Synthesis of 1-O-acyl- and 1-oxo-kamebanin analogues and their cytotoxic activity.

A series of 1-O-acyl- and 1-oxo-kamebanin analogues were prepared from kamebanin, isolated from Rabdosia excisa and their cytotoxicity was assayed on HL60 promyelocytic leukemia cells and HCT116 human colon cancer cells. The structure-activity relationship study showed that the presence of 1-O-acyl groups of a C3-C5 carbon chain increased the cytotoxic activity.

ORMDL3 overexpression facilitates FcεRI-mediated transcription of proinflammatory cytokines and thapsigargin-mediated PERK phosphorylation in RBL-2H3 cells.

INTRODUCTION: The chromosomal region 17q21 harbors the human orosomucoid-like 3 (ORMDL3) gene and has been linked to asthma and other inflammatory diseases. ORMDL3 is involved in the unfolded protein response (UPR), lipid metabolism, and inflammatory reactions. We investigated the effects of ORMDL3 overexpression in RBL-2H3 cells to determine the contribution of ORMDL3 to inflammatory disease development. METHODS: We generated ORMDL3 stably overexpressing RBL-2H3 cells to assess degranulation, transcriptional upregulation of interleukin-4 (IL-4), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and mitogen-activated protein kinase (MAPK) phosphorylation via FcεRI. In addition, we examined the effects of ORMDL3 overexpression on thapsigargin (TG)-mediated proinflammatory cytokine transcription and UPR by monitoring MAPK, protein kinase-like endoplasmic reticulum kinase (PERK), and inositol-requiring enzyme 1 (IRE1) phosphorylation. RESULTS: Overexpression of ORMDL3 enhanced IL-4, TNF-α, and MCP-1 expression after FcεRI cross-linking, whereas the sphingosine-1-phosphate (S1P) agonist FTY720 suppressed this enhancement. There was no significant difference in degranulation and MAPK phosphorylation via FcεRI-mediated activation between vector-transfected and ORMDL3-overexpressing cells. ORMDL3 overexpression accelerated TG-mediated PERK phosphorylation, while MAPK phosphorylation and proinflammatory cytokine expression showed no significant changes in ORMDL3-overexpressing cells. CONCLUSIONS: Our findings suggest that ORMDL3 plays an important role in regulating proinflammatory cytokine expression via the S1P pathway and selectively affects the UPR pathway in mast cells.

Increased expression of L-plastin in nasal polyp of patients with nonsteroidal anti-inflammatory drug-exacerbated respiratory disease.

BACKGROUND: Most patients with nonsteroidal anti-inflammatory drug-exacerbated respiratory disease (NERD) suffer from recurrence of nasal polyps. However, little is known about the specific cellular and molecular mechanisms contributing to the pathogenesis of nasal polyp development in patients with NERD in particular, especially at baseline when cyclooxygenase 1 inhibitors are not present. The objectives of this study were to identify proteins involved in the pathogenesis of nasal polyps in patients with NERD. METHODS: We collected nasal polyp tissue from patients with NERD and from patients with aspirin-tolerant chronic rhinosinusitis with nasal polyps (CRSwNP). Protein profiles were analyzed by 2-dimensional electrophoresis and identified several proteins, including L-plastin, as highly expressed. We examined L-plastin and tissue factor (TF) expression by immunohistochemical and immunofluorescence analyses. To examine the role of L-plastin in eosinophils, we knocked down L-plastin expression in Eol-1 cells by using siRNA transfection. RESULTS: L-plastin protein levels in nasal polyp tissue were increased in patients with NERD relative to those in patients with aspirin tolerant CRSwNP. Immunofluorescence analysis revealed that L-plastin was dominantly expressed in eosinophils and L-plastin and TF were co-expressed in eosinophils in NERD nasal polyp tissue. Knockdown of L-plastin in Eol-1 cells disrupted the cell surface distribution of TF by stimulation with granulocyte macrophage colony-stimulating factor. CONCLUSION: Increased expression of L-plastin by eosinophils may contribute to abnormal fibrin deposition through TF translocation to the eosinophil cell surface in NERD nasal polyp tissue, which in turn may contribute to the pathogenesis of NERD.

Genome-wide association study identifies eight new susceptibility loci for atopic dermatitis in the Japanese population.

Atopic dermatitis is a common inflammatory skin disease caused by interaction of genetic and environmental factors. On the basis of data from a genome-wide association study (GWAS) and a validation study comprising a total of 3,328 subjects with atopic dermatitis and 14,992 controls in the Japanese population, we report here 8 new susceptibility loci: IL1RL1-IL18R1-IL18RAP (P(combined) = 8.36 × 10(-18)), the major histocompatibility complex (MHC) region (P = 8.38 × 10(-20)), OR10A3-NLRP10 (P = 1.54 × 10(-22)), GLB1 (P = 2.77 × 10(-16)), CCDC80 (P = 1.56 × 10(-19)), CARD11 (P = 7.83 × 10(-9)), ZNF365 (P = 5.85 × 10(-20)) and CYP24A1-PFDN4 (P = 1.65 × 10(-8)). We also replicated the associations of the FLG, C11orf30, TMEM232-SLC25A46, TNFRSF6B-ZGPAT, OVOL1, ACTL9 and KIF3A-IL13 loci that were previously reported in GWAS of European and Chinese individuals and a meta-analysis of GWAS for atopic dermatitis. These findings advance the understanding of the genetic basis of atopic dermatitis.

ORAI1 genetic polymorphisms associated with the susceptibility of atopic dermatitis in Japanese and Taiwanese populations.

Atopic dermatitis is a chronic inflammatory skin disease. Multiple genetic and environmental factors are thought to be responsible for susceptibility to AD. In this study, we collected 2,478 DNA samples including 209 AD patients and 729 control subjects from Taiwanese population and 513 AD patients and 1027 control subject from Japanese population for sequencing and genotyping ORAI1. A total of 14 genetic variants including 3 novel single-nucleotide polymorphisms (SNPs) in the ORAI1 gene were identified. Our results indicated that a non-synonymous SNP (rs3741596, Ser218Gly) associated with the susceptibility of AD in the Japanese population but not in the Taiwanese population. However, there is another SNP of ORAI1 (rs3741595) associated with the risk of AD in the Taiwanese population but not in the Japanese population. Taken together, our results indicated that genetic polymorphisms of ORAI1 are very likely to be involved in the susceptibility of AD.

Variants of C-C motif chemokine 22 (CCL22) are associated with susceptibility to atopic dermatitis: case-control studies.

Atopic dermatitis (AD) is a common inflammatory skin disease caused by multiple genetic and environmental factors. AD is characterized by the local infiltration of T helper type 2 (Th2) cells. Recent clinical studies have shown important roles of the Th2 chemokines, CCL22 and CCL17 in the pathogenesis of AD. To investigate whether polymorphisms of the CCL22 gene affect the susceptibility to AD, we conducted association studies and functional studies of the related variants. We first resequenced the CCL22 gene and found a total of 39 SNPs. We selected seven tag SNPs in the CCL22 gene, and conducted association studies using two independent Japanese populations (1(st) population, 916 cases and 1,032 controls; 2(nd) population 1,034 cases and 1,004 controls). After the association results were combined by inverse variance method, we observed a significant association at rs4359426 (meta-analysis, combined P = 9.6×10⁻6; OR, 0.74; 95% CI, 0.65-0.85). Functional analysis revealed that the risk allele of rs4359426 contributed to higher expression levels of CCL22 mRNA. We further examined the allelic differences in the binding of nuclear proteins by electrophoretic mobility shift assay. The signal intensity of the DNA-protein complex derived from the G allele of rs223821, which was in absolute LD with rs4359426, was higher than that from the A allele. Although further functional analyses are needed, it is likely that related variants play a role in susceptibility to AD in a gain-of-function manner. Our findings provide a new insight into the etiology and pathogenesis of AD.

Genome-wide association study identifies three new susceptibility loci for adult asthma in the Japanese population.

Bronchial asthma is a common inflammatory disease caused by the interaction of genetic and environmental factors. Through a genome-wide association study and a replication study consisting of a total of 7,171 individuals with adult asthma (cases) and 27,912 controls in the Japanese population, we identified five loci associated with susceptibility to adult asthma. In addition to the major histocompatibility complex and TSLP-WDR36 loci previously reported, we identified three additional loci: a USP38-GAB1 locus on chromosome 4q31 (combined P = 1.87 × 10(-12)), a locus on chromosome 10p14 (P = 1.79 × 10(-15)) and a gene-rich region on chromosome 12q13 (P = 2.33 × 10(-13)). We observed the most significant association with adult asthma at rs404860 in the major histocompatiblity complex region (P = 4.07 × 10(-23)), which is close to rs2070600, a SNP previously reported for association with FEV(1)/FVC in genome-wide association studies for lung function. Our findings offer a better understanding of the genetic contribution to asthma susceptibility.

Genome-wide association studies of asthma.

Bronchial asthma is a common inflammatory disease caused by a combination of genetic and environmental factors. To discover the genes and cellular pathways underlying asthma, a large number of genetic studies have been conducted. Genome-wide association studies (GWAS), which comprehensively assess genes related to multifactorial diseases and drug reactivity, have enhanced understanding of human diseases. From 2007, GWAS of susceptibility to asthma in Caucasian, Mexican, and African-ancestry populations have been conducted and several susceptible loci were identified. Recently, much larger consortium-based GWAS analyses of collaborative samples with adequate statistical power were performed, and the implicated genes suggested a role for communication of epithelial damage to the adaptive immune system and activation of airway inflammation. Furthermore, GWAS identified candidate loci associated with natural variations in lung function, blood eosinophilia and eosinophilic esophagitis, which is inflammation of the esophagus with abnormal infiltration of eosinophils in an allergic reaction. Comparing GWAS in asthma and these clinical phenotypes might help to clarify the mechanisms underlying asthma. Pharmacogenomics analyses using GWAS regarding genetic factors related to the effectiveness of inhaled corticosteroid (ICS) therapy and inhaled beta(2)- adrenergic agonists are ongoing now. Although a more complete collection of associated genes and pathways is needed, biologic insights revealed by GWAS provide valuable insights into the pathophysiology of asthma and contribute to the development of better treatment and preventive strategies.

Aberrant methylation of the RIZ1 gene in myelodysplastic syndrome and acute myeloid leukemia.

We performed methylation specific PCR analysis on the RIZ1 promoter in MDS and AML. Methylation was detected in 17 of 34 MDS (50%) and 22 of 72 AML (31%) (p=0.053). Methylation was detected in eleven of 17 secondary AML from MDS (65%), and eleven of 55 de novo AML (20%) (p=0.0005). Bisulfite sequence revealed methylation at many CpG sites in the promoter. Decreased RIZ1 expression was accompanied by methylation in six of nine samples examined, while it was also observed in seven of 13 without methylation. Treatment of AML cells, that have RIZ1 methylation, with 5-Aza-dC, induced growth suppression with RIZ1 restoration. Our results suggest that the RIZ1 gene is inactivated in MDS and AML in part by methylation, whereas another mechanism should be involved in others.

JAK2 V617F mutation is rare in idiopathic erythrocytosis: a difference from polycythemia vera.

A single mutation 1849G>T in the JAK2 gene (V617F) has recently been described in classical myeloproliferative disorders (MPD). To investigate the incidence and clinical significance of the JAK2 mutation, we performed allele-specific polymerase chain reaction (PCR) and enzyme-based assessment in 11 idiopathic erythrocytosis (IE) and 15 polycythemia vera (PV) patients. Aberrant bands indicating the V617F mutation were detected in only one of 11 patients with IE, whereas all of the 15 patients with PV showed the JAK2 mutation. Sequence analysis was subsequently performed in the IE patient showing aberrant bands on allele-specific PCR, and a nucleotide change corresponding to the V617F mutation was detected in four of 29 clones tested. This patient might have progressed to PV according to the new WHO diagnostic criteria proposed in 2007, since a gradual increase in platelet counts was observed 4 years after the time of diagnosis. A further longitudinal study monitoring V617F positive-cells will clarify the process of progression from IE to PV in such a patient.

DL-alpha-tocopherol acetate mitigates maternal hyperthermia-induced pre-implantation embryonic death accompanied by a reduction of physiological oxidative stress in mice.

Maternal hyperthermia induces pre-implantation embryo death, which is accompanied by enhanced physiological oxidative stress. We evaluated whether the administration of DL-alpha-tocopherol acetate (TA) to hyperthermic mothers mitigated pre-implantation embryo death. Mice were exposed to heat stress (35 degrees C, 60% relative humidity) for 12 h or not heated (25 degrees C) on the day of mating. Twelve hours before the beginning of temperature treatment, TA was injected intraperitoneally at a dose of 1 g/kg body weight. After the treatment, zygotes were recovered and the developmental abilities and intracellular glutathione (GSH) levels were evaluated. Another set of mice, with or without TA treatment, was exposed to heat stress for 12, 24 and 36 h, and the urinary levels of the oxidative stress marker 8-hydroxy-2'-deoxyguanosine (8-OHdG) were measured. Heat stress significantly decreased the blastocyst development rate and the GSH content in zygotes, as compared with the non-heat-stressed embryos, while TA administration significantly mitigated the deleterious effects of heat stress with regard to both parameters. Moreover, although the urinary levels of 8-OHdG gradually increased according to the duration of heat exposure, with or without TA administration, the levels were lower in the TA-administered group than in the placebo-injected mice. These results suggest that heat stress enhances physiological oxidative stress, and that TA administration alleviates the hyperthermia-induced death of pre-implantation embryos by reducing physiological oxidative stress.