Induction and activation of the aryl hydrocarbon receptor by IL-4 in B cells

It is widely known that IL-4 and IL-13 act on various kinds of cells, including B cells, resulting in enhancement of proliferation, class switching to IgE and expression of several surface proteins. These functions are important for the recognition of the various antigens in B cells and are known to be involved in the pathogenesis of allergic diseases. However, it has not been known whether IL-4/IL-13 is involved in the metabolism of various kinds of xenobiotics including 2,3,7,8-tetra-chlorodibenzo-p-dioxin (TCDD), and it remains undetermined whether TCDD, an environmental pollutant, influences IgE production in B cells, exaggerating allergic reactions. We identified IL-4- or IL-13-inducible genes in a human Burkitt lymphoma cell line, DND-39, using microarray technology, in which the AHR gene was included. The AHR gene product, the aryl hydrocarbon receptor (AhR), was induced by IL-4 in both mouse and human B cells in a STAT6-dependent manner. IL-4 alone had the ability to translocate the induced AhR to the nuclei. TCDD, a ligand for AhR, rapidly degraded the induced AhR by the proteasomal pathway, although IL-4-activated AhR sustained its expression. AhR activated by IL-4 caused expression of a xenobiotic-metabolizing gene, CYP1A1, and TCDD synergistically acted on the induction of this gene by IL-4. However, the induction of AhR had no effect on IgE synthesis or CD23 expression. These results indicate that the metabolism of xenobiotics would be a novel biological function of IL-4 and IL-13 in B cells, whereas TCDD is not involved in IgE synthesis in B cells.     

Retrograde dopaminergic neuron degeneration following intrastriatal proteasome inhibition

Recent studies have suggested that defects in the ubiquitin-proteasome system (UPS) contribute to the etiopathogenetic mechanisms underlying dopaminergic neuronal degeneration in Parkinson's disease. The present study aims to study the effects of proteasome inhibition in the nerve terminals of nigrostriatal dopaminergic neurons in the substantia nigra pars compacta (SNpc). Following a unilaterally intrastriatal injection of lactacystin, a selective proteasome inhibitor, dopaminergic neurons in the ipsilateral SNpc progressively degenerated with alpha-synuclein-immunopositive intracytoplasmic inclusions. When lactacystin was administered at a high concentration, the striatum was simultaneously involved, and alpha-synuclein-immunopositive extracytoplasmic granules appeared extensively within the SN pars reticulata (SNpr). In addition, during the retrograde neuron degeneration in SN, the level of heme oxygenase-1 immunopositivity, an oxidative stress marker, was markedly increased in SNpc neurons. These results reveal that intrastriatal proteasome inhibition sufficiently induces retrograde dopaminergic neuronal degeneration with abundant accumulation of alpha-synuclein in the SN.     

The cytokines NAP-1 (IL-8), MCP-1, IL-1 beta,and GRO in rabbit inflammatory skin lesions produced by the chemical irritant sulfur mustard

Developing and healing dermal inflammatory lesions were produced in rabbits by the topical application of dilute sulfur mustard (SM),⁹ the military vesicant. In tissue sections of such lesions, cells containing the mRNA of important cytokines were identified with in situ hydridization techniques. These cytokines were neutrophil attractant/activation protein-1 (NAP-1 (also called IL-8)), monocyte chemoattractant (activating) protein 1 (MCP-1), interleukin 1 (beta) (IL-1 (beta)), and GRO (a growth factor and chemokine). Mononuclear cells (mainly macrophages and activated fibroblasts) contained the mRNA of all four of these cytokines. A higher percentage of cytokine-producing mononuclear cells (macrophages and activated fibroblasts) was present in lesions at 2 days (their peak size) than at 6 days, when they were almost healed. Granulocytes emigrated from the bloodstream, passed through the lesions, and were the major constituent of the protective crust. This sequence correlated with the distribution of cells able to produce NAP-1: At 2 days and 6 days, the mononuclears that contained messenger RNA for this granulocyte chemoattractant were found mainly in the upper part of the dermis. At 2 days and 6 days, cells containing the mRNA of IL-1, a primary cytokine, were also found predominantly in the upper dermis, i.e., nearest the site of injury. In contrast, mononuclears containing the mRNA of MCP-1 (a monocyte chemoattractant), and the mRNA of GRO (a granulocyte chemoattractant) were more equally distributed throughout the dermis. SM stimulated hair follicle epithelial cells to up-regulate GRO mRNA and, to a lesser degree, NAP-1 mRNA. Apparently, the irritation produced by SM directly or indirectly induces such epithelial cells to manufacture these growth factors. In the rabbit, hair follicles are known to be the main source of new epithelial cells after the covering epithelium has been destroyed. Therefore, GRO is probably a major autocrine-paracrine stimulus for such repair. A brief review of the role of cytokines in dermal inflammation is presented.Abbreviations SM Sulfur mustard: bis(2-chloroethyl)sulfide - GM-CSF Granulocyte-Macrophage Colony Stimulating Factor - GRO A member of the CXC subfamily of chemokines that promotes the multiplication of cells, formerly called melanoma growth stimulating activity (MGSA) - IFN (gamma)-Interferon-gamma - IL-1 Interleukin 1 - IL-8 Interleukin 8 (same as NAP-1)-a CXC chemokine - MCP-1 Monocy te Chemoattractant (Activating) Protein-1-a C-C chemokine - NAP-1 Neutrophil Attractant/Activating Protein-1 (same as IL-8)-a C-X-C chemokine - TGF (beta) Transforming Growth Factor (beta) - TNF (alpha) Tumor Necrosis Factor (alpha) - EDTA Ethylenediamine tetraacetate - DEPC Diethylpyrocarbonate - PBS Phosphate-buffered saline solution - PGE₂ Prostaglandin E₂ - PGI₂ Prostaglandin I₂ (prostacyclin) - SSC Sodium chloride-sodium citrate solution On leave of absence from the Institute for Medical Immunology, Kumamoto University School of Medicine, Kumamoto, Japan.On leave of absence from the Department of Internal Medicine, Oita Medical University, Oita, Japan.     

Genetic Conservation of Hemagglutinin Gene of H9 Influenza Virus in Chicken Population in Mainland China

The hemagglutinin (HA) genes of 12 H9N2 influenza virus strains isolated from chickens in Mainland China during the period 1995–2002 were genetically analyzed. All the isolates possessed the same amino acid motif -R-S-S-R/G-L- at the cleavage site of HA. Except for the conserved amino acids, as is the case in the other avian influenza viruses, located in the receptor binding site, all of the 12 isolates possessed N at amino acid position 183; A, T, or V at position 190; K at position 137, whereas the representative strains of the other lineage (except Dk/HK/Y280/97-like lineage) virus of H9N2 viruses had H, E, and R at these positions respectively. These could be considered as the partial molecular markers of the H9 viruses isolated from chickens in Mainland China. Phylogenetic analyses showed HA genes of these isolates belonged to that of A/duck/Hong Kong/Y280/97-like virus lineage. No A/quail/Hong Kong/Gl/97-like virus was found in chicken, population since the outbreak of H9N2 influenza in Mainland China in 1992. The available evidence indicates that HA genes of H9 influenza virus circulating in Mainland China during the past years were well conserved.     

Selective haploinsufficiency of longer isoforms of PTCH1 protein can cause nevoid basal cell carcinoma syndrome

Nevoid basal cell carcinoma syndrome (NBCCS) is an autosomal dominant disorder characterized by developmental defects and tumorigenesis. The gene responsible for NBCCS is PTCH1. The PTCH1 gene has five alternatively used first exons resulting in the translation of three isoforms of the PTCH1 protein; that is, PTCHL, PTCHM and PTCHS. However, the biological significance of each isoform is unclear. Here we show an individual with NBCCS carrying a nonsense mutation in PTCH1 exon2, c.387G>A (p.W129X). As the mutation lay upstream of the ATG codon used for PTCHS translation, the mutant allele still expressed RNA isoforms that encode PTCHS. These results clearly demonstrate that a selective haploinsufficiency of longer isoforms of the PTCH1 protein, PTCHL and PTCHM, but not PTCHS is sufficient to cause NBCCS. Although mice selectively deficient in PTCHS isoforms are currently unavailable, this study sheds light on the complex in vivo roles of PTCH1 isoforms.     

Coordinated expression of REG4 and aldehyde dehydrogenase 1 regulating tumourigenic capacity of diffuse‐type gastric carcinoma‐initiating cells is inhibited by TGF‐β

Aldehyde dehydrogenase 1 (ALDH1) has been shown to serve as a marker for cancer‐initiating cells (CICs), but little is known about the regulation of the CIC functions of ALDH1+ cancer cells. We isolated ALDH1+ cells from human diffuse‐type gastric carcinoma cells and characterized these cells using an Aldefluor assay. ALDH1+ cells constituted 5–8% of the human diffuse‐type gastric carcinoma cells, OCUM‐2MLN and HSC‐39; were more tumourigenic than ALDH1? cells; and were able to self‐renew and generate heterogeneous cell populations. Using gene expression microarray analyses, we identified REG4 (regenerating islet‐derived family, member 4) as one of the genes up‐regulated in ALDH1+ cells, and thus as a novel marker for ALDH1+ tumour cells. Induced expression of REG4 enhanced the colony‐forming ability of OCUM‐2MLN cells, while knockdown of REG4 inhibited the tumourigenic potential of ALDH1+ cells. We further found that TGF‐β signalling reduces the expression of ALDH1 and REG4, and the size of the ALDH1+ cell population. In human diffuse‐type gastric carcinoma tissues, the expression of ALDH1 and REG4 correlated with each other, as assessed by immunohistochemistry, and ALDH1 expression correlated inversely with Smad3 phosphorylation as a measure of TGF‐β signalling. These findings illustrate that, in diffuse‐type gastric carcinoma, REG4 is up‐regulated in ALDH1+ CICs, and that the increased tumourigenic ability of ALDH1+ cells depends on REG4. Moreover, TGF‐β down‐regulates ALDH1 and REG4 expression, which correlates with a reduction in CIC population size and tumourigenicity. Targeting REG4 in ALDH1+ CICs may provide a novel strategy in the treatment of diffuse‐type gastric carcinoma. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Yeast transformation mediated by Agrobacterium strains harboring an Ri plasmid: comparative study between GALLS of an Ri plasmid and virE of a Ti plasmid

Agrobacterium strains containing a Ti plasmid can transfer T-DNA not only to plants but also to fungi, including the yeast Saccharomyces cerevisiae. However, no Agrobacterium strain harboring an Ri plasmid has been evaluated in fungal transformation. Some Ri plasmids have GALLS , instead of virE1 and virE2. GALLS protein can functionally substitute in plant transformation for a structurally different protein VirE2. In this study, we compared the yeast transformation ability among Agrobacterium donors: a strain containing a Ti plasmid, strains harboring either an agropine-type or a mikimopine-type Ri plasmid, and a strain having a modified Ri plasmid supplemented with a Ti plasmid type virE operon. Agrobacterium strains possessing GALLS transformed yeast cells far less efficiently than the strain containing virE operon. Production of GALLS in recipient yeast cells improved the yeast transformation mediated by an Agrobacterium strain lacking neither GALLS nor virE operon. A reporter assay to detect mobilization of the proteins fused with Cre recombinase revealed that VirE2 protein is much more abundant in yeast cells than GALLS. Based on these results, we concluded that the low yeast transformability mediated by Agrobacterium strains having the Ri plasmid is because of low amount of mobilized GALLS in yeast cells.     

Hepatocyte growth factor concentration in the early second-trimester amniotic fluid does not predict fetal growth at birth.

The purpose of this study was to evaluate whether hepatocyte growth factor (HGF) concentrations in the early second-trimester amniotic fluid predict fetal growth at birth. HGF and insulin-like growth factor-I (IGF-I) concentrations in the early second-trimester amniotic fluid were measured in 12 pregnancies with small for gestational age (SGA) infants, 84 pregnancies with appropriate for gestational age (AGA) infants, and eight pregnancies with large for gestational age (LGA) infants. HGF concentrations were measured from the early second-trimester amniotic fluid samples using an enzyme-linked immunosorbent assay. IGF-I concentrations were measured from the early second-trimester amniotic fluid samples using an immunoradiometric assay. Maternal age in AGA group (34.2 +/- 5.5 years) was significantly lower than in SGA (37.9 +/- 3.0 years) and LGA (37.6 +/- 3.3 years) groups (P < 0.05). There were no significant differences for parity or gestational age at amniocentesis among the groups. There were significant differences for birth age, birth weight, neonatal height, and placental weight among the groups (P < 0.05). HGF concentrations in SGA, AGA and LGA groups were 16.9 +/- 6.6, 16.7 +/- 9.0 and 20.2 +/- 14.8 ng/ml respectively (not significant). There was no correlation between amniotic fluid HGF concentrations and birth weight, height or placental weight. There were also no significant differences for amniotic fluid IGF-I concentrations among the three groups. These results suggest that differences in HGF concentrations in the early second-trimester amniotic fluid do not predict fetal growth at birth. Further study is needed to clarify the role of high HGF concentrations in early second-trimester amniotic fluid during pregnancy.     

Comparison of alterations in fetal regional arterial vascular resistance in appropriate-for-gestational-age singleton, twin and triplet pregnancies.

The objective of this longitudinal study was to evaluate alterations in fetal vascular resistance of fetal peripheral arteries with advancing gestation in singleton appropriate-for-gestational-age (S-AGA), twin appropriate-for-gestational-age (Tw-AGA) and triplet appropriate-for-gestational-age (Tri-AGA) infants. Colour Doppler flow imaging and pulsed Doppler ultrasonographic examinations were performed on 35 S-AGA, 52 Tw-AGA and 12 Tri-AGA fetuses. The pulsatility index for middle cerebral artery (MCAPI), umbilical artery (UAPI), descending aorta (DAPI), splenic artery (SAPI), renal artery (RAPI) and femoral artery (FAPI) was measured as vascular resistance every 2 weeks after 15 weeks of menstrual age until delivery. Optimal models and normal ranges for pulsatility index for each artery in each group were generated. The alterations in various fetal regional arterial pulsatility indices with advancing gestational age showed no significant differences in S-AGA, Tw-AGA and Tri-AGA infants, respectively. These results suggest that there is no significant difference for regional arterial vascular resistance in AGA fetuses among singleton, twin, and triplet pregnancies, whereas there was a slight difference in fetal growth pattern among singleton, twin, and triplet pregnancies described in our previous investigation.