve ...maximal inhibition with cyclosporin A (CsA). TMC specifically blocked tyrosine phosphorylation of intracellular signal mediators downstream of Src tyrosine kinases in a T cell-specific manner, leading to apoptosis due to cleavage of Bcl-2, caspase-9, caspase-3, and poly(ADP-ribose) polymerase, but not caspase-1. In TMC-treated rats that received a heterotopic cardiac allograft, the graft survived more than 160 days, comparable to graft survival in allografted rats treated with CsA. Thus, TMC, whose mechanism of action is different from that of CsA or FK506, can be used as a potent T cell-specific immunosuppressor.
Hyper-IgM syndrome type 1 (HIGM1) is a primary immunodeficiency characterized by recurrent bacterial and opportunistic infections, associated with normal or high serum level of IgM and decreased ...serum levels of IgG, IgA and IgE due to the defect of class switch recombination.
CD40LG, located in Xq26, has been reported to be mutated in male HIGM1 patients. Here, we report the second case of a female HIGM1 with the defect of CD40 ligand (CD40L) expression and of soluble serum CD40L. Clinical course and HIGM phenotype was indistinguishable from that of male HIGM1 including severe neutropenia. High-resolution chromosome banding revealed that this patient's karyotype is 46, X, t(X;14)(q26.3;q13.1), and FISH analysis demonstrated that the break point of the chromosomal translocation is within
CD40LG. Using four chimeric cDNA clones obtained by 3′ RACE method, the break point was identified within the intron 4 of
CD40LG on X chromosome and non-coding region of chromosome 14. We also found an extremely skewed X-chromosome inactivation pattern by methylation-specific PCR. Thus, the reciprocal translocation caused the disruption of
CD40LG, resulting in defective CD40L expression in the female patient with an extremely skewed X-inactivation pattern in T cells leading to the HIGM1 phenotype.
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