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Mar. Drugs 2013, 11(1), 81-98; https://doi.org/10.3390/md11010081
Fucoidan Extract Enhances the Anti-Cancer Activity of Chemotherapeutic Agents in MDA-MB-231 and MCF-7 Breast Cancer Cells
Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan
Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan
Yoshida Clinic, 6-18-27 Higashi Mikuni, Yodogawa-ku, Osaka 532-0002, Japan
Daiichi Sangyo Co., Ltd., 6-7-2 Nishitenman, Kita-ku, Osaka 530-0037, Japan
Author to whom correspondence should be addressed.
Received: 12 November 2012; in revised form: 13 December 2012 / Accepted: 14 December 2012 / Published: 9 January 2013
Fucoidan, a fucose-rich polysaccharide isolated from brown alga, is currently under investigation as a new anti-cancer compound. In the present study, fucoidan extract (FE) from Cladosiphon navae-caledoniae Kylin was prepared by enzymatic digestion. We investigated whether a combination of FE with cisplatin, tamoxifen or paclitaxel had the potential to improve the therapeutic efficacy of cancer treatment. These co-treatments significantly induced cell growth inhibition, apoptosis, as well as cell cycle modifications in MDA-MB-231 and MCF-7 cells. FE enhanced apoptosis in cancer cells that responded to treatment with three chemotherapeutic drugs with downregulation of the anti-apoptotic proteins Bcl-xL and Mcl-1. The combination treatments led to an obvious decrease in the phosphorylation of ERK and Akt in MDA-MB-231 cells, but increased the phosphorylation of ERK in MCF-7 cells. In addition, we observed that combination treatments enhanced intracellular ROS levels and reduced glutathione (GSH) levels in breast cancer cells, suggesting that induction of oxidative stress was an important event in the cell death induced by the combination treatments.
Keywords:fucoidan extract; chemotherapeutic agents; anti-cancer activity; apoptosis; breast cancer
1. Introduction
Natural dietary compounds have been widely and safely consumed over centuries, and preclinical studies suggest that most of them have potential applications in pharmacology and cancer therapy. Fucoidan is a naturally occurring polysaccharide compound in brown alga, such as Fucus vesiculosus, Cladosiphon okamuranus and Laminaria saccharina [1,2,3]. Studies in vitro have indicated that fucoidan provides protection against various cancers, including human lymphoma, promyelocytic leukemia, colon carcinoma, breast carcinoma, hepatoma and melanoma [4,5,6,7,8,9]. It was found that fucoidan inhibits angiogenesis of melanoma, and it has anti-metastatic activity against Lewis lung adenocarcinoma and 13762 MAT rat mammary adenocarcinoma in mouse xenograft models [10,11,12]. Clinical studies have shown that fucoidan causes tumor regression and subjective improvement of overall survival in cancer patients [13]. These findings confirm the efficacy of fucoidan against human cancers. Fucoidan exerts pleiotropic effects on cancer cells involving the induction of apoptosis through caspase-cascade activation, regulation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38 signaling, Bcl-2 protein expression and Akt signaling, inhibition of angiogenesis by suppressing vascular endothelial growth factor expression and inhibition of cell transformation induced by epidermal growth factor receptor [2,4,5,6,7,14,15]. In our previous studies, we showed that fucoidan extract (FE) induced apoptosis in MCF-7 cancer cells via ROS-dependent JNK activation and a mitochondrion-mediated pathway [8]. Since fucoidan has anti-tumor properties both in vitro and in vivo, a combination of fucoidan with chemotherapeutic drugs might be an intriguing option in the therapy of cancer patients. Ikeguchi et al. [16] have performed a clinical trial in patients with unresectable advanced or recurrent colorectal cancer. The patients who received 150 mL/day of fucoidan were able to endure prolonged chemotherapy without fatigue. The survival of patients with fucoidan treatment was longer than that of patients without fucoidan treatment, although the difference was not significant [16]. Therefore, the application of combination approaches involving chemotherapeutic agents could improve drug absorption and enhance the clinical response.
Low molecular weight FE was used in this study, which was obtained by enzymatic digestion of a high molecular weight FE purified from Cladosiphon navae-caledoniae Kylin. The digested low molecular weight FE is more water-soluble than undigested high molecular weight FE, which affects absorption in vivo and, thus, bioavailability [17,18,19,20]. Cisplatin (CDDP) is a widely used chemotherapeutic agent for various types of cancers. It has been confirmed that CDDP exerts its cytotoxicity by interference with transcription or DNA replication mechanisms, leading to cell cycle checkpoint activation and sustained G2 arrest [21]. CDDP has been reported to cause apoptosis mediated by the activation of distinct signal pathways, including death receptor signaling, mitogen-activated protein kinases (MAPKs) signaling, protein kinase Akt signaling, p53 signaling and the activation of mitochondrial pathways [22]. Tamoxifen (TAM) is a selective estrogen receptor (ER) antagonist that is extensively used in the treatment of both advanced-stage and early-stage estrogen receptor-positive breast cancers [23]. Clinical response to TAM has been shown to be associated with both decreased proliferation and increased apoptosis. Mechanisms associated with apoptosis have been described, including production of oxidative stress and activation/expression of modulation proteins, such as ERK and JNK, transforming growth factor-β, protein kinase C, as well as Bcl-2 protein family members [24]. Paclitaxel (TAXOL), a natural chemotherapeutic drug isolated from the bark of the pacific yew, is currently used in the treatment of breast cancer and ovarian cancer. TAXOL-treated cancer cells undergo cell cycle arrest and apoptosis [25]. The activities of TAXOL have been described and include effects on cell signaling and gene expression, activation of MAPKs, Raf-1, protein tyrosine kinases and regulation of Bcl-2-related proteins, such as Bcl-2, Bcl-xL and Bad [26,27].
The data presented here show that low molecular weight FE in combination with CDDP, TAM or TAXOL significantly enhanced cell death of MDA-MB-231 and MCF-7 breast cancer cells by regulating the expression of Bcl-2 family proteins, modulating ERK and Akt signaling and regulating the production of oxidative stress.
2. Results and Discussion
2.1. Enhanced Cytotoxicity by Combination of FE and Chemotherapeutic Agents
MDA-MB-231 and MCF-7 breast cancer cells were exposed to FE or FE plus one of the three commonly used chemotherapeutic agents, namely, CDDP, TAM or TAXOL. In the absence of chemotherapeutic agents, FE exhibited a dose-dependent cytotoxicity to the cells (Figure 1). MCF-7 cells were much more sensitive than MDA-MB-231 cells. Among the three agents, MCF-7 cells were relatively resistant to CDDP treatment and relatively sensitive to TAM treatment compared to MDA-MB-231 cells. In the presence of 400 μg/mL FE, more than 80% cell growth inhibition was induced by 10 μM CDDP in both MDA-MB-231 and MCF-7 cells. FE also enhanced TAM-induced (20 μM) cell growth inhibition from 29% to 84% in MDA-MB-231 cells. When FE was combined with TAXOL, inhibition of MCF-7 cell growth was similar to that of MDA-MB-231 cells. The synergistic effect of FE on chemotherapeutic agent-induced cell proliferation reduction was dose-dependent.
Figure 1. Synergistic inhibition of cell growth. Cell proliferation was analyzed using the MTT assay. Cells were treated with fucoidan extract (FE), Cisplatin (CDDP), Tamoxifen (TAM) or Paclitaxel (TAXOL) alone or in combination for 48 h (n = 5). Results represent the mean ± SD from three independent experiments. A significant difference from control is indicated by p < 0.05 (#) or p < 0.01 (##); a significant difference from single treatments is indicated by p < 0.05 (*) or p < 0.01 (**).
2.2. Synergistic Induction of Apoptosis and Cell Cycle Arrest by FE and Chemotherapeutic Agents
To determine if the combination treatment-induced cytotoxicity occurred through necrosis and/or apoptosis, cells were assayed with Hoechst 33342 staining and annexin V-FITC staining. As shown in Figure 2A, in untreated cells, nuclear chromatin was homogeneously dispersed throughout the whole nucleus. When combining FE with chemotherapeutic agents, cells exhibited a shrunken nucleus, peripheral clumping and nuclear fragments with bright chromatin. Enhanced apoptosis of tumor cells by the combination treatments was evident by an increased number of annexin V-positive cells in a time-dependent manner (Figure 2B). We found that 200 μg/mL FE enhanced 5 μM CDDP-induced apoptosis from 32.9% to 52.4% in MDA-MB-231 cells and from 20.4% to 47.6% in MCF-7 cells. Combining FE with TAM (10 μM) enhanced cell death from 17.9% to 57.9% in MDA-MB-231 cells and from 31% to 66% in MCF-7 cells after 48 h of treatment. Apoptotic cell death increased about two-fold over 48 h in the presence of both FE and TAXOL (2.5 nM) relative to TAXOL alone in both MDA-MB-231 cells and MCF-7 cells.
Figure 2. Synergistic induction of apoptosis by co-treatment. (A) Hoechst 33342 staining of cells treated with 200 μg/mL FE alone or 200 μg/mL FE in combination with 5 μM CDDP, 10 μM TAM or 2.5 nM TAXOL for 48 h. Each experiment shown is representative of 20 randomly observed fields; (B) Analysis of apoptotic cells by annexin/PI double-staining using the IN Cell Analyzer 1000. MDA-MB-231 and MCF-7 cells were treated for different times with 200 μg/mL FE alone or 200 μg/mL FE in combination with 5 μM CDDP, 10 μM TAM or 2.5 nM TAXOL. All results were obtained from three independent experiments. A significant difference from control is indicated by p < 0.05 (#) or p < 0.01 (##); a significant difference from single treatments is indicated by p < 0.05 (*) or p < 0.01 (**).
We next performed flow cytometry analysis to investigate changes in cell cycle distribution upon co-treatment. As shown in Table 1, CDDP alone or in combination with FE strongly induced the accumulation of MDA-MB-231 and MCF-7 cells in the G2/M phase. The combination of both FE and TAM slightly increased the number of MCF-7 cells in the G1 phase and had little effect on the cell cycle distribution in MDA-MB-231 cells. TAXOL or combination treatment in both cell lines caused an increase in the number of cells in the G2/M phase. The percentage of cells in the sub-G1 phase was significantly increased by the three co-treatments in the two cell lines, suggesting that the combination of two agents induced cell death more effectively than any single agent did.
Table 1. Analysis of cell cycle distribution. Cells were treated with FE, CDDP, TAM or TAXOL alone or in combination with FE for 48 h. Cells were then measured by flow cytometry as described in the Methods section. The cell distribution was analyzed by flow cytometry analysis software.
|65.9 ± 2.8
|21.9 ± 2.0
|12.2 ± 1.8
|1.9 ± 0.2
|54.6 ± 5.2
|31.7 ± 2.5
|13.7 ± 0.5
|5.0 ± 0.2
|FE (200 μg/mL)
|68.7 ± 1.0
|18.8 ± 1.6
|12.5 ± 0.9
|10.7 ± 0.8
|57.3 ± 4.8
|27.5 ± 1.5
|15.2 ± 1.2
|13.6 ± 1.0
|CDDP (5 μM)
|13.7 ± 0.3
|29.4 ± 3.5
|56.9 ± 3.2
|17.5 ± 4.3
|37.1 ± 1.4
|14.1 ± 1.0
|48.8 ± 1.5
|8.9 ± 1.5
|19.9 ± 0.7
|21.4 ± 0.6
|58.7 ± 0.4
|37.8 ± 6.2
|26.2 ± 2.5
|29.0 ± 2.6
|44.8 ± 2.0
|32.4 ± 3.6
|TAM (10 μM)
|66.6 ± 3.6
|23.3 ± 2.1
|10.1 ± 1.6
|4.9 ± 0.6
|63.7 ± 2.2
|18.9 ± 1.5
|17.4 ± 0.9
|12.6 ± 0.9
|60.8 ± 2.0
|22.7 ± 1.5
|16.5 ± 1.5
|18.3 ± 2.1
|67.8 ± 2.4
|17.6 ± 1.6
|14.6 ± 1.1
|46.5 ± 4.3
|TAXOL (2.5 nM)
|54.5 ± 2.0
|20.5 ± 1.5
|25 ± 0.8
|6.4 ± 0.4
|54.2 ± 2.2
|16.9 ± 1.3
|29.1 ± 0.8
|8.4 ± 0.6
|53.7 ± 1.9
|22.1 ± 2.1
|24.2 ± 1.6
|25.8 ± 2.4
|49.8 ± 2.6
|21.8 ± 1.0
|28.4 ± 0.7
|20.6 ± 0.3
2.3. Synergistic Changes in the Expression of Bcl-2 Proteins
To investigate the cell death mechanism induced by the combination treatments, Western blot analysis of the breast cancer cells was performed. Our previous experiments revealed that FE affected the regulation of expression of Bcl-2 family proteins [8]. Therefore, we first tested whether co-treatment with chemotherapeutic agents could further regulate the effect of FE on the expression of Bcl-2 proteins. The data in Figure 3 show that 200 μg/mL FE alone efficiently reduced the expression of the anti-apoptotic proteins Bcl-xL and Mcl-1 in both MDA-MB-231 and MCF-7 cells. In combination, CDDP, TAM and TAXOL further reduced Bcl-xL and Mcl-1 expression. Significant up-regulation of the expression of the pro-apoptotic protein Bax was observed only in MCF-7 cells and was observed only with FE, CDDP or combination treatment.
Figure 3. Regulation of expression of Bcl-2 proteins by combination treatments. MDA-MB-231 and MCF-7 cells were treated with 200 μg/mL FE or 200 μg/mL FE in combination with 5 μM CDDP, 10 μM TAM or 2.5 nM TAXOL for 48 h. The expression of Bcl-xl, Mcl-1 and Bax was determined by Western blotting using specific antibodies. Experiments were conducted three times; representative blots are shown.
2.4. Modulations in ERK and Akt Phosphorylation
We next determined the effects of co-treatment on the levels of survival receptor ERK. The data in Figure 4A,B show that combination treatments partially decreased the phosphorylation of ERK in MDA-MB-231 cells compared with untreated controls or FE treatment alone. Unexpectedly, an obvious increase in ERK phosphorylation was observed by the addition of FE in MCF-7 cells; this change was further increased by combination treatments. Among the three drugs, FE in combination with TAM resulted in a four-fold increase in ERK phosphorylation, compared to the untreated control. To evaluate the role of ERK phosphorylation in the induction of cell death, MCF-7 and MDA-MB-231 cells were treated with 50 μM PD98059 prior to their exposure to the agents (Figure 4C). We found that PD98059 not only increased the extent of cell death induced by CDDP, TAM or TAXOL alone, but also increased the cell death caused by combination treatment in MDA-MB-231 cells. Interestingly, in the presence of PD98059, the combination of FE and CDDP, TAM or TAXOL resulted in partially decreased levels of cell death in MCF cells. A combination of FE and TAM induced an approximately 50% increase in cell viability.
Figure 4. Modulation of ERK phosphorylation by combination treatments. (A) MDA-MB-231 and MCF-7 cells were treated with FE or FE in combination with CDDP, TAM or TAXOL for 48 h. ERK1/2 and its phosphorylated forms were determined by Western blotting using specific antibodies; (B) The relative amount of protein expression was quantified using Quantity One software (Bio-Rad) and normalized by the band intensity of total ERK; (C) MDA-MB-231 and MCF-7 cells were pretreated with 50 μM PD98059 for 1 h and then exposed to FE or FE in combination with CDDP, TAM or TAXOL for 48 h. After treatment, MTT was used to analyze cell survival. All results were obtained from three independent experiments. Differences with p < 0.05 (*) or p < 0.01 (**) were considered statistically significant.
Activation of the Akt signaling pathway has been reported to mediate drug resistance and promote survival of cancer cells [28]. Therefore, we examined the status of Akt after drug treatment. As shown in Figure 5, FE alone produced an increase in the level of Akt phosphorylation in MCF-7 cells, with little change in Akt phosphorylation in MDA-MB-231 cells. Combination of FE plus CDDP, TAM or TAXOL exhibited inhibition of Akt phosphorylation compared with FE treatment alone in MDA-MB-231 cells. Phosphorylation of Akt was suppressed to approximately 50% by treatment of the cells with CDDP plus FE relative to FE treatment alone (Figure 5B).
Figure 5. Modulation of Akt phosphorylation by combination treatments. (A) MDA-MB-231 and MCF-7 cells were treated with FE or FE in combination with CDDP, TAM or TAXOL for 48 h. Akt and its phosphorylated forms were determined by western blotting using specific antibodies; (B) The relative amount of protein expression was quantified using Quantity One software and normalized by the band intensity of total Akt.
2.5. Synergistic Increases in ROS Levels and Reduction in GSH Levels
Our previous study showed that FE induced ROS generation in MCF-7 cells. To examine the effect of combination treatments on ROS generation, DCFH-DA was used as a molecular probe. As shown in Figure 6A, treatment of MDA-MB-231 cells with FE alone had a slight impact on ROS generation, similar to that in MCF-7 cells. The combination treatment with FE and the chemotherapeutic agents led to enhanced generation of ROS in the two cell lines. To determine whether ROS contributed to cell death, we tested the effect of NAC on drug-treated cancer cells (Figure 6B). As expected, pretreatment with NAC partially attenuated the cell cytotoxicity induced by FE in combination with chemotherapeutic agents in MDA-MB-231 cells. However, in MCF-7 cells, significant inhibition of cell cytotoxicity was observed only with a combination of FE and CDDP after treatment with NAC.
Oxidative stress can be facilitated by reduced intracellular GSH levels that promote apoptosis. Therefore, we measured total GSH levels in the breast cancer cells exposed to FE or in combination with chemotherapeutic agents (Figure 7A). A slight, but statistically significant, reduction in GSH levels of 10.5% was observed for FE-treated MCF-7 cells. FE in combination with the chemotherapeutic agents enhanced the reduction of GSH levels more than chemotherapeutic agents alone in the two cell lines. Treatment with the combination of FE and TAM further reduced GSH levels by up to 31.5% in MDA-MB-231 cells and by 33% in MCF-7 cells. Furthermore, the intracellular GSH content was replenished by the addition of externally supplied GSH. As shown in Figure 7B, in the presence of 5 mM GSH, a large increase in cell viability was observed in the cells treated with a combination of FE and chemotherapeutic agents.
Figure 6. Synergistic increases in ROS levels. (A) MDA-MB-231 and MCF-7 cells were treated with FE or FE in combination with CDDP, TAM or TAXOL for 48 h. Then, the cells were labeled with 5 μM DCFH-DA and the fluorescence was measured using the IN Cell Analyzer 1000, as described in the Methods section; (B) MDA-MB-231 and MCF-7 cells were pretreated with 5 mM NAC for 1 h and then exposed to FE or FE in combination with CDDP, TAM or TAXOL for 48 h. After treatment, MTT was used to analyze cell survival. All results were obtained from three independent experiments. Differences with p < 0.05 (*) or p < 0.01 (**) were considered statistically significant.
Figure 7. Synergistic reduction of GSH levels. (A) MDA-MB-231 and MCF-7 cells were treated with FE or FE in combination with CDDP, TAM or TAXOL for 48 h. Total GSH levels were quantified using the GSH Assay Kit (Trevigen Inc., Gaithersburg, MD, USA); (B) MDA-MB-231 and MCF-7 cells were incubated with FE or FE in combination with CDDP, TAM or TAXOL for 48 h in the presence or absence of 5 mM GSH; cell survival was analyzed using the MTT assay. All results were obtained from three independent experiments. Differences with p < 0.05 (*) or p < 0.01 (**) were considered statistically significant.
2.6. Global Discussion
A growing body of evidence indicates that fucoidans function as prospective anti-cancer agents in vitro, as well as in vivo. However, the in vivo concentration achieved after oral administration of fucoidan is much lower than the concentration that demonstrated efficacy in vitro [29]. Although the reported efficacy of fucoidan may be achieved by the application of higher oral or intravenous doses, it has the potential to be an anti-cancer drug based on its synergistic activity with other chemotherapeutic drugs.
In the present study, we investigated the combination of FE and three chemotherapeutic drugs, which exhibit highly synergistic inhibition effects on cancer cell growth, thus allowing the effective concentration of FE to be reduced relative to our previous study. We noted that FE significantly enhanced cell proliferation reduction induced by CDDP, TAM or TAXOL alone in MDA-MB-231 and MCF-7 breast cancer cells. Morphological changes of the nucleus and the number of time-dependent annexin V-positive cells increased, indicating that the combination treatments caused apoptotic cell death. FE itself did not arrest cell cycle progression in MCF-7 cells in our previous study [8]. However, the combination treatment induced modifications in the cell cycle distribution in both MDA-MB-231 and MCF-7 cells, suggesting additive or synergistic effects occurred with the co-treatment.
In this study, the combination treatments relative to FE or chemotherapeutic drugs alone partially reduced Bcl-xL and Mcl-1 expression in MDA-MB-231 and MCF-7 cells. It has been reported that concomitant downregulation of Mcl-1 and Bcl-xL is sufficient to induce massive cell death in human mesothelioma cells [30], and reduction of Bcl-xL expression caused reduction of tumor growth in vivo, as well as increased survival in mouse models [31]. Furthermore, pro-apoptotic Bak is sequestered by Mcl-1 and Bcl-xL; therefore, inactivation of these proteins by BH3-only proteins is necessary and sufficient for Bak-mediated cell death [32]. These findings indicate that Mcl-1 and Bcl-xL downregulation may serve as a convergence point for inducing massive apoptotic cell death and avoiding therapeutic failure of FE in combination with chemotherapeutic drug treatments. In addition, our previous studies showed that FE treatment induced mitochondrion-dependent apoptosis in MCF-7 cells, as demonstrated by the regulation of Bcl-2 proteins and the release of cytochrome c and AIF. The results presented in this study regarding regulation of Mcl-1, Bcl-xL and Bax expression in cells treated by the combination therapy is likely to cause the induction of Bcl-2 protein-mediated, mitochondrion-dependent cell death. Further experiments will be required to determine if the mitochondria is the major target for the FE-enhanced increased efficacy of chemotherapy.
ERK activation induced by fucoidan has been shown to contribute to the signal transduction pathways leading to cell apoptosis [4,6,15]. We have previously demonstrated that ERK phosphorylation occurred rapidly and persisted for a period after FE exposure in MCF-7 cells [8]. The present study characterized the effects of a combination of FE and chemotherapeutic agents on cell death with respect to ERK activation. Surprisingly, our data showed that a combination of FE and chemotherapeutic agents decreased ERK phosphorylation induced by FE in MDA-MB-231 cells; however, it was increased in MCF-7 cells. Furthermore, pretreatment with PD98059 promoted cell death in MDA-MB-231 cells treated by combination therapy, but blocked cell death in similarly treated MCF-7 cells. These results suggest that a combination of FE and chemotherapeutic agents modified the ERK activation induced by FE and demonstrated different effects on transduction pathways in MDA-MB-231 cells and MCF-7 cells. It has been reported that fucoidan exerts activation of the ERK1/2 signaling pathway associated with cell apoptosis in HL-60 cells and HCT-15 cells [6,15]. On the other hand, the effect of fucoidan on ERK inhibition was observed in HS-Sultan cells [4]. These findings indicate that it is likely that chemotherapeutic agents exhibit various synergistic effects on cell death due to cell type-specific factors. MEK/ERK activity has been shown to act on mitochondrion-dependent apoptosis by modulating expression of Bcl-2 family proteins, such as Bax and Bcl-xL, and they regulate cell cycle arrest associated with cyclin D1 transcription and cyclin B/cdc2 nuclear translocation [33,34,35]. Our studies concerning the involvement of cell cycle arrest and apoptosis in combination treatments indicated that ERK phosphorylation might mediate cell cycle arrest and apoptosis in breast cancer cells. In addition, mechanisms of estrogen-mediated cellular actions have been shown to be very complex [36]. TAM has been shown to activate ERK rapidly in ER-positive MCF-7 and T47D cells, but not significantly in ER-negative MDA-MB-231 cells, which demonstrates a correlation between activated ERK and ER expression in the induction of rapid death of breast cancer cells by TAM [37]. Doxorubicin has been shown to decrease the phosphorylation of ERK in ER-negative MDA-MB-231 cells; however, phosphorylation was increased in ER-positive MCF-7 cells [38]. It is possible that FE and in combination with anti-cancer drugs can activate different receptor and signaling pathways in ER-negative MDA-MB-231 cells compared to ER-positive MCF-7 cells.
There is abundant evidence that activation of Akt signaling mediates cancer cell growth and may serve as a potential target for drug development. Fucoidan has been reported to inhibit Akt phosphorylation in lymphoma HS-Sultan cells [4]. However, significant inhibition of Akt phosphorylation in FE-treated breast cancer cells was not observed in the present study. We noted that combination treatment involving FE and anti-cancer drugs reduced Akt phosphorylation in MDA-MB-231 cells, but had no effect in MCF-7 cells. Cross-talk between the Ras-Raf-MEK-ERK and PI3K-Akt pathways has been demonstrated in several cellular systems [34,39,40]. It is possible that disruption of Akt signaling has a greater impact on cell death under conditions of MEK/ERK inactivation, because the combination treatment resulted in decreased ERK phosphorylation in MDA-MB-231 cells. It has been reported that estrogen promotes association of ERK with the IGF-1 receptor and the p85 subunit of PI3-kinase in the plasma membrane, which leads to AKT activation [41,42]. In addition, estrogen-mediated Akt activation has been reported to be blocked by the estrogen antagonist ICI 182780 in ER-positive MCF-7 cells [43]. The discrepancy between the two cell lines might be attributed to differences in ER status. Inactivation of Akt seems to be important, at least in the apoptotic response of ER-negative breast cancer cells against combination treatments. Activation of Akt was observed in ER-positive MCF-7 cells, suggesting that FE might act as an ER agonist to stimulate Akt. This hypothesis suggests a negative impact of FE as a single drug on ER-positive breast cancer patients, but a potential positive interaction when combined with chemotherapeutic drugs for ER-negative breast cancer patients. However, further in vivo studies are required to address this possibility.
Cellular redox status or oxidative stress has been shown to be involved in cell proliferation, growth arrest or apoptotic cell death based on the extent of redox imbalance [44]. In our study, combination treatments somewhat enhanced ROS generation induced by FE in both MDA-MB-231 and MCF-7 cells. Pretreatment with NAC partially increased cell viability for all combination treatments in MDA-MB-231 cells, suggesting that ROS generation is related to cell death. Meanwhile, in MCF-7 cells, significant inhibition of cell cytotoxicity failed to be observed with the combination of FE and TAM or the combination of FE and TAXOL after treatment with NAC. It is possible that ROS generation is not necessary for cell death by these two combination treatments in MCF-7 cells. High intracellular GSH levels have been associated with apoptotic resistance, and GSH depletion itself has been found to trigger cell death cascades [45]. We observed that GSH was further reduced with combination treatment relative to FE treatment alone. The addition of external GSH to the medium protected against cell death. These findings suggest that oxidative stress is involved in the progression of cell death caused by a combination of FE with the three chemotherapeutic agents. In addition, early GSH depletion has been reported to cause dissipation of the ΔΨm with simultaneous ROS generation prior to induction of apoptosis [46]. We previously found that ROS is involved in mitochondria dysfunction in FE-treated MCF-7 cells [8]. It is likely that generation of intracellular ROS and depletion of GSH are related to the induction of mitochondria-dependent cell death in breast cancer cells. Thus, these findings support the hypothesis that oxidative stress plays a role as a common mediator of cell death by treatment with a combination of FE and chemotherapeutic agents.
3. Experimental Section
3.1. Cell Culture
Estrogen receptor (ER)-positive MCF-7 and ER-negative MDA-MB-231 cells were obtained from the American Type Culture Collection (Manassas, VA, USA). Cells were maintained in DME medium supplemented with 10% fetal bovine serum in an incubator in a humidified atmosphere of 5% CO2 at 37 °C.
3.2. Materials
The abalone glycosidase-digested FE was generously donated for this study by the Daiichi Sangyo Corporation (Osaka, Japan). FE was prepared as previously described [2]. Briefly, high molecular weight fucoidan extract was purified to 85% from seaweed of Cladosiphon navae-caledoniae Kylin and digested with glycosidases to obtain FE. FE consists of a digested small molecular weight fraction (72%, MW < 500 Da) and non-digested fractions (less than 28%, peak MW: 800 kDa). FE consisted of mostly fucose (73%), xylose (12%) and mannose (7%). The ratio of sulfation was 14.5%.
Cisplatin, tamoxifen citrate, paclitaxel, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and propidium iodide (PI) were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). Cellstain®-Hoechst 33342 was obtained from DOJINDO Laboratories (Kumamoto, Japan). The annexin V-FITC apoptosis kit was obtained from Medical and Biological Laboratories Co., Ltd. (Nagoya, Japan). PD98059 and 2′,7′-dichlorofluorescin diacetate (DCFH-DA) were purchased from Merck KGaA (Darmstadt, Germany). The ROS inhibitor NAC was purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). The GSH Assay Kit was purchased from Trevigen Inc. (Gaithersburg, MD, USA). All antibodies used in the study were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA).
3.3. MTT Assay
Cells were seeded in 96-well plates at densities of 2000 cells/well for 24 h. The cells were then incubated with FE or chemotherapeutic drugs plus FE. After 48 h of incubation, 10 μL of MTT solution (5 mg/mL) was added to each well and incubated for an additional 4 h at 37 °C. The medium was aspirated and replaced with 200 μL/well of DMSO to dissolve the formazan salt. The color intensity of the formazan solution was measured at 595 nm using a microplate reader (Tecan Group Ltd., Männedorf, Switzerland).
3.4. Hoechst Staining
Cells were seeded onto 96-well plates (5 × 103 cells/well) and cultured for 24 h. After 48 h of treatment with drugs, cells were washed with PBS and stained with 2 μg/mL Hoechst 33342 for 15 min. Images were then captured with the IN Cell Analyzer 1000 (GE Healthcare UK Ltd., Buckinghamshire, UK). Each image shown is representative of 20 randomly observed fields.
3.5. Annexin V Binding Assay
FITC-labeled annexin V was applied and analyzed by the IN Cell Analyzer 1000. After drug treatment, cells were stained with the annexin V-FITC solution (annexin V-fluorescein in binding buffer containing PI and Hoechst 33342). The cells were further incubated for 15 min in the dark, and then, images of the cells were acquired using the IN Cell Analyzer 1000. Cells with apoptotic morphology of the nuclei (condensation/fragmentation) or annexin V-positive cells were analyzed using the Developer Toolbox software (GE Healthcare UK Ltd.). For each analysis, 3000 cells were recorded.
3.6. Flow Cytometry Analysis
Cell cycle distribution was performed by flow cytometry. After treatment, the cells were collected by trypsin, washed in cold PBS and fixed with 70% ethanol on ice for 30 min. The fixed cells were washed with PBS and incubated in 10 μg/mL RNase A for 30 min at room temperature. PI (10 μL of 1 mg/mL) was added to the cell suspension and cells were incubated for 10 min in the dark. The DNA content was analyzed by flow cytometry (Beckman Coulter, Inc., Brea, CA, USA). The proportion of cells in G1, S and G2/M phases was determined using FlowJo software (Tree Star, Inc., Ashland, OR, USA). For each sample, 1 × 104 cells were recorded.
3.7. Western Blot Analysis
Cell protein lysates were prepared and Western blot analyses were performed as described previously [8]. The proteins were separated by SDS-polyacrylamide gel electrophoresis. The proteins were then transferred onto polyvinylidene difluoride membranes (GE Healthcare UK Ltd.) and blotted with each antibody. Protein bands were visualized using enhanced chemiluminescence, as described by the supplier (GE Healthcare UK Ltd.).
3.8. Measurement of Intracellular ROS
To detect generation of intracellular ROS, cells were incubated with drugs for 5 h and labeled with 5 μM DCFH-DA at 37 °C for 30 min. Then, the cellular fluorescence intensity was measured with the IN Cell Analyzer 1000 after washing the cells twice with PBS. For each sample, 3000 events were recorded. Image analysis was carried out using the IN Cell Investigator software using either the Developer Toolbox or the Multi-Target Analysis (MTA) Module (GE Healthcare UK Ltd.).
3.9. GSH Assay
The GSH concentration was measured by the GSH Assay Kit, according to the manufacturer's instructions. Briefly, the drug-treated cells were collected in cold 5% metaphosphoric acid. The samples were sonicated and then incubated on ice for 5 min. The suspension was then centrifuged at 12,000× g for 5 min at 4 °C. Total GSH content was measured with a microplate reader at 405 nm.
3.10. Statistical Analysis
Each experiment was performed at least in triplicate and repeated three times. The results are presented as the mean ± standard deviation (SD) values. The difference between the two groups was analyzed using the two-tailed Student's t-test; differences among three or more groups were analyzed by one-way analysis of variance multiple comparisons.
4. Conclusions
In conclusion, the present study demonstrates that FE, an enzymatically digested polysaccharide, cooperates with the chemotherapeutic agents CDDP, TAM and TAXOL to induce cell growth inhibition through the induction of apoptosis and cell cycle arrest in the human MDA-MB-231 and MCF-7 breast cancer cell lines. These studies highlight the potential regarding the achievable efficacy of FE in combination with chemotherapeutic agents in cancer treatment. Further in vivo and clinical studies are needed to evaluate the safety and utility of these combination treatments in cancer patients.
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- Samples Availability: Available from the authors.
© 2013 by the authors; licensee MDPI, Basel, Switzerland. This article is an open-access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).


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You are here: Who we are > Management team
Rachel Cook
Client Assurance Director
Rachel joined Capita in May 2011 via the acquisition of Red Procurement where she was the Operations Director, and has subsequently become accountable for Client Assurance which includes the accountability for the People Strategy and Resource Landscape and Delivery Assurance across Capita Procurement Solutions. In addition to this, she has led a number of a client transformation programmes deploying her key skills in organisational change, process improvement, project management and benefits realisation.
Rachel has gained her procurement expertise over the last 16 years having held both procurement leadership and delivery roles working across both private and public sectors having worked both within consultancy and in house procurement teams.
In her spare time, Rachel enjoys spending time with her young family, as well as travelling and spending time outdoors walking and running.All staff

subset: commoncrawl


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Electronic - Get The Message Chords & Tabs
Get The Message Chords & Tabs
#----------------------------------PLEASE NOTE---------------------------------# #This file is the author's own work and represents their interpretation of the # #song. You may only use this file for private study, scholarship, or research. # #------------------------------------------------------------------------------## From uunet!mcsun!uknet!bradford.ac.uk!N.J.Dixon Sun Oct 25 13:45:47 PST 1992 Article: 3062 of alt.guitar.tab Path: nevada.edu!uunet!mcsun!uknet!bradford.ac.uk!N.J.Dixon From: firstname.lastname@example.com (NJ DIXON) Newsgroups: alt.guitar.tab Subject: CRD: Get The Message (ELECTRONIC) Message-ID: <firstname.lastname@example.com> Date: 20 Oct 92 12:44:48 GMT Organization: University of Bradford, UK Lines: 46 Originator: 90468713@ccw204 Nntp-Posting-Host: ccw204 X-Newsreader: TIN [version 1.1 PL6] Well I'm no musical genius, but I think it's like this: INTRO: D | C | BbM7 | BbM7 C {repeat this a few times} VERSE: [ Tab from: https://www.guitartabs.cc/tabs/e/electronic/get_the_message_crd.html ] |D |G |BbM7 | | I've always thought of|you as my re-|-ward, built like an| |C |G |G |C D |D | |angel, six feet|tall.| |Six feet tall.| | {etc} CHORUS: |D |G |BbM7 | I|don't know where to be-|-gin, living in|sin, how can you| |C | G |D |G | |talk?| Look where you've|been. I've counted the|nights, living in| |G |BbM7 C |D |sin, how can you|talk? Look where we've|been. Basically, the song's made up of those four chords, played like this: D G C Bbmaj7 e--2------3-----0--------1------| B--3------0-----1--------3------| G--2------0-----0--------2------| D--0------0-----2--------3------| A---------2-----3---------------| E---------3---------------------| I've probably got this all wrong, but it sounds OK to me. Have fun! - Nick


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By Darlene Senger, Candidate for Congress, IL-11th
In the last few weeks I have toured the 11th Congressional District meeting with business owners, managers and employees to talk about the current economic climate in our community. What I heard was both uplifting and very concerning.
Every day there are amazing people who decide to take a risk and start their own business. It is a tireless and often thankless task, one that leads to many sleepless nights worrying about paying bills and making payroll. But these small business owners help drive our local and national economy by providing a service and creating jobs. They truly exemplify the spirit of the American Dream.
However these same people who are working towards the American Dream are being held back by the American government. Poor fiscal policies, a cumbersome tax code and oppressive government regulations have stymied our local economic growth. We can and must do better, and it starts with changing the culture in Washington DC.
Businesses large are small are worried about the out of control federal spending. For too long politicians have spent recklessly, leaving us with over $17 trillion in debt. We need a long-term balanced budget solution that brings stability and accountability to our spending so that businesses feel secure in their efforts to grow in America.
One of the major frustrations faced by business today is the 70,000 pages of federal income tax code. Job creators are spending more and more time making sure they are compliant with the latest tax policies and less time focused on growth. We need to lower the tax rates, eliminate the death tax and simplify the tax code. I want companies spending more on research and development and putting people to work instead of sending more tax lawyers to court.
All the business owners I talked were upset about new government regulations that were being placed on their businesses. From the federal healthcare requirements all the way down to local ordinances on merchants, business owners are being over-regulated. Government's culture of red tape is strangling our local businesses. It's time to cut the red tape and get Illinois back to work!
We live in a great region, filled with enterprising and innovative people who help make this country such a great place to live. Unfortunately, our politicians are failing us, leaving us with record high taxes and unemployment while jobs are disappearing. We need to start putting our country back to work by implementing the policies needed to chart a new direction. And that starts with electing people willing to make the hard decisions necessary to start moving towards a brighter more prosperous future, and that's why I am running for Congress.
Darlene Senger is the Republican candidate for the Illinois 11th Congressional District. Senger has lived in Naperville for 20 years and before that resided in Clarendon Hills. Darlene is a graduate of Purdue University and received her MBA from DePaul University, having worked her way through school. Darlene's children, Eric and Michelle are graduates of Waubonsie Valley High School and the University of Illinois.


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Auxo Now Available for iOS 5.1 & iOS 6 | Cydia
Auxo the awesome Cydia tweak which transforms the iOS app switcher has now been released for iOS 5.1 and above. You can read more about the tweak here.
The tweak was at first available only for iOS 6. But the developers quickly saw the demand for it on the iPhone 4S which can only be jailbroken at iOS 5.1.1. They very quickly worked to get it working on both versions of iOS.
The wait now is for the iPhone 5 to be jailbroken, which this tweak was primarily made for, because it has a larger screen.
The tweak is available on Cydia right now for $1.99.


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Ethanol vs. Biodiesel: Just the Facts
What are they?
Ethanol is "an alcohol product produced from corn, sorghum, potatoes, wheat, sugar cane, even biomass such as cornstalks and vegetable waste. When combined with gasoline, it increases octane levels while also promoting more complete fuel burning that reduces harmful tailpipe emissions such as carbon monoxide and hydrocarbons." (Abengoa Bioenergy)
Biodiesel is "a domestic, renewable fuel for diesel engines derived from natural oils like soybean oil, and which meets the specifications of ASTM D 6751." Or, for those of you who want a more technical definition, it is "a fuel comprised of mono-alkyl esters of long chain fatty acids derived from vegetable oils or animal fats..." (National Biodiesel Board)How are the produced?
According to the Renewable Fuels Association (the US ethanol trade organization), there are two processes for making ethanol: wet milling and dry milling. Each process is explained in detail on the RFA's site.
Biodiesel is made through a chemical process called transesterification "whereby the glycerin is separated from the fat or vegetable oil. The process leaves behind two products -- methyl esters (the chemical name for biodiesel) and glycerin (a valuable byproduct usually sold to be used in soaps and other products). (NBB)
Which one is better for the environment?
Both forms of biofuel have definite environmental advantages over petroleum-based gasoline and diesel fuel.
According to the RFA, "Ethanol contains 35% oxygen. Adding oxygen to fuel results in more complete fuel combustion, thus reducing harmful tailpipe emissions. Ethanol also displaces the use of toxic gasoline components such as benzene, a carcinogen. Ethanol is non-toxic, water soluble and quickly biodegradable."
Biodiesel, on the other hand, "is the only alternative fuel to have fully completed the health effects testing requirements of the Clean Air Act. The use of biodiesel in a conventional diesel engine results in substantial reduction of unburned hydrocarbons, carbon monoxide, and particulate matter compared to emissions from diesel fuel. In addition, the exhaust emissions of sulfur oxides and sulfates (major components of acid rain) from biodiesel are essentially eliminated compared to diesel." (NBB)
Which biofuel should I use?
Unless you already own or plan to buy a vehicle with a diesel engine (which is no small feat these days), ethanol-blended gasoline is likely your best option for biofuel use. Biodiesel, however, could provide a great environmental benefit to trucking fleets, which consist almost entirely of vehicles with diesel engines.
If you're a do-it-yourselfer, though, biodiesel can be made at home -- a real DIYer can even make his/her own biodiesel reactor with open-source plans available on the Internet. Ryan Thibodaux, a political blogger, recorded his own efforts to build a reactor, buy diesel cars and actually start producing the stuff at his blog The Higher Pie
Still have questions?
A quick Google search can provide you with plenty of information on biofuels. Both the Renewable Fuels Association and the Biodiesel Board have tons of educational materials on their respective sites. And, as reader Mat pointed out, Wikipedia has extensive entries on ethanol and biodiesel.
UPDATE: In response to Mike's question below, Gristmill has a handy chart showing estimated fuel yields of certain crops. Those numbers look like they tend to favor ethanol.


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It's Labour Day weekend and if you have kids and you live in Canada that means back to school is literally around the corner. It's that time of year where we all know we need to starting planning. Figuring out lunches, signing up for activities (late of course), and feeling like a hot mess for the next week or two. Or is that just me? While you are starting to make a schedule and organize your life now is the perfect time to add "Update Family Portrait" to your list. The leaves will soon be changing, adding beautiful colours to the background and now is the perfect time to book an outdoor photo session with your favourite photographer!
Here is some inspiration for those interested in doing a fall session!
Interested in booking with us? Shoot us an email, text us, or give us a call (information below)!
Have an awesome long weekend!
Love, LOCPS xoxo
Left of Centre Photography Services www.locps.com email@example.com
Get social with us!


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Congratulations, you found the ultimate groom, you got engaged, you have your wedding ring, and now the next big task at hand is finding your wedding dress. Some brides have envisioned walking down the aisle in their wedding gown since they can remember, just waiting for the right man to walk into their lives, while others never thought about getting married. Finding a wedding dress is a fabulous experience, yet may be stressful and overwhelming.
After 25 years in the wedding business as a wedding planner, finally decided to sit down and write this blog to get you to say YES to the DRESS in an informed and easy way, without settling!
Wedding planners (by the way, I am not "just a wedding planner", yet a master bridal consultant – currently 1 of 61 in the world), normally don't have input on the final wedding gown selection. Yet now, after dealing with bride's 911s on "my gown doesn't fit, they are going to have to sew in a side seam," or "I don't trust them for alterations after they botched up my wedding gown order," and so much more, it IS about the entire experience at the bridal shop, which is purchase, alterations, service after the sale, and happily ever after!
I am known as being one of the best in the industry for getting you in your wedding gown and making you look and feel fabulous on your wedding day! It may be all the years of TV experience working with different body types and no matter what, the show must go on!
WHAT SHOULD YOU REMEMBER WHEN SHOPPING FOR A WEDDING GOWN?  Three important words will help you when shopping for your wedding gown: fit, flatter, and budget.
WHEN SHOULD YOU START SHOPPING? Many brides start shopping as soon as they get engaged. Would highly recommend you start shopping at least 9 months before your wedding date. Consider most bridal shops take an average of 4 – 6 months (some may take longer) to get your gown in, then you add 2 months for alterations. Don't wait to start shopping 6 months prior to your wedding date to be told "if you would have started shopping 9 months ahead of time, we would have been able to get that gown for you, now it is too late to order the gown, or we could still order the gown if you are willing to pay rush charges."
WHAT IS ONE WEDDING GOWN ITEM YOU DEFINITELY WANT? Having a small pocket in your wedding gown is priceless to hold your lipstick for touch-ups! This may be included in your gown, or a small additional alteration item, yet will make your wedding day touch-ups a breeze!
WHO SHOULD YOU TAKE WITH YOU TO SHOP? Shop with your mom, and maid or matron of honor, they know you better than anyone! If you take too many people it gets overwhelming, and there may be too many opinions.
SHOULD YOU MAKE AN APPOINTMENT, OR JUST STOP IN? For the best customer service, make an appointment, and ask to be scheduled with the manager or another employee who knows their inventory, and will be able to make recommendations to you based on your body type and style of your wedding.
THE GOWN ON THE HANGER MAY LOOK BETTER ON YOU. When you make an appointment to try on gowns, ask for an appointment with the manager or another employee who knows their inventory. This person will have seen a zillion brides with your same body type in the gowns in their store. Knowledge is power. Allow this person to make suggestions to you on what may look best on your body type to accentuate your positives, and to take away from any negatives. If you are lucky, you are one of those brides with a fabulous body where every gown you try on looks amazing on you!
WHAT WEDDING DRESS WILL FIT THE SEASON AND STYLE OF MY WEDDING? If you are getting married in the hot Arizona summer, you will want a lightweight dress with airy fabrics and minimal layers to keep you from feeling hot and sticky. If you are getting married in the cooler months (most popular wedding months in Arizona are March, April, October, and November), you may want a heavier fabric like satin or taffeta with stylish wraps or jackets when the sun goes down. If you are having a church wedding vs. an outdoor garden wedding, these are two different styles. If you are having a destination wedding, you need to think about logistics on taking your wedding gown with you on your flight.
HOW MANY GOWNS IS "ENOUGH" TO TRY ON? When I got married, I tried on two wedding gowns, and purchased the second one I tried on. Yes, being in the business, I knew what I wanted, and knew where to buy, so it was easy! Some brides make a "day" of it, making appointments (normally 1.5 hours for one appointment) at multiple bridal shops. When you do the math, if you schedule 4 appointments, you are trying on gowns for 6 hours and are in transit for 2 hours. How much do you want to drive, and how many gowns do you want to try on? Do your research, each bridal shop has a different ambiance, style, and price range. Many bridal shops overlap in the "middle" zone, yet some are less expensive (normally meaning less customer service), and some are more expensive (carrying one of a kind gowns). Some may be GREAT at selling you a gown, yet how are their alterations and service AFTER the sale? This is important!
WHAT SHOULD YOU INCLUDE IN YOUR WEDDING GOWN BUDGET? A look, feel, and quality of a dress priced at $800 will be extremely different from the look, feel, and quality of a gown priced at $3,000. It will make a difference if you are planning to wear one gown for the ceremony and change gowns for your dancing. If you are purchasing one gown vs. two gowns, it makes a difference. If you are not finding gowns at a price you love, it may be time to add some to your gown budget (reduce your budget in another category) to get what you want, since you do get what you pay for. Include these items in your budget: the wedding gown, wedding shoes, veil, headpiece/comb/tiara/headband, alterations, and undergarments (bra, hosiery).
WHAT SHOULD I WEAR TO SHOP FOR WEDDING GOWNS? Wear something which is comfortable and easy to get in and out of for quick changes. Wear your hair in a way you are thinking of wearing it on the day of your wedding. The neckline of your gown will make all the difference in the world!
WHAT WEDDING DRESS FITS MY BODY TYPE THE BEST? There are 5 popular styles of wedding gowns. All are gorgeous, yet choosing the right one to accentuate your best assets and diminish areas you are more conscious of, no matter what your shape or size, may seem overwhelming.
1). A-LINE/PRINCESS. The A-line wedding gown features a shape which is fitted around the bodice (part of the gown above the waist) and flows out to the ground, resembling the outline of an uppercase A. This A-line dress flows from the bust to the hem with an unbroken line. Due to its classic and simple elegant style, the A-line dress is appropriate for any occasion, from a quiet backyard wedding to a traditional church wedding. Its length may range from above the knee to full length that flows into a large train. A princess cut is the most dramatic variation of the A-line gown. This version of the A-line dress emphasizes the style's clean lines by utilizing vertical panels that run from the hemline to the hem. The princess style provides elongating lines that create a slimming effect since your eye focuses on the length created by these long, smooth seams.
The A-line wedding dress is flattering on almost all body types. The long lines of the princess cut create the illusion of height on a shorter body, as well as slim down a rounded figure. The full skirt of the A-line may also hide a larger lower body, or create the illusion of curves on a narrow frame. With the A-line being versatile, simple modifications to the neckline, waistline, or fabric can accentuate your best features and hide those you're not so fond of.
2). EMPIRE. The empire wedding gown is defined by a raised waistline which sits just below the bust line, and the rest of the dress flows down to the hem. The empire style is very versatile and can accommodate a wide variety of formality.Â
The empire wedding dress is flattering on almost all body types. It is two fold on small busted brides and large busted brides. On small busted brides the empire wedding gown draws attention to your neckline and creates definition. Yet oddly enough, this also may work to conceal a large bust by controlling and minimizing the size of the breasts due to the neckline of the empire dress. The design of the empire dress is also ideal for covering other unwanted body features such as a long torso, short legs, or a pear shaped figure. Since the dress flows from the bust line, it easily leaves these areas undefined. The structure of the empire dress is also ideal for pregnant brides since the fluidity of the dress may accommodate a growing belly.
3). COLUMN/SHEATH. The column/sheath wedding gown has a narrow shape that flows straight down from the neckline to the hem. The column/sheath gown tends to hug the body and show any and all of your curves. This column/sheath gown is appropriate for all types of weddings, especially casual weddings on the beach, or intimate evening weddings.Â
The column/sheath gown is flattering on lean figures, both short and tall. The elongating nature of this design help petite brides appear taller. The column/sheath dress does not allow much room for hiding problem areas, such as a large lower body. It may conceal minor flaws if you opt for a constructed column that will stay in place, but the slip style may be unforgiving.
4). BALL GOWN. The ball gown is a perfect wedding gown for the bride envisioning a fairytale wedding. This silhouette has a fitted bodice (part of the gown above the waist) with a full skirt. The ball gown is most appropriate for traditional weddings where the dress will be consistent with the formality of the event.Â
The ball gown is flattering for almost all body types. The ball gown is ideal for slender or pear-shaped figures as the full skirt helps to accentuate the waist and hide the lower body. If you have a large bust, this style will help to create an hourglass look since it emphasizes the waistline due to the fitted bodice (part of the gown above the waist) and natural or dropped waist.
5). MERMAID. The mermaid silhouette contours the body from the chest to the knee, then flares out to the hem. This is a sexy look that highlights curves of your body. For this reason, you must be confident and comfortable to pull off this dramatic style.Â
The mermaid gown is not a style for everyone. This daring design is best worn by slender figures, both short and tall. The mermaid dress is made to hug the body and show every curve, it is not recommended for those brides with full figures, however if you are on the voluptuous side and want to accentuate your curves, this is the perfect dress for you. When considering a mermaid, you will want to hide embarrassing underwear lines, so be careful of your undergarment selections.
Copyright 2013, Kim Horn, MBC™
To hire Kim as your wedding planner, call 480.921.7891 or text your name, wedding date, and wedding planner request to 602.418.9089.
Kim M. Horn, MBC™
Master Bridal Consultant | 1 of 61 in the World
Scottsdale, Paradise Valley, Chandler, Phoenix and Destination Weddings
Publisher | Pres. | ArizonaBridalSource.com
AZ State Coordinator Assoc. of Bridal Consultants May 2003 – May 2012
2010 Annual Conference Chair (worked with David Tutera) – Assoc. of Bridal Consultants
E email@example.com


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These Six Babies Dressed As Tiny Disney Princesses Is The Best Thing You'll See Today.
We all know that our daughters are princesses in our hearts and the apple of our eyes, but photographer Karen Marie has taken it to the next level with her Belly Beautiful Portraits project and turned a group of babies into royalty, ahem, make that Disney royalty. The results were magical as she transforms six babies into teensy-weensy versions of the most popular Disney princesses ever. We doubt even the Blue Fairy could have done better.
At a studio in Roseville, California, photographer Karen Marie was busy working her magic.
In the process, she transformed these itty-bitty babies into Ariel, Snow White, Sleeping Beauty, Cinderella, Belle, and Jasmine, which included kids from her current client list as well as baby models.
Karen created the perfect setting to go with the costumes for each of the Disney princesses in training.
Forest creatures watching over this beloved Snow White are making our heart ache from so much adorableness. But Karen could thank those magic gowns to Valerie Best of Sew Trendy Accessories.
Karen, who has two daughters of her own are now 10 and 12, in other words, too old to be princesses.
But the photographers loves kids, princesses, and Disney so much that this project allowed her to bring something back into her life that she had missed so much with a collection of pint-size Disney princesses.
Have you ever seen an Ariel who's lovelier than this precious little one?
We just love the way Karen thought of everything from the setting of the fork, and even the plush Flounder who watches over this little mermaid.
While Sew Trendy Accessories helped with the costumes, Wooly Dreams brought magic to the studio.
Wooly Dreams provided the felted flowers and animals to accentuate the enchanted world of these oh-so popular Disney princesses. This collection was ultimately a dream come true for Karen.
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Whatever the occasion, makeup can enhance or destroy your look. If you get it right, the overall look can be amazing; Professional Makeup Companies in Woodmead get it wrong and it all looks like an artificial mess. For a bride to be all these considerations are multiplied in importance so as to look as beautiful as possible on the big day.
One of the major mistakes brides tend to make is to wear no or very little makeup. Even if a bride is lucky enough to have a gorgeous natural look, Average Price For Makeup Artist it would still make sense to wear a little. There will likely be no other day when you will be photographed more than your wedding day, and covering up slight blemishes will help you treasure those photos the rest of your life.
Finally, remember to take the weather and ambient heat into account. If your lucky day promises to be a hot one, keep the makeup as light as possible so it does not run or cake. Remember you will be standing there in front of two entire families and their friends; Best Makeup Artist you are going to be hot.
Professional Makeup Companies in Woodmead ?
Wedding is the most auspicious day in a bride's life. She wants to look extraordinary beautiful on her wedding day. Bridal makeup can help you to get the desired looks on your special day. You should choose the best wedding makeup artist to get the desired look. Experience and perfect skills of a makeup artists can help you to get the desired results. A wedding artist can help you to look simple and beautiful by applying the right makeup techniques. There are some artists who do not know the right techniques and they just cover your face with several layers of makeup that affect your natural looks badly. Therefore, it is very important to choose the best wedding makeup artists.You can make your wedding a memorable event by choosing the best wedding makeup artist. You have to put some efforts in choosing the right makeup artist. You have to do research on the internet and meet different artists to select the most appropriate artist for your wedding.
Want to Become a Make-Up Artist? Know It's Pros and Cons, Make Up Schools, and Courses
As I've worked in the cosmetics industry I've had the opportunity to work with makeup artists, makeup schools and professional makeup companies; living in Hollywood, California for the last 10 years has increased my exposure to those various companies. Throughout that time I've also had the opportunity to learn how to apply the makeup that I've been creating for years and even learn some of the fun aspects of makeup artistry including working with latex and silicone FX makeup. I've been fortunate in that all the additional knowledge I've acquired has been a result of work that I've done with these companies. Many of the students I've met, haven't been as fortunate in that they've had to pay full price (sometimes upwards of $30,000) for the same knowledge. At the end of their studies they are generally given the opportunity to venture out and apply their new trade for free, maybe if they're lucky than can get a kit fee (usually around $45) for a day's worth of work. I've heard the term "paying their dues" applied to this over the years, of course the people saying this were the same ones charging the 30 thousand dollars. I for one have always felt like the new makeup artists are being taken advantage of, perhaps it has more to do with my inner capitalist screaming at the thought of a person giving away their time. The old adage "why buy the cow when you can get the milk for free" comes to mind: When you start down the path of giving your services or product away for free, it is incredibly difficult to ever break that cycle. Whereas it may take a little longer for you to be a makeup artist when you charge for your skills, I believe that when we earn payment for our work we carry our heads a little higher and the have more pride in our work in the end gain greater respect from our customers.As companies like By Jove Cosmetics continue to expand their custom blending kits to include Blush, Eye shadow and Lip colors, any color in the rainbow will be achievable by everyone who can see color.With free access to the internet being what it is, there is such a wealth of knowledge and networking access (via Facebook, Twitter and MySpace) that is available today that anyone who desires to be a professional makeup artist can do so from virtually anywhere in the world and you should not feel like you have to go to Hollywood or New York and spend tens of thousands of dollars to get your start.


text: STOP Smoking Now with Self-Hypnosis STOP Smoking Now with Self-Hypnosis PLDZ-25 In Stock $ 19.95 USD Buy and Download > Description "Most of us have a tendency towards anxiety or even panic in some areas of life. Deciding to stop smoking is definitely one of these! Sometimes we understand the fear & other times not. Often it is illogical & we may even question our own ability to own our own life & control outcomes. Although you may feel the urge to run away & hide from the inevitable, the answer is to run towards it & own it. Then you are free to manage both the physiological & emotional aspects of it.".........Elizabeth Bohorquez, RN, C.Ht No Butt's About It....There is Nothing Good to Say About Smoking....This audio program is designed to assist smokers in diminishing & finally ending this dangerous habit. There are several keys to finalizing an addiction, whether it be smoking, alcohol, drugs, food, etc. One is truly seeing it for what it is & what it represents. This is accomplished through High Level Awareness coupled with Interactive Self-Hypnosis. Each individual has different motivators & de-motivators. These need to be programmed hypnotically. Next, one needs to climb the steps to cessation & to do so in order. This means ordering one's nutrition which is very connected to smoking cessation & the development of stress. Next comes emotional & thought management, ending on the top step of inner imagery or imaginology for managing moments when cravings or negative motivators may have slipped through the cracks. This audio can be utilized for any nicotine problem including tobacco chewing, cigar and/or pipe smoking. Note: All programs produced by International Medical & Health Writers, Ltd. are written & recorded by Elizabeth Bohorquez, RN, C.Ht. Each program has 2 or 3 separate original Interactive Self-Hypnosis Sessions, each with it's own induction. All programs include directions for listening & other important information. "Most of us have a tendency towards anxiety or even panic in some areas of life. Deciding to stop smoking is definitely one of these! Sometimes we understand the fear & other times not. Often it is illogical & we may even question our own abi Direct Link https://store.payloadz.com/go/?id=1361928 https://store.payloadz.com/details/1361928-audio-books-health-and-well-being-stop-smoking-now-with-self-hypnosis.html Tags smokingtobaccohypnosisaddiction Health and Well Being Audio Books


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Home > Giving > Legacy Society
Named in honor of Edward A. Langenbach, who was instrumental in the formation of Stark Community Foundation, the Legacy Society recognizes and thanks individuals and families who plan to leave a gift to the community through a bequest, trust, life insurance policy, retirement plan or charitable annuity. Members of our Legacy Society care about the future of the community and realize that their gifts will strengthen the projects and programs of local nonprofit organizations and the people they serve. We thank the following members for their foresight and generosity.
|Anonymous (6)
|Ron and Susan Amedeo
|Paul R. Arena*
|Arlene A. Bartlow*
|Robert E.* and Mary B. Basta
|Julia B. Belden*
|Robert F. and Kathleen K. Belden
|Murray M.* and Hanna V. Bett
|Jane A. and Paul R. Bishop
|Warren M. and Sheila Markley Black
|William F. Jr.* and Kathleen M. Brauchler
|Anne S. Bretean*
|Jeanne M. Cullen*
|William T. Cumler*
|Joyce and Andrew Deuble
|Don* and Phyllis Dickes
|Jocelyn Dobransky
|Cindy and Jon Elsasser
|William and Julieann Farber
|Robert F. Fay
|Dwain and Kathy Foltz
|Robert I. and Diane J. Friedman
|Geoff and Bobbi Gordon
|Jim* and Shirley Green
|Steven M. Gyalai
|Kathleen Fernandez Hillibish
|David and Susan Hoecker
|Dr. Grace Hofsteter*
|Douglas L. Houck*
|Dorothy R. Householder
|Joseph F. Jadick
|Eugene B. Jester*
|James W. and Carol A. Johnson
|David P. Kielson
|Thomas A. Kolp
|Christine and Howard* Kruman
|Dean G.* and Jane G. Lauritzen
|William J. Lemmon
|Robert Lewis
|William L. and Sharon K. Luntz
|Army* and Barbara Marzilli
|James H. McVay
|Donald M. and Cynthia A. Miller
|R.C. Miller Family
|Phyllis and Jack Milligan
|Jean E. Naughtrip*
|John and Pamela Frease Wynn
|John B. Nolan
|Stephen J. Ondrejas
|Charles J. Paquelet
|Lou dell A. Printz
|Mr.* and Mrs.* Giles B.J. Raisner, Jr.
|David and Ann Renkert
|Edie Sanders
|Chuck and Judy Scheurer
|Earl C. and Nancy N. Sheehan
|Gary and Linda Sirak
|Dr. Mike Shreffler
|Gary R. and Jan Smith
|Carl and Sally Sorenson
|Charles P. Sturgis*
|Peggy and John Stratton
|Beth Ann Toalston
|Ralph E. Toalston
|Joe and Marlene Toot
|Ken* and Joan Voyles
|James R. and Ida A. (Tenaglia) Warner
|Brandon* and Paula Wehl
|Harold L.* and Edna M. Weil
|Fred and A.J. Whittier
|Joseph Wojcik
If Stark Community Foundation is part of your future giving plan or if you would like further information about our Legacy Society, please contact Bridgette Neisel, vice president of advancement, at 330-454-7992 or email@example.com.
Why Donate? learn more
Creating a lasting legacy.Start a Fund
Contact online or call
330 454 7992


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Interview with Bree Despain
Hi Bree! So what first sparked the idea of The Dark Divine?
Well, the idea for the story was actually the combination of lots of ideas that all came together one night as I was driving and a conversation between a brother and a sister popped into my head. They were discussing a former friend who had recently reappeared in their lives. "Stay away from him" the brother warned, "He's not who he used to be. He's dangerous." That conversation was so haunting; I had to know what was going on and so that night I went home and starting writing.
Can you tell us about how your cover came about?
Back in February my editor asked me if I had any thoughts about what I wanted on the cover. What he got back was a two-page, detailed letter explaining what mood and atmosphere I wanted the cover to convey, and a list of different book covers I thought were good examples of I wantedJ. I was hoping for something beautiful and striking, but also a little eerie at the same time.
I wish I could take credit for the beautiful cover, but the designer, Joel Tippe, deserves all the praise. After exploring a few different concepts, he came up with the cover we all loved. They told me to consider it an abstract representation of the mood of the book rather than of a literal event. However, the cover and the colors used do have more of a connection with the final version of the novel as some minor manuscript revisions were made after the ARCs went to press.
Was Daniel's character inspired by anyone you know of? Or were any characters inspired by someone you knew in real life?
The idea for Daniel comes from three different guys from my past that pulled similar disappearing acts on me, but he is very much his own personality. Daniel was such a strong character from the beginning. Grace has a little of me in her, but we are definitely different people too. I like to think of her as a stronger version of teenage me. The other characters are not really based on anyone in particular, but do share similar traits with some of the people in my life.
If The Dark Divine was made into a movie, who would you pick to play Grace, Daniel and Jude?
Hey I actually did a post about this (with pictures!) over on The Tenners site. It was a fun conversation.
Bree picked for Daniel as first choice a younger TAYLOR KITSCH, second choice would be ZAC EFRON (I would love to see Zac as Daniel!). For Jude Divine it would be a young TOM WELLING from the early Smallville or CHACE CRAWFORD from Gossip Girl. I totally approve of Chace! And lastly for Grace Divine, Bree picked EMMA ROBERTS whom I reckon is an excellent choice!
Can you share with us why Grace still loved Daniel after all those years. Most girls would have moved on and fallen for another guy.
The relationship that Daniel and Grace share is a lot deeper that simple attraction. It was and is still rooted in a deep friendship and concern for each other's happiness. Some people will just always be with you and be a part of who you are.
Grace has certainly dated other guys, and she thought she had moved on and gotten over Daniel. That is, until she saw him again and all those feelings and thoughts of what could have been began to return.
Are you currently working on anything?
Yes! I am in the middle of writing the sequel to THE DARK DIVINE. And I have several more stories I hope to tell.
Who are some of your favourite YA authors?
So, so many. Laurie Halse Anderson, Meg Cabot, Sarah Zarr, Emily Wing Smith, Suzanne Collins, to name a few. I've recently read HUSH HUSH by Becca Fitzpatrick too and really enjoyed it.
Any advice to people participating in NaNoWriMo? *yes I know November is already over but take it as an advice for next year*
Um… keep writing!
I couldn't be happier with the cover and was so excited that the designer was able to put my thoughts into print. I hope you all love it as much as I do.
Thanks for the interview! It was a lot of fun.


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Dining With Denise Neil
Celeb chef Lidia Bastianich will swing through Wichita while checking on KC
Italian chef Lidia Bastianich is one of the rare food celebrities who has a restaurant close enough for Wichitans to visit regularly. Her Lidia's Kansas City at 101 W. 22nd St. in Kansas City, Mo., serves homey Italian dishes in an upscale setting.
Bastianich, 68, is also a television star and cookbook author, and she's making a visit to Wichita. She'll appear at Abode Venue at 1330 E. Douglas on Tuesday, where she'll sign copies of her new cookbook, "Lidia's Mastering the Art of Italian Cuisine: Everything You Need to Know to Be a Great Italian Cook," and speak at an event moderated by Wichita's Scott Redler, co-owner of Freddy's Frozen Custard.
During a recent phone interview from New York City, where she lives and owns four restaurants, Bastianich talked about her new cookbook, her love for Kansas City, and her thoughts on Americanized Italian dishes.
Bastianich, whose cooking show, "Lidia's Kitchen," airs at 1 p.m. Tuesdays on KPTS, Channel 8, said she likes to get back to Kansas City frequently to speak with the staff, make menu revisions, and taste everything her restaurant is serving. While in town, she said, she tries to schedule an appearance or two nearby.
When her Kansas City restaurant opened 17 years ago in the Crossroads, a former railroad freight house, it was kind of isolated, she said. Now, it has two high-end next-door neighbors – Fiorella's Jack Stack Barbecue and German restaurant Grunauer – plus a whole arts district that's developed around it.
"I like coming back to Kansas City," Bastianich said. "It's a great city. I like the feel. I especially like the area where we are at now. It's really evolved. When we first started there, it was all dead. It was kind of a nostalgic, romantic setting. Now it's kind of a happening setting."
Her new cookbook, which was released in October, is her 10th. Bastianich calls it her "magnum opus." The hardbound book contains more than 400 Italian recipes, and that's not all.
"This one is kind of a collection of a lot of how-tos and whys," Bastianich said. "The first 100 pages is about my philosophy, history and relationship with food and about understanding Italian food and traditional Italian products."
Bastianich offers, for example, a guide to Italian cheeses, a glossary of pasta varieties and information about the benefits of braising, broiling and brining.
She includes recipes for exotic-sounding Italian recipes like salt cod fritters and bread and prune gnocchi. She also includes recipes for some of the Americanized Italian dishes popular in the states, like manicotti, spaghetti and meatballs and eggplant Parmesan.
"I think the Italian-American cuisine is a great cuisine," she said. "It's wonderful. It's tasty. It's good. It's not what you would eat in Italy, though. It's the cuisine of Italian immigrants adapting themselves to this new world they found. But I love it. A Sunday sauce is delicious. Manicotti is delicious."
Her presentation on Tuesday, Bastianich said, will appeal to anyone interested in food. It will start with Redler asking her a list of questions, which she said usually leads to her talking about immigrating to America as a young child, becoming interested in cooking and growing into roles as a businesswoman, a mother, a grandmother and a television chef.
She'll then take questions from members of the audience, who usually want to know how to choose the right olive oil and which pastas go with which sauces, she said.
Lidia Bastianich
What: The celebrity chef and cookbook author will make an appearance in Wichita
When: 6 p.m. Tuesday. Doors open at 5:15 p.m.
Where: Abode Venue, 1330 E. Douglas
Tickets: $37.50, which includes a copy of the new cookbook and a free companion ticket, at Watermark Books, 1330 E. Douglas, or online at http://www.watermarkbooks.com/, where if you buy a copy of the cookbook, you'll also be signed up for tickets
Also: A cash bar will be available
Win a copy of Lidia's cookbook
Want your very own copy of celeb chef Lidia Bastianich's new cookbook, which she'll be discussing at an appearance in Wichita next week? I have a copy, and I'm giving it away.
All you have to do is make a comment on my post about the giveaway at Facebook.com/diningwithdenise . On Monday, I'll randomly choose a comment, and its author will win my copy of "Lidia's Mastering the Art of Italian Cuisine," a $37.50 value.
Give the Dining with Denise Facebook page a like while you're there, if you're so inclined.


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Bobby Buckle
Blue Plaque Application
Bobby Buckle was a founder, first captain, scorer of the first recorded Spurs goal, Treasurer, Secretary and Director of Tottenham Hotspur Football Club.
Without Bobby Buckle, there would be no Spurs! He wasn't just one of the founders of the Hotspur football team he was at the centre of every major moment in its development off the field during the first twenty years of its existence.
While on the pitch he was the team captain ( just 13 years old) in that first season and is the first known goal scorer in the clubs history. A speedy winger he remained a main stay of the side for ten years
He attended Tottenham Grammar School from 1881 to 1883. Some Tottenham Grammar Old Boys have set up a small committee to try to get his influence on the founding and development of Spurs recognised.
This website brings together all the information that has been gathered to use in the application to get a Blue Plaque placed on Bobby Buckle's family home at White Cottage, 7 White Hart Lane, Tottenham. This address was used as the first postal address for Tottenham Hotspur Football Club.
To help the application we need to establish the level of support from those living and working in Tottenham, those involved with the Club and fans both in the UK and overseas
If you would like to support the application please go to the 'Support Us' page.
If you would like more information about London's Blue Plaques you can click here: www.theheritage-foundation.co.uk/


text: On 3/26/14, 10:47 AM, Paul Kyzivat wrote: > On 3/25/14 2:36 PM, Tony Hansen wrote: > >> Are you saying that you don't know how to set the datatracker to show an >> individual the entire document by default? >> >> (upper left corner, under Settings is New Account and Options. Click >> Options, and set your default for Show full document text in document >> page) > > I didn't. Thanks! > > But I assume that setting will only apply when logged in. Most of the > time when I retrieve a doc from the data tracker I'm not logged in. It uses a cookie in your browser. So you need to change the setting again if you switch to a different browser. You don't need to be logged in. Tony


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The Grass Shack is a locally owned and family (ohana) operated dispensary, that welcomes OMMP patients, cardholders and everyone 21 years or older. We are committed to providing great service and providing the finest lab tested flower and other products at a fair price. We serve everyone in the spirit of aloha and wish to share with them our knowledge of the health benefits and healing power of the wonderful Cannabis plant. We also have a variety of edibles, concentrates, wax, vape pens and topical products.
We embrace new relationships with patients, caregivers, customers and high quality vendors. Our goal is to help as many individuals as possible lead more productive and pain free lives. We look forward to meeting you and taking care of your medical and recreational marijuana needs.
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A Plumbing Permit is required for all changes to plumbing, including installation or replacement of a water softener, water heater, or back flow prevention system for lawn irrigation. For water heater replacements, a fresh air intake must be installed if not existing.
Plumbers must have a current state license and bond on file with the city.
*Please call at least 24-48 hours ahead of desired inspection time.*Building Permit Technician: 651-552-4116 or email@example.com with following information: -Permit Number -Type of Inspection -Day/Time Requesting -Contact name & Phone Number


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NIX Advance frames allow you to play HD videos and photos in the same Slideshow
720p HD Video Formats: MP4 (H.264 HD 720p AAC)
Fast processor: View thousands of photos and videos from your inserted USB stick or SD/SDHC card
Nix Advance turns on when you enter the room to display the most recent photos & videos and turns off when you leave.
For the ultimate convenience, Nix Advance allows you to set the frame to switch off after sensing no movement for 5 minutes up to 1 hour.
Why NIX? NIX is the leading brand for Digital Photo Frames in the U.S. With 7 years of experience and dedicated customer service team, we are committed to delivering high quality frames and services that allow you to display digital memories in a sophisticated way. To assure you of the quality and service we provide, we give you a full 1 Year Part's & Labor Warranty at the time of purchase. Need to display all your digital photos? The NIX Digital Photo Frame is the most simple and affordable solution to display all your digital moments. Just plug in your USB stick or SD/SDHC card and see all your photos play in front of you! It's that easy. This frame is perfect for everyone from professionals, to parents and grandparents who just want a frame that works with the convenience of not having to actually operate it. Combined with an intuitive remote, it's a breeze to get started. NIX Digital Frame With a High Resolution LED Backlit Display, enjoy all your (JPEG) photos with amazing...


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« Back to Events
25 August, 2019 - 29 August, 2019
Dublin, Ireland
This year's EMBS will take place in Dublin on August 25 to 29. It will be hosted by University College Dublin, in the state-of-the-art O'Brien Centre for Science. The social programme will provide opportunities to sample the the city's famous cultural life and explore the UNESCO Dublin Bay Biosphere.
2019 will be the International Year of the Salmon, a fish that has a special place in the mythology of Ireland. Look out for salmon throughout the scientific and social programme. Find out more about the Symposium themes for 2019, the keynote speakers and the key dates. The call for abstracts is open until 3 April 2019.


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The ball was changed three times during the Bangladesh innings, twice more than is mandatory under the regulations, as the humid night-time atmosphere made it hard for the England bowlers to hold the ball.
"Graeme Swann couldn't grip the ball at all," Strauss said. "I think there's something not quite right if a spinner can't grip the ball in this part of the world, where spin plays such an important role.
"Graeme was obviously very frustrated, because the dew meant he couldn't grip the ball. He felt he had a big role to play in the game, and for a period there we had to take him off until later on when he could grip it.
"It seems slightly strange to have the first ever day-nighter at a certain ground in a World Cup. So perhaps lessons can be learned there."
Strauss had to intervene as Swann became involved in an argument with umpire Daryl Harper and at one stage commentators on television apologised after the stump microphone picked up the England spinner swearing.
"It was frustrating for him, and all of us, that the ball got as wet as it did but those are the conditions we encountered," Strauss said. "I think Graeme was asking to change the ball. I wasn't there so didn't hear what was said. But once the exchange happened I told Graeme to calm down and get on with it – and he did do so."
At no stage during this World Cup have England managed to produce consistent cricket that would suggest they have the ability to win the tournament.
It has been a long winter on the road although Strauss refused to blame fatigue and feels England can still regroup to become a force in the World Cup.
"We still believe we can win the World Cup," he said. "We've put in some very inconsistent performances and still haven't had a game when our batting and bowling fires at the same time. We're going to have to do it in that last game against the West Indies.
"World Cups are funny things. You can gain confidence just at the right moment, and get in a real run of form just when you need it. But we need to win that final group game to qualify for the quarter-finals. Clearly we need consistency from now on if we want to progress in the tournament."
Thousands of Bangladesh fans swarmed into the streets to celebrate victory, in sharp contrast to the public mood after their defeat to the West Indies, when the opposition team bus was stoned.