IL‐23 plays a significant role in the augmentation of particulate matter‐mediated allergic airway inflammation

It has been recently that particulate matter (PM) exposure increases the risk and exacerbation of allergic asthma. However, the underlying mechanisms and factors associated with increased allergic responses remain elusive. We evaluated IL‐23 and IL‐23R (receptor) expression, as well as changes in the asthmatic phenotype in mice administered PM and a low dose of house dust mite (HDM). Next, changes in the phenotype and immune responses were evaluated after intranasal administration of anti‐IL‐23 antibody during co‐exposure to PM and low‐dose HDM. We also performed in vitro experiments to investigate the effect of IL‐23. IL‐23 expression was significantly increased in Epcam+CD45− and CD11c+ cells, while that of IL‐23R was increased in Epcam+CD45− cells only in mice administered PM and low‐dose HDM. Administration of anti‐IL‐23 antibody led to decreased airway hyperresponsiveness, eosinophils, and activation of dendritic cells, reduced populations of Th2 Th17, ILC2, the level of IL‐33 and granulocyte‐macrophage colony‐stimulating factor (GM‐CSF). Inhibition of IL‐23 in PM and low‐dose HDM stimulated airway epithelial cell line resulted in decreased IL‐33, GM‐CSF and affected ILC2 and the activation of BMDCs. PM augmented the phenotypes and immunologic responses of asthma even at low doses of HDM. Interestingly, IL‐23 affected immunological changes in airway epithelial cells.     

Current state and perspectives on erythropoietin production

Erythropoietin is a major regulator of erythropoiesis which maintains the body's red blood cell mass and tissue oxygenation at an optimum level. Recombinant human erythropoietin (rhEPO), which is a widely used therapeutic agent for the treatment of anemia and which represents one of the largest biopharmaceuticals markets, is produced from recombinant Chinese hamster ovary cells. rhEPO is a glycoprotein with complex glycan structure, which is responsible for its therapeutic efficacy, including the in vivo activity and half-life. In order to obtain an optimal and consistent glycoform profile of rhEPO and concurrently maintain a high production yield, various approaches in drug development and cell culture technology have been attempted. Recent advances in rhEPO production are classified into three types: the development of improved rhEPO molecules by protein engineering; improvement of production host cells by genetic engineering; and culture condition optimization by fine control of the production mode/system, process parameters, and culture media. In this review, we focus on rhEPO production strategies as they have progressed thus far. Furthermore, the current status of the market and outlook on rhEPO and its derivatives are discussed.     

DNA ligase I is recruited to sites of DNA replication by an interaction with proliferating cell nuclear antigen: identification of a common targeting mechanism for the assembly of replication factories.

In mammalian cells, DNA replication occurs at discrete nuclear sites termed replication factories. Here we demonstrate that DNA ligase I and the large subunit of replication factor C (RF-C p140) have a homologous sequence of approximately 20 amino acids at their N-termini that functions as a replication factory targeting sequence (RFTS). This motif consists of two boxes: box 1 contains the sequence IxxFF whereas box 2 is rich in positively charged residues. N-terminal fragments of DNA ligase I and the RF-C large subunit that contain the RFTS both interact with proliferating cell nuclear antigen (PCNA) in vitro. Moreover, the RFTS of DNA ligase I and of the RF-C large subunit is necessary and sufficient for the interaction with PCNA. Both subnuclear targeting and PCNA binding by the DNA ligase I RFTS are abolished by replacement of the adjacent phenylalanine residues within box 1. Since sequences similar to the RFTS/PCNA-binding motif have been identified in other DNA replication enzymes and in p21(CIP1/WAF1), we propose that, in addition to functioning as a DNA polymerase processivity factor, PCNA plays a central role in the recruitment and stable association of DNA replication proteins at replication factories.     

Endothelial Transient Receptor Potential Conical Channel (TRPC)-3 Activation Induces Vasogenic Edema Formation in the Rat Piriform Cortex Following Status Epilepticus

Transient receptor potential canonical channel (TRPC) is a nonselective cation channel permeable to Ca²⁺, which express in many cell types, including neurons. However the alterations in TRPC receptor expressions in response to status epilepticus (SE) have not been explored. Therefore, the present study was designated to elucidate the roles of TRPC3 in neuronal death and vasogenic edema within the rat piriform cortex (PC) following SE. In non-SE animals, TRPC3 immunoreactivity was abundantly detected in the PC. Following SE, TRPC3 immunoreactivity was increased in neurons. Furthermore, TRPC3 expression was detected in endothelial cells that did not contain it in non-SE animals. Loss of SMI-71 (a blood–brain barrier antigen) immunoreactivity was also observed in TRPC3 positive endothelial cells. In addition, FJB positive neurons and vasogenic edema were noticeably detected in the PC. To directly determine whether TRPC3 activation is correlated to SE-induced vasogenic edema formation and neuronal damages in the PC, the effect of Pyr-3 (a TRPC3 antagonist) on SE-induced insults were investigated. Pyr-3 infusion effectively attenuated vasogenic edema in the PC as compared to the vehicle. Therefore, our findings indicate that TRPC3 activation/overexpression induced by SE may involve BBB disruption and neuronal damages in the rat PC following SE. Therefore, the present study was TRPC3 may play an important role in SE-induced vasogenic edema formation through BBB disruptions in the rat PC.     

Partial protection from cyclical selection generates a high level of polymorphism at multiple non‐neutral sites

Temporally varying selection is known to maintain genetic polymorphism under certain restricted conditions. However, if part of a population can escape from selective pressure, a condition called the “storage effect” is produced, which greatly promotes balanced polymorphism. We investigate whether seasonally fluctuating selection can maintain polymorphism at multiple loci, if cyclically fluctuating selection is not acting on a subpopulation called a “refuge.” A phenotype with a seasonally oscillating optimum is determined by alleles at multiple sites, across which the effects of mutations on phenotype are distributed randomly. This model resulted in long‐term polymorphism at multiple sites, during which allele frequencies oscillate heavily, greatly increasing the level of nonneutral polymorphism. The level of polymorphism at linked neutral sites was either higher or lower than expected for unlinked neutral loci. Overall, these results suggest that for a protein‐coding sequence, the nonsynonymous‐to‐synonymous ratio of polymorphism may exceed one. In addition, under randomly perturbed environmental oscillation, different sets of sites may take turns harboring long‐term polymorphism, thus making trans‐species polymorphism (which has been predicted as a classical signature of balancing selection) less likely.     

Involvement of caveolin‐1 in fibronectin‐induced mouse embryonic stem cell proliferation: Role of FAK,RhoA, PI3K/Akt,and ERK 1/2 pathways

Fibronectin (FN) is the foremost proliferation‐associated extracellular matrix component promoting cell adhesion, migration, and survival. We examined the effect of FN on cell proliferation and the related signaling pathways in mouse embryonic stem (ES) cells. FN increased integrin β1, Src, focal adhesion kinase (FAK), and caveolin‐1 phosphorylation levels in a time‐dependent manner. Phosphorylation of Src, FAK, and caveolin‐1 was attenuated by integrin β1 neutralizing antibody. Integrin β1, Src, and FAK coimmunoprecipitated with caveolin‐1 in the presence of FN. In addition, FN increased RhoA and Rho kinase activation, which were completely blocked by PP2, FAK small interfering RNA (siRNA), caveolin‐1 siRNA, or the caveolar disruptor methyl‐β‐cyclodextrin (MβCD). FN also increased phosphorylation of Akt and ERK 1/2, which were significantly blocked by either FAK siRNA, caveolin‐1 siRNA, MβCD, GGTI‐286 (RhoA inhibitor), or Y‐27632 (Rho kinase inhibitor). FN‐induced increase of protooncogenes (c‐fos, c‐myc, and c‐Jun) and cell‐cycle regulatory proteins (cyclin D1/CDK4 and cyclin E/CDK2) expression levels were attenuated by FAK siRNA or caveolin‐1 siRNA. Furthermore, inhibition of each pathway such as integrin β1, Src, FAK, caveolin‐1, RhoA, Akt, and ERK 1/2 blocked FN‐induced [³H]‐thymidine incorporation. We conclude that FN stimulates mouse ES cell proliferation via RhoA‐PI3K/Akt‐ERK 1/2 pathway through caveolin‐1 phosphorylation. J. Cell. Physiol. 226: 267–275, 2010. © 2010 Wiley‐Liss, Inc.     

Heterogeneity in human interleukin-3 receptors. A subclass that binds human granulocyte/macrophage colony stimulating factor

125I-Labeled recombinant human interleukin-3 (IL-3) was used to study the characteristics and distribution of receptors for IL-3 on human cells. Receptors were found on primary monocytes, on some strains of KG-1 cells, and on pre-B cell lines. Binding was rapid at 37 degrees C, while requiring several hours to reach equilibrium at 4 degrees C. Equilibrium binding studies indicated that IL-3 bound to a single class of high affinity receptor (less than 500 receptors/cell) with a Ka of approximately 1 x 10(10) M-1. Inhibition studies revealed that human granulocyte/macrophage colony stimulating factor partially inhibited the binding of 125I-IL-3 to human monocytes but not JM-1 cells. Additional analysis showed that on KG-1 cells, both IL-3 and GM-CSF partially competed specific binding of heterologous radiolabeled ligand, with approximately equivalent capacities. This competition occurred at both 37 and 4 degrees C. These results suggest heterogeneity in the binding sites for IL-3 and GM-CSF in which a subset of receptors binds only IL-3, a subset only GM-CSF, and another subset can bind both, all with high affinity. Additional heterogeneity was suggested by equilibrium binding of 125I-IL-3 to KG-1 cells which revealed a biphasic Scatchard plot containing a low affinity component not observed on monocytes and JM-1 cells.     

NF-kappaB activates fibronectin gene expression in rat hepatocytes

Resveratrol (RSVL), an edible polyphenolic stilbene, claims a myriad of cardiovascular benefits. However, the molecular underpinnings of such actions are poorly understood. Currently, in sheep coronary arteries (SCA), RSVL markedly (threefold) enhanced cGMP formation (t(1/2): 6.5 min; EC(50): 3 microM). This response was not abrogated by the phosphodiesterase inhibitor (IBMX, 0.5 mM), but was partly sensitive (20-30%) to either removal of the endothelium, treatment with the nitric oxide synthase-inhibitor (L-NMMA, 10 microM), or with the soluble GC (sGC)-inhibitor (ODQ, 10 microM). In membrane preparations from denuded SCA, either RSVL or the pGC agonist atrial natriuretic peptide (ANP, 0.1-1 microM) activated GC in the particulate, but not in the soluble, membrane fraction. By contrast, the nitric oxide donor, sodium nitroprusside (SNP, 1-10 microM), stimulated GC only in the soluble fraction. Further, pretreatment with RSVL partly desensitized the ANP response, but was additive to that of SNP. In arterial tension studies, RSVL relaxed PGF(2alpha)-precontracted denuded rings in a concentration-dependent manner, a response that was markedly enhanced (approximately 18 fold) in the presence of IBMX. Conversely, precontraction with phorbol ester, which also desensitizes pGC, blunted relaxations to RSVL but not to forskolin or SNP. These findings demonstrate that RSVL increases cGMP in coronary arteries, mostly by activation of pGC. This pathway triggers vasorelaxant responses that remain effective in endothelium-disrupted arteries.     

Solid cultures of thrips-pathogenic fungi Isaria javanica strains for enhanced conidial productivity and thermotolerance

Entomopathogenic fungi have great potential to control agricultural and horticultural insect pests, however optimizing conidial production systems to demonstrate high productivity and stability still needs additional efforts for successful field application and industrialization. Although many virulent entomopathogenic fungal isolates have been viewed as potential candidates in a laboratory environment, very few of the isolates are being used in practice for application in agricultural fields as commercial products. I. javanicus is an entomopathogenic fungus that is parasitic to various diverse coleopteran and lepidopteran insects and thought good candidate as biopesticdes. In this work, the basic characteristics of two entomopathogenic fungi, I. javanica FG340 and Pf04, were investigated in morphological examinations, genetic identification, and virulence against Thrips palmi, and then the feasibility of various grains substrates for conidial production was assessed, particularly focusing on conidial productivity and thermotolerance. Isaria javanica FG340 and Pf04 conidia were solid-cultured on 12 grains for 14?days in a Petri dish. Of the tested Italian millet, perilla seed, millet and barley-based cultures showed high conidial production. The four-grain media yielded >1?×?10⁹ conidia/g of I. javanica FG340 and Pf04. Pf04 strain had enhanced thermotolerance up to 45?°C when cultured on Italian millet. In application, it was easy to make a conidial suspension using the cultured grains, and several surfactants were tested to release the conidia. This work suggests several possible inexpensive grain substrates by which to promote conidial production combined with enhanced stability against exposure to high temperature.