Complement C3a-induced IL-17 plays a critical role in an IgE-mediated late-phase asthmatic response and airway hyperresponsiveness via neutrophilic inflammation in mice

Allergen-specific IgE plays an essential role in the pathogenesis of allergic asthma. Although there has been increasing evidence suggesting the involvement of IL-17 in the disease, the relationship between IL-17 and IgE-mediated asthmatic responses has not yet been defined. In this study, we attempted to elucidate the contribution of IL-17 to an IgE-mediated late-phase asthmatic response and airway hyperresponsiveness (AHR). BALB/c mice passively sensitized with an OVA-specific IgE mAb were challenged with OVA intratracheally four times. The fourth challenge caused a late-phase increase in airway resistance associated with elevated levels of IL-17(+)CD4(+) cells in the lungs. Multiple treatments with a C3a receptor antagonist or anti-C3a mAb during the challenges inhibited the increase in IL-17(+)CD4(+) cells. Meanwhile, a single treatment with the antagonist or the mAb at the fourth challenge suppressed the late-phase increase in airway resistance, AHR, and infiltration by neutrophils in bronchoalveolar lavage fluid. Because IL-17 production in the lungs was significantly repressed by both treatments, the effect of an anti-IL-17 mAb was examined. The late-phase increase in airway resistance, AHR, and infiltration by neutrophils in bronchoalveolar lavage fluid was inhibited. Furthermore, an anti-Gr-1 mAb had a similar effect. Collectively, we found that IgE mediated the increase of IL-17(+)CD4(+) cells in the lungs caused by repeated Ag challenges via C3a. The mechanisms leading to the IgE-mediated late-phase asthmatic response and AHR are closely associated with neutrophilic inflammation through the production of IL-17 induced by C3a.     

Gain of function by phosphorylation in Presenilin 1-mediated regulation of insulin signaling

During pregnancy, activation of the maternal immune system results in inflammation in the foetal nervous system. The causative agents are pro-inflammatory cytokines like interleukin-1β (IL-1β), produced by the foetus. In this study, we examine the effect of IL-1β on the proliferation and differentiation of neural progenitor cells (NPCs) to better understand its potential effects on the developing brain. We find that the IL-1β receptor (IL-1R1) is expressed in the ventral mesencephalon of the developing brain. Furthermore, IL-1R1 is expressed on Nestin-positive, Sox-2-positive NPCs. IL-1β treatment reduced the numbers of proliferating NPCs, an effect prevented by the IL-1R1 receptor antagonist. LDH and MTT assays, and western blot analysis for cleaved caspase 3 and poly(ADP-ribose) polymerase, confirmed that this was not due to an increase in cell death but rather an induction of differentiation. To further study the effects of IL-1β on cell fate determination, we differentiated NPCs in the presence and absence of IL-1β. Il-1β promoted gliogenesis and inhibited neurogenesis, an effect that required p38-MAPK kinase signalling. In summary, these data show that exposure of NPCs to IL-1β affects their development. This necessitates an examination of the consequences that maternal immune system activation during pregnancy has on the cellular architecture of the developing brain.     

Long-term expression of glial cell line-derived neurotrophic factor slows, but does not stop retinal degeneration in a model of retinitis pigmentosa

J. Neurochem. (2012) 122, 1047-1053. ABSTRACT: Retinitis pigmentosa is a group of diseases in which one of hundreds of mutations causes death of rod photoreceptor cells and then cones gradually die from oxidative damage. As different mutations cause rod cell death by different mechanisms, mutation-specific treatments are needed. Another approach is to use a neurotrophic factor to promote photoreceptor survival regardless of the mechanism of cell death, and previous studies have demonstrated encouraging short-term results with gene transfer of glial cell line-derived neurotrophic factor (GDNF). We generated rd10 mice with doxycycline-inducible expression of GDNF in photoreceptors (Tet/IRBP/GDNF-rd10 mice) or retinal pigmented epithelial cells (Tet/VMD2/GDNF-rd10 mice). In doxycycline-treated Tet/IRBP/GDNF-rd10 mice, there was a 9.3?×?10(4) -fold increase in Gdnf mRNA at P35 and although it decreased over time, it was still increased by 9.4?×?10(3) -fold at P70. Gdnf mRNA was increased 4.5?×?10(2) -fold in doxycycline-treated Tet/VMD2/GDMF-rd10 mice at P35 and was not significantly decreased at P70. GDNF protein levels were increased about 2.3-fold at P35 and 30% at P70 in Tet/IRBP/GDNF-rd10 mice, and in Tet/VMD2/GDNF-rd10 mice they were increased 30% at P35 and not significantly increased at P70. Despite the difference in expression, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 mice had comparable significant increases in outer nuclear layer thickness and mean photopic and scotopic ERG b-wave amplitudes compared with rd10 mice at P35 which decreased, but was still significant at P70. Compared with rd10 mice, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 mice had comparable significant improvements in cone density at P50 that decreased, but were still significant at P70. These data indicate that despite a large difference in expression of GDNF, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 provide comparable slowing of photoreceptor degeneration, but cannot stop the degeneration.     

Glutamate receptor δ1 induces preferentially inhibitory presynaptic differentiation of cortical neurons by interacting with neurexins through cerebellin precursor protein subtypes

Glutamate receptor (GluR) δ1 is widely expressed in the developing forebrain, whereas GluRδ2 is selectively expressed in cerebellar Purkinje cells. Recently, we found that trans-synaptic interaction of postsynaptic GluRδ2 and pre-synaptic neurexins (NRXNs) through cerebellin precursor protein (Cbln) 1 mediates excitatory synapse formation in the cerebellum. Thus, a question arises whether GluRδ1 regulates synapse formation in the forebrain. In this study, we showed that the N-terminal domain of GluRδ1 induced inhibitory presynaptic differentiation of some populations of cultured cortical neurons. When Cbln1 or Cbln2 was added to cultures, GluRδ1 expressed in HEK293T cells induced preferentially inhibitory presynaptic differentiation of cultured cortical neurons. The synaptogenic activity of GluRδ1 was suppressed by the addition of the extracellular domain of NRXN1α or NRXN1β containing splice segment 4. Cbln subtypes directly bound to the N-terminal domain of GluRδ1. The synaptogenic activity of GluRδ1 in the presence of Cbln subtypes correlated well with their binding affinities. When transfected to cortical neurons, GluRδ1 stimulated inhibitory synapse formation in the presence of Cbln1 or Cbln2. These results together with differential interactions of Cbln subtypes with NRXN variants suggest that GluRδ1 induces preferentially inhibitory presynaptic differentiation of cortical neurons by interacting with NRXNs containing splice segment 4 through Cbln subtypes.     

The highly conserved arginine residues at positions 76 through 78 of influenza A virus matrix protein M1 play an important role in viral replication by affecting the intracellular localization of M1

Influenza A virus matrix protein (M1) plays an important role in virus assembly and budding. Besides a well-characterized basic amino acid-rich nuclear localization signal region at positions 101 to 105, M1 contains another basic amino acid stretch at positions 76-78 that is highly conserved among influenza A and B viruses, suggesting the importance of this stretch. To understand the role of these residues in virus replication, we mutated them to either lysine (K), alanine (A), or aspartic acid (D). We could generate viruses possessing either single or combination substitutions with K or single substitution with A at any of these positions, but not those with double substitutions with A or a single substitution with D. Viruses with the single substitution with A exhibited slower growth and had lower nucleoprotein/M1 quantitative ratio in virions compared to the wild-type virus. In cells infected with a virus possessing the single substitution with A at position 77 or 78 (R77A or R78A, respectively), the mutated M1 localized in patches at the cell periphery where nucleoprotein and hemagglutinin colocalized more often than the wild-type did. Transmission electron microscopy showed that virus possessing M1 R77A or R78A, but not the wild-type virus, was present in vesicular structures, indicating a defect in virus assembly and/or budding. The M1 mutations that did not support virus generation exhibited an aberrant M1 intracellular localization and affected protein incorporation into virus-like particles. These results indicate that the basic amino acid stretch of M1 plays a critical role in influenza virus replication.     

Chemical modification of triplex-forming oligonucleotide to promote pyrimidine motif triplex formation at physiological pH

Extreme instability of pyrimidine motif triplex DNA at physiological pH severely limits its use in wide variety of potential applications, such as artificial regulation of gene expression, mapping of genomic DNA, and gene-targeted mutagenesis in vivo. Stabilization of pyrimidine motif triplex at physiological pH is, therefore, crucial for improving its potential in various triplex-formation-based strategies in vivo. To this end, we investigated the effect of 3'-amino-2'-O,4'-C-methylene bridged nucleic acid modification of triplex-forming oligonucleotide (TFO), in which 2'-O and 4'-C of the sugar moiety were bridged with the methylene chain and 3'-O was replaced by 3'-NH, on pyrimidine motif triplex formation at physiological pH. The modification not only significantly increased the thermal stability of the triplex but also increased the binding constant of triplex formation about 15-fold. The increased magnitude of the binding constant was not significantly changed when the number and position of the modification in TFO changed. The consideration of the observed thermodynamic parameters suggested that the increased rigidity of the modified TFO in the free state resulting from the bridging of different positions of the sugar moiety with an alkyl chain and the increased hydration of the modified TFO in the free state caused by the introduction of polar nitrogen atoms may significantly increase the binding constant at physiological pH. The study on the TFO viability in human serum showed that the modification significantly increased the resistance of TFO against nuclease degradation. This study presents an effective approach for designing novel chemically modified TFOs with higher binding affinity of triplex formation at physiological pH and higher nuclease resistance under physiological condition, which may eventually lead to progress in various triplex-formation-based strategies in vivo.     

Direct Absorption of Methyl Mercury by Lymph

Methyl mercury is contained in fish and seafood products and is taken up into the body in food. While the central nervous system is known as a target organ, methyl mercury also induces autoimmunity and acts as a potent immunosuppressor. The aim of the present study is to know whether methyl mercury is directly absorbed by lymph. Conscious rats were infused with methyl mercury (4 mg/kg) via duodenal tubing as a single pulse infusion, followed by the continuous infusion of saline, and lymphatic fluids were continuously collected from the thoracic lymph duct every 30 min until 360 min after infusion. Mercury was detected immediately after infusion, and total mercury contents in lymph gradually increased until 90–120 min, remained steady, and then gradually decreased until 360 min; however, the amount of mercury collected during 330–360 min was about twofold higher than during 0–30 min. The amount of cumulative mercury in lymph at 360 min was 1.4 μg. In contrast, blood mercury concentration was 2.4 μg/ml 5 min after infusion, with the value at 360 min being 12.6 times higher than at 5 min. Plasma mercury concentration was 56 ng/ml at 5 min, with hundreds of nanograms per milliliter of mercury detected until 360 min. From the present study, it is concluded that some methyl mercury is directly absorbed by lymph and remains steady 6 h after infusion.     

Are Additional Trace Elements Necessary in Total Parenteral Nutrition for Patients with Esophageal Cancer Receiving Cisplatin-Based Chemotherapy?

It is known that cisplatin induces the excretion of zinc from the urine and thereby reduces its serum concentration. However, the fluctuation of these trace elements during or after cisplatin-based chemotherapy has not been evaluated. To answer this question, we performed a clinical study in esophageal cancer patients undergoing cisplatin-based chemotherapy. Eighteen patients with esophageal cancer who were not able to swallow food or water orally due to complete stenosis of the esophagus were evaluated. The patients were divided into a control group [total parenteral nutrition (TPN) alone for 28?days, ten cases] and an intervention group (TPN with additional trace elements for 28?days, eight cases). The serum concentrations of zinc, iron, copper, manganese, triiodothyronin (T3), and thyroxin (T4), as alternative indicators of iodine, were measured on days?0, 14, and 28 of treatment, and statistically analyzed on day?28. In the control group, the serum concentration of copper was significantly decreased from 135.4 (day?0) to 122.1???g/ml (day?14), and finally to 110.6???g/ml (day?28, p?=?0.015). The concentration of manganese was also significantly decreased from 1.34 (day?0) to 1.17???g/ml (day?14) and finally to 1.20 (day?28, p?=?0.049). The levels of zinc, iron, T3, and T4 were not significantly changed. In the intervention group, the supplementation with trace elements successfully prevented these decreases in their concentrations. TPN with supplementary trace elements is preferable and recommended for patients who are undergoing chemotherapy in order to maintain the patients?? nutrient homeostasis.     

Abiotic Formation of Valine Peptides Under Conditions of High Temperature and High Pressure

We investigated the oligomerization of solid valine and the stabilities of valine and valine peptides under conditions of high temperature (150–200 °C) and high pressure (50–150 MPa). Experiments were performed under non-aqueous condition in order to promote dehydration reaction. After prolonged exposure of monomeric valine to elevated temperatures and pressures, the products were analyzed by liquid chromatography mass spectrometry comparing their retention times and masses. We identified linear peptides that ranged in size from dimer to hexamer, as well as a cyclic dimer. Previous studies that attempted abiotic oligomerization of valine in the absence of a catalyst have never reported valine peptides larger than a dimer. Increased reaction temperature increased the dissociative decomposition of valine and valine peptides to products such as glycine, β-alanine, ammonia, and amines by processes such as deamination, decarboxylation, and cracking. The amount of residual valine and peptide yields was greater at higher pressures at a given temperature, pressure, and reaction time. This suggests that dissociative decomposition of valine and valine peptides is reduced by pressure. Our findings are relevant to the investigation of diagenetic processes in prebiotic marine sediments where similar pressures occur under water-poor conditions. These findings also suggest that amino acids, such as valine, could have been polymerized to peptides in deep prebiotic marine sediments within a few hundred million years.