Genetic structure of Japanese Chattonella marina (Raphidophyceae) populations revealed using microsatellite markers

Chattonella marina var. antiqua and C. marina var. marina (Raphidophyceae) are red tide‐forming, harmful phytoplankton species. We investigated the genetic diversity and genetic relationship among the populations using microsatellite markers to identify putative sources of C. marina var. antiqua and C. marina var. marina in Japanese coastal populations. A positive correlation between genetic divergence and geographical distance (isolation by distance) was recognized for C. marina var. antiqua. The C. marina var. antiqua populations were established throughout a geological time scale, and genetic divergence had progressed in each population with gene flow depending on geographic distances. In contrast, isolation by distance was not observed for C. marina var. marina populations, and the genetic divergence among populations was extremely high. The Tokyo Bay population of C. marina var. marina, which was first recognized in 2008, had many private alleles but was related to the Kagoshima Bay population. The Tokyo Bay population may have been established by several invasions from the Kagoshima Bay population and other regions.     

Therapeutic effect of IL-12/23 and their signaling pathway blockade on brain ischemia model

Recently, T cell cytokines such as IL-17 and IFN-γ have been shown to play important roles in the progression of brain injury induced by ischemia. We have shown that IL-23 from infiltrated macrophages activates γδT cells, thereby inducing IL-17 from these cells. However, deletion of the IL-23 gene in mice showed a more dramatic protective effect against brain ischemia reperfusion (I/R) model than γδT cell depletion did, suggesting that IL-23 plays some other pivotal role in brain injury in addition to its role in IL-17 induction. To develop therapeutic methods based on these findings, we examined the effect of the JAK kinase inhibitor CP-690550 and an anti-IL12/23 monoclonal antibody on an I/R model. CP-690550 efficiently inhibited IL-17 production from memory T cells in vitro and partly suppressed infarct volume increase after I/R. Anti-p40 antibody, which blocks both IL-12 and IL-23, efficiently suppressed I/R injury and improved recovery of neurological deficits. The number of IL-17-producing cells was decreased by anti-p40 antibody treatment. Thus the JAK inhibitor and anti-p40 antibody, both of which have already been under trial for the treatment of several human inflammatory diseases, appear to be promising therapeutic agents for the amelioration of stroke.     

Cephalometric study of elderly with nearly intact dental arches

Objective: The purpose of this study was to investigate the craniofacial morphology of elderly people with many remaining teeth using cephalometric analysis. Subjects and methods: The subjects were 30 Japanese elderly who participated in the ‘8020 campaign 2001’ in Bunkyo Ward, Tokyo, organised by The Dental Association of Tokyo, as well as 30 Japanese young adults with normal occlusion. Lateral cephalograms of all subjects were analysed using the Coben method. Results: In the female elderly group, the lower face depth was smaller than in the younger adults. In the male elderly group, the height and depth of both the total face and the lower face were longer than in the younger group. In comparing the 8020 achievers with the younger group, the proportion of the lower facial height was greater than the upper facial height, and this finding was more pronounced in women than in men. Conclusion: For the lateral facial pattern of the elderly, a reduction of lower facial height because of tooth occlusal reduction was not apparent. It was clear that there are age differences for males and females; in addition, differences in the total face and lower face area of the elderly group were due to their having many remaining teeth over a long time period. Also, these changes were more apparent in women than in men, and it is clear that there is a male–female difference in ageing.     

The regulator of the F1 motor: inhibition of rotation of cyanobacterial F1-ATPase by the epsilon subunit

The chloroplast-type F(1) ATPase is the key enzyme of energy conversion in chloroplasts, and is regulated by the endogenous inhibitor epsilon, tightly bound ADP, the membrane potential and the redox state of the gamma subunit. In order to understand the molecular mechanism of epsilon inhibition, we constructed an expression system for the alpha(3)beta(3)gamma subcomplex in thermophilic cyanobacteria allowing thorough investigation of epsilon inhibition. epsilon Inhibition was found to be ATP-independent, and different to that observed for bacterial F(1)-ATPase. The role of the additional region on the gamma subunit of chloroplast-type F(1)-ATPase in epsilon inhibition was also determined. By single molecule rotation analysis, we succeeded in assigning the pausing angular position of gamma in epsilon inhibition, which was found to be identical to that observed for ATP hydrolysis, product release and ADP inhibition, but distinctly different from the waiting position for ATP binding. These results suggest that the epsilon subunit of chloroplast-type ATP synthase plays an important regulator for the rotary motor enzyme, thus preventing wasteful ATP hydrolysis.     

Substrate-free structure of a monomeric NADP isocitrate dehydrogenase: an open conformation phylogenetic relationship of isocitrate dehydrogenase

Both monomeric and dimeric NADP+-dependent isocitrate dehydrogenase (IDH) belong to the metal-dependent beta-decarboxylating dehydrogenase family and catalyze the oxidative decarboxylation from 2R,3S-isocitrate to yield 2-oxoglutarate, CO2, and NADPH. It is important to solve the structures of IDHs from various species to correlate with its function and evolutionary significance. So far, only two crystal structures of substrate/cofactor-bound (isocitrate/NADP) NADP+-dependent monomeric IDH from Azotobacter vinelandii (AvIDH) have been solved. Herein, we report for the first time the substrate/cofactor-free structure of a monomeric NADP+-dependent IDH from Corynebacterium glutamicum (CgIDH) in the presence of Mg2+. The 1.75 A structure of CgIDH-Mg2+ showed a distinct open conformation in contrast to the closed conformation of AvIDH-isocitrate/NADP+ complexes. Fluorescence studies on CgIDH in the presence of isocitrate/or NADP+ suggest the presence of low energy barrier conformers. In CgIDH, the amino acid residues corresponding to the Escherichia coli IDH phosphorylation-loop are alpha-helical compared with the more flexible random-coil region in the E. coli protein where IDH activation is controlled by phosphorylation. This more structured region supports the idea that activation of CgIDH is not controlled by phosphorylation. Monomeric NADP+-specific IDHs have been identified from about 50 different bacterial species, such as proteobacteria, actinobacteria, and planctomycetes, whereas, dimeric NADP+-dependent IDHs are diversified in both prokaryotes and eukaryotes. We have constructed a phylogenetic tree based on amino acid sequences of all bacterial monomeric NADP+-dependent IDHs and also another one with specifically chosen species which either contains both monomeric and dimeric NADP+-dependent IDHs or have monomeric NADP+-dependent, as well as NAD+-dependent IDHs. This is done to examine evolutionary relationships.     

Antihypertensive effect of angiotensin I-converting enzyme inhibitory peptides from a sesame protein hydrolysate in spontaneously hypertensive rats

Sesame peptide powder (SPP) exhibited angiotensin I-converting enzyme (ACE) inhibitory activity, and significantly and temporarily decreased the systolic blood pressure (SBP) in spontaneously hypertensive rats (SHRs) by a single administration (1 and 10 mg/kg). Six peptide ACE inhibitors were isolated and identified from SPP. The representative peptides, Leu-Val-Tyr, Leu-Gln-Pro and Leu-Lys-Tyr, could competitively inhibit ACE activity at respective Ki values of 0.92 microM, 0.50 microM, and 0.48 microM. A reconstituted sesame peptide mixture of Leu-Ser-Ala, Leu-Gln-Pro, Leu-Lys-Tyr, Ile-Val-Tyr, Val-Ile-Tyr, Leu-Val-Tyr, and Met-Leu-Pro-Ala-Tyr according to their content ratio in SPP showed a strong antihypertensive effect on SHR at doses of 3.63 and 36.3 microg/kg, which accounted for more than 70% of the corresponding dosage for the SPP-induced hypotensive effect. Repeated oral administration of SPP also lowered both SBP and the aortic ACE activity in SHR. These results demonstrate that SPP would be a beneficial ingredient for preventing and providing therapy against hypertension and its related diseases.     

Prognostic Value of EGFR Mutation and ERCC1 in Patients with Non-Small Cell Lung Cancer Undergoing Platinum-Based Chemotherapy

Background

In order to improve the outcome of patients with non-small cell lung cancer (NSCLC), a biomarker that can predict the efficacy of chemotherapy is needed. The aim of this study was to assess the role of EGFR mutations and ERCC1 in predicting the efficacy of platinum-based chemotherapy and the outcome of patients with NSCLC.

Methods

We conducted a retrospective study to analyze the relationships between EGFR mutations or ERCC1 expression and progression-free survival (PFS) in patients with NSCLC who received platinum-based chemotherapy. EGFR mutation status was determined using the peptide nucleic acid-locked nucleic acid polymerase chain reaction clamp method, and immunohistochemistry was used to examine the expression of ERCC1 in tumor samples obtained from the patients.

Results

Among the NSCLC patients who received platinum-based chemotherapy, the median PFS was significantly better in those who had never smoked and those with exon 19 deletion, and the median overall survival (OS) was significantly better in those who had never smoked, those with exon 19 deletion, and women. Cox regression analysis revealed that exon 19 deletion and having never smoked were significantly associated with both PFS and OS. Subset analysis revealed a significant correlation between ERCC1 expression and EGFR mutation, and ERCC1-negative patients with exon 19 deletion had a longer PFS than the other patients; ERCC1-positive patients without exon 19 deletion had a shorter PFS than the other patients.

Conclusions

Our results indicate that among NSCLC patients receiving platinum-based chemotherapy, those with exon 19 deletion have a longer PFS and OS. Our findings suggest that platinum-based chemotherapy is more effective against ERCC1-negative and exon 19-positive NSCLC.     

Effects of low molecular weight soybean peptide on mRNA and protein expression levels of differentiation markers in normal human epidermal keratinocytes

Low molecular weight soybean peptide (LSP) was applied to normal human epidermal keratinocytes, and the results showed a significant increase in the gene expression levels of involucrin, transglutaminase, and profilaggrin. Filaggrin protein levels were also significantly higher. It is possible that LSP has an epidermal cell differentiation-promoting effect and may be able to regulate metabolism of the epidermis.     

Cell death of Nicotiana benthamiana is induced by secreted protein NIS1 of colletotrichum orbiculare and is suppressed by a homologue of CgDN3

Colletotrichum orbiculare, the causal agent of cucumber anthracnose, infects Nicotiana benthamiana. Functional screening of C. orbiculare cDNAs in a virus vector-based plant expression system identified a novel secreted protein gene, NIS1, whose product induces cell death in N. benthamiana. Putative homologues of NIS1 are present in selected members of fungi belonging to class Sordariomycetes, Dothideomycetes, or Orbiliomycetes. Green fluorescent protein-based expression studies suggested that NIS1 is preferentially expressed in biotrophic invasive hyphae. NIS1 lacking signal peptide did not induce NIS1-triggered cell death (NCD), suggesting apoplastic recognition of NIS1. NCD was prevented by virus-induced gene silencing of SGT1 and HSP90, indicating the dependency of NCD on SGT1 and HSP90. Deletion of NIS1 had little effect on the virulence of C. orbiculare against N. benthamiana, suggesting possible suppression of NCD by C. orbiculare at the postinvasive stage. The CgDN3 gene of C. gloeosporioides was previously identified as a secreted protein gene involved in suppression of hypersensitive-like response in Stylosanthes guianensis. Notably, we found that NCD was suppressed by the expression of a CgDN3 homologue of C. orbiculare. Our findings indicate that C. orbiculare expresses NIS1 at the postinvasive stage and suggest that NCD could be repressed via other effectors, including the CgDN3 homologue.     

Effects of interleukin-8 on estradiol and progesterone production by bovine granulosa cells from large follicles and progesterone production by luteinizing granulosa cells in culture

Interleukin 8 (IL-8) is a chemoattractant involved in the recruitment and activation of neutrophils and is associated with the ovulate process. We examined the possible role of IL-8 in steroid production by bovine granulosa cells before and after ovulation. The concentration of IL-8 in the follicular fluid of estrogen-active dominant (EAD) and pre-ovulatory follicles (POF) was higher than that of small follicles (SF). CXCR1 mRNA expression was higher in the granulosa cells of EAD and POF than that of SF. In contrast, CXCR2 mRNA expression was lower in granulosa cells of EAD and POF than in SF. IL-8 inhibited estradiol (E2) production in follicle-stimulating hormone (FSH)-treated granulosa cells at 48 h of culture. IL-8 also suppressed CYP19A1 mRNA expression in FSH-treated granulosa cells. IL-8 stimulated progesterone (P4) production in luteinizing hormone (LH)-treated granulosa cells at 48 h of culture. Although IL-8 did not alter the expression of genes associated with P4 production, it induced StAR protein expression in LH-treated granulosa cells. The expression of CXCR1 mRNA in corpus luteum (CL) did not change during the luteal phase. In contrast, the expression of CXCR2 mRNA in middle CL was significantly higher than in early and regression CL during the luteal phase. In luteinizing granulosa cells, an in vitro model of granulosa cell luteinization, CXCR2 mRNA expression was downregulated, whereas CXCR1 mRNA expression was unchanged. IL-8 also stimulated P4 production in luteinizing granulosa cells. These data provide evidence that IL-8 functions not only as a chemokine, but also act as a regulator of steroid synthesis in granulosa cells to promote luteinization after ovulation.