Expression of adeFGH was not the cause of resistance in the clinical isolates of MDR A. baumannii, DB and R2. This method allows the impact of each efflux system on antimicrobial resistance to be clearly defined. Methods Bacterial strains, plasmids and LY2603618 cost culture conditions Bacterial strains and plasmids used in this study are listed in Table  3. Acinetobacter baumannii R2 (TTSH6013 654325/06) and DB (DB15354/07) were clinical isolates from a collection by the Network for Antimicrobial Resistance Surveillance, Singapore. According to the interim standard find more definitions for acquired resistance, both DB and R2 are classified as MDR as they are

non-susceptible to ≥1 agent in ≥3 antimicrobial categories (aminoglycosides, fluoroquinolones, carbapenems, tetracycline, extended spectrum cephalosporins, folate pathway inhibitors) [17]. DB and R2 carry and express bla OXA-23-like and bla OXA-51-like, do not carry bla OXA-24-like and bla OXA-58-like (data not shown). A. baumannii and E. coli were cultured under aerobic conditions at 37°C in Luria-Bertani Miller (LB) agar or LB broth (Becton Dickinson, Cockeysville, U.S.A.). Antibiotics used were at the following concentrations for E. coli: kanamycin, 10 mg/L; tellurite

6 mg/L; and for A. baumannii: tellurite, 30 mg/L. Table 3 List of bacterial strains and plasmids used in this study Strain or plasmid Relevant characteristics Reference or source A. baumannii strains     R2 Wild-type clinical MDR isolate TTSH6013 624325/06 Network for Antimicrobial Resistance Surveillance (Singapore) Apoptosis Compound Library research buy DB Wild-type clinical MDR isolate DB15354/07 Network for Antimicrobial Resistance Surveillance (Singapore) R2ΔadeFGH Sucrase R2 with deletion in adeFGH operon This study R2ΔadeIJK R2 with deletion in adeIJK operon This study R2ΔadeFGHΔadeIJK R2 with deletion in adeFGH and adeIJK operons This study DBΔadeFGH DB with deletion in adeFGH operon This

study DBΔadeIJK DB with deletion in adeIJK operon This study DBΔadeFGHΔadeIJK DB with deletion in adeFGH and adeIJK operons This study E. coli strains     DH5α F– Φ80lacZΔM15 Δ(lacZYA-argF) U169 recA1 endA1 hsdR17 phoA supE44 λ– thi-1 gyrA96 relA1 Invitrogen S17-1 Genotype: recA pro hsdR RP4-2-Tc::Mu-Km::Tn7, GmS [16] Plasmids     pMo130 Suicide plasmid, xylE +, sacB +, KmR [8] pwFRT-TelR Donor of tellurite resistance cassette [10] pMo130-TelR pMo130 plasmid containing 3.26 kb XmaI-digested tellurite-resistance cassette from pwFRT-TelR This study pMo130-TelR-P8(UP/DWN) pMo130-TelR containing a 1 kb UP fragment (promoterless adeL) and 1 kb DOWN fragment (3’ partial adeH) This study pMo130-TelR-adeJ(Up/Down) pMo130-TelR containing a 1 kb UP fragment (5’ partial adeI) and 0.9 kb DOWN fragment (3’ partial adeK) This study DNA manipulations Bacterial genomic DNA was extracted using a rapid procedure described by Pitcher et al[18].

Based on the previous studies, we hypothesized that PDCD4 might also play a role on the inhibition of HCC metastasis. To testify this hypothesis, we first examined the expressions of PDCD4 in three human HCC cell lines with different metastasis potentials, then we transfected

a plasmid encoding the PDCD4 gene into HCC cells with lowest PDCD4 expression level and further investigated the effects of PDCD4 on the gene expression of MTA1 and migration and invasion of HCC cells. Methods Cell lines and cell culture Three human HCC cell lines, MHCC-97H (high metastatic potential), MHCC-97L (low metastatic potential), Hep3B(no metastatic potential) [14], were obtained from the Liver Cancer Institute of Zhongshan Hospital, Fudan University, Shanghai, China. One normal human liver cell line L02 [15]and one mouse fibroblast cell line NIH3T3[16] was obtained from the Central Laboratory of Shandong Provincial Hospital. find more HCC cells were routinely cultured in Dulbecco’s modified Eagle’s medium (DMEM) with high glucose (Hyclone, USA). L02 and NIH3T3 cells were cultured in RPMI 1640 medium (Hyclone, USA). Both the DMEM and the RPMI 1640 medium were

https://www.selleckchem.com/products/Roscovitine.html RG-7388 in vivo supplemented with 10% fetal bovine serum (FBS, Gibco, USA), antibiotics (100 U/ml penicillin, 2 μg/ml streptomycin) and 2 mmol/L glutamine, at 37°C in a humidified, 5% CO2 atmosphere. Immunocytochemistry MHCC-97H, MHCC-97L and Hep3B cells were cultured in 24-well plates with one glass slide in each well. Twenty four hours later, the slides were washed with PBS, fixed with 4% paraformaldehyde for 30 min and permeablized with 0.2% Triton X-100 for 20 minutes. In order to inhibit the endogenous peroxidase activity, the slides were treated with 3% H2O2 for 15 min. The nonspecific binding sites were blocked by incubation in a solution of 5% bovine serum albumin (BSA) for 20 min. The primary rabbit polyclonal antibody Immune system to PDCD4 (Santa Cruz Biotechnology, Santa Cruz, California, USA. diluted by 1:30 in phosphate-buffered saline, PBS) was applied and incubated

at 4°C, overnight. Slides were washed twice with PBS and incubated with biotinylated goat anti-rabbit IgG (Santa Cruz Biotechnology, Santa Cruz, California, USA) at a 1:100 dilution. Slides were then incubated for 30 min with HRP-conjugated streptavidin (Zhongshan Biotechnology, Beijing, China). The avidin/biotin complexes were revealed with a diaminobenzidine (DAB) kit (Zhongshan Biotechnology, Beijing, China) according to the manufacturer’s instructions. Hematoxylin was used to counterstain the slides which were then dehydrated and cover-slipped. Equal volume of PBS was used instead of the primary antibody and served as a negative control [17]. A semi-quantitative scoring method was used to assess the expression level of PDCD4.

98 ±15% for mean power. Participants practiced the anaerobic capacity test during the familiarization session to minimize learning effects. Side effect assessment Participants were given weekly questionnaires on how well they tolerated the supplement,

how well they followed the RO4929097 clinical trial supplement protocol, and if they experienced any medical problems/symptoms during the study. Compliance to the supplementation protocol was monitored by turning in empty weekly supplement containers, supplement logs and verbal confirmation. After completing C188-9 the compliance procedures, subjects were given the required supplements and dosages for the following supplementation period. Data analysis Participant baseline demographic data were analyzed by one-way Analysis of Variance (ANOVA). Study data were analyzed by Multivariate Belinostat Analysis

of Variance (MANOVA) with repeated measures. Overall MANOVA effects were examined using the Wilks’ Lamda time and group x time p-levels as well as MANOVA univariate ANOVA group effects. Greenhouse-Geisser univariate tests of within-subjects time and group x time effects and between-subjects univariate group effects were reported for each variable analyzed within the MANOVA model. In some instances, repeated measures ANOVA was run on variables not included in a MANOVA design with univariate group, time, and group x time interaction effects reported. Data were considered statistically significant when the probability of type I error was 0.05 or less and statistical trends were considered when the probability pheromone of error ranged between p > 0.05 to p < 0.10. If a significant group, treatment and/or interaction alpha level was observed, Tukey’s least

significant differences (LSD) post-hoc analysis was performed to determine where significance was obtained. A priori power analysis of the design indicated that an n-size of 12 per group was sufficiently powered to identify previously reported changes in muscle creatine content and training adaptations in responses to creatine supplementation (>0.70). Results Subject demographics Forty-one participants were initially recruited for the study, completed consent forms and participated in the required familiarization session. Of the original 41 participants, 36 completed the 28-day research study. Three participants dropped out due to time constraints, one due to an unrelated illness, and one due to apprehension of the muscle biopsy procedure. None of the participants dropped out of the study due to side effects related to the study protocol. Table 3 shows the baseline demographics for the participants. Overall, participants were 20.2 ± 2 years, 181 ± 7 cm, 82.1 ± 12 kg, and 14.7 ± 5% fat with 3.8 ± 3 years of resistance training experience. One-way ANOVA revealed no significant differences among groups in baseline demographic variables.

RA is working as

an assistant professor in the Interdisciplinary Research Center in Biomedical Materials (IRCBM) at COMSATS Institute of Information Technology, Lahore, Pakistan. His research interests are in the field of artificially designed DNA nanostructures and their applications in different fields, especially in biosensor applications, nanodevices designing and fabrication, and tissue selleck chemicals engineering, especially in assisting burn patients. Acknowledgments learn more This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (2012-005985). References 1. Sekhon BS: Nanobiotechnology: an overview of drug discovery, delivery and development. Pharmacol Ther 2005, 69:13. 2. Seeman NC: Nanomaterials based on DNA. Annu Rev Biochem 2010, 79:65–87.CrossRef 3. ACS: Redefining DNA: Darwin from the atom up . In American Chemical Society’s 237th National Meeting: selleck inhibitor March 22–29 2009; Salt Lake City. Edited by: Bernstein M. Washington DC: ACS; 2009:237. 4. Kallenbach NR, Ma RI, Seeman NC: An immobile nucleic acid junction constructed from oligonucleotides. Nature 1983,305(5937):829–831.CrossRef 5. Pinheiro AV, Han D, Shih WM, Yan H: Challenges and opportunities for structural DNA nanotechnology. Nat Nanotechnol 2011,6(12):763–772.CrossRef 6. Aldaye FA, Palmer AL, Sleiman HF: Assembling materials with DNA

as the guide. Science 2008,321(5897):1795–1799.CrossRef 7. Shih WM, Lin C: Knitting complex weaves with DNA origami. Curr Opin Struct Biol 2010,20(3):276–282.CrossRef 8. Seeman NC: Nucleic acid junctions and lattices. J Theor Biol 1982,99(2):237–247.CrossRef 9. Seeman NC: DNA in a material world. Nature 2003,421(6921):427–431.CrossRef 10. Yurke B, Turberfield AJ, Mills AP, Simmel FC, Neumann

JL: A DNA-fuelled molecular machine made of BCKDHA DNA. Nature 2000,406(6796):605–608.CrossRef 11. Mao C, Sun W, Shen Z, Seeman NC: A nanomechanical device based on the B-Z transition of DNA. Nature 1999,397(6715):144–146.CrossRef 12. Kay ER, Leigh DA, Zerbetto F: Synthetic molecular motors and mechanical machines. Angew Chem Int Ed 2007,46(1–2):72–191.CrossRef 13. Keller S, Marx A: The use of enzymes for construction of DNA-based objects and assemblies. Chem Inform 2012,40(12):5690–5697. 14. Hemminga MA, Vos WL, Nazarov PV, Koehorst RB, Wolfs CJ, Spruijt RB, Stopar D: Viruses: incredible nanomachines. New advances with filamentous phages. Eur Biophys J 2010,39(4):541–550.CrossRef 15. Park SH, Yin P, Liu Y, Reif JH, LaBean TH, Yan H: Programmable DNA self-assemblies for nanoscale organization of ligands and proteins. Nano Lett 2005,5(4):729–733.CrossRef 16. Lund K, Liu Y, Lindsay S, Yan H: Self-assembling a molecular pegboard. J Am Chem Soc 2005,127(50):17606–17607.CrossRef 17.

05 versus

respective untreated cells) (mean±SD, n = 3). (g) Significant decreases in TER were also seen in the transfected LOXO-101 cells MDA CL5exp after treatment with HGF (using ANOVA p ≤ 0.05 versus respective untreated cells) (mean±SD, n = 3) and in MDA CL5rib2 (h) (using ANOVA p ≤ 0.05versus respective untreated cells) (mean±SD, n = 3). Low levels of Claudin-5 reduces the cell adhesion to an artificial Matrigel basement membrane The ability of MDACl5exp and MDACL5rib2 cells to adhere to matrix was assessed in an in vitro Matrigel adhesion assay (Figure 4b). There was a significant difference between the adherence of MDACL5rib2 and MDApEF6 with MDACL5rib2 cells being less adherent to matrix. In the case of MDACl5exp, the opposite effect was seen, however differences did not reach statistical significance when compared to the control. Claudin-5 did not alter the invasive phenotype of transfected human breast cancer cells The invasive potential of the transfected cells MDACl5exp and MDACL5rib2 was examined using an in vitro Matrigel invasion assay (Figure 4c). Both cell lines were found to have no significant

differences when compared to the control MDApEF6 and invaded as individual MLN2238 clinical trial cells, with no apparent difference in invasion patterns. Claudin-5 did not alter the in vivo tumour growth of human breast cancer cells The growth and capability of developing tumours of MDACl5exp in an in vivo model was examined and compared to the control MDApEF6 cells after subcutaneous injection into the athymic nude mouse model. Over the period of 33 days, no significant difference was observed between the two groups, the control (injected with MDApEF6) and those injected with MDACl5exp (Figure 4d). Low levels of Claudin-5 confers increased trans-epithelial resistance (TER) in human breast cancer cells Transepithelial resistance was measured to assess the effect of over-expressing or knocking-down Claudin-5

on TJ functionality in MDA-MB-231 breast cancer cells (Figure 4e). If the cells were to produce a higher resistance, this is interpreted as them having increased Tight Junction function; conversely, reduced resistance implies a loss of cell-cell contact and a reduced Tight Junction function. MDACl5exp showed increased TER over a period of 4 hours in comparison others with the control MDApEF6. Changes in TER were more evident in MDACL5rib2 when compared to the control. Treatment of cells with HGF (50 ng/ml) resulted in a significant reduction of the transepithelial resistance in transfected and in control cells when Momelotinib clinical trial compare to untreated cells over a period of 4 hours (Figure 4f, g, h). Low levels of Claudin-5 retarded the motility and migration of human breast cancer cells Transfected and control cells, either untreated or treated with HGF, were evaluated for their motility using a Cytodex-2 bead motility assay to explore the possibility of Claudin-5 involvement in motility.

Therefore, further studies are required for a better understanding of our results. Brigatinib solubility dmso Figure 5 Magnetoresistivity measurements ρ xx (B) at various driving currents

I. The lattice temperature is constantly fixed at T ≈ 2 K. Figure 6 The magnetoresistivity measurements ρ xx (B) at different T for sample 1. The inset shows the Hall measurements ρ xy (B) at different T for sample 1. Figure 7 The determined exponent α in the power law T DF ∝ I α versus magnetic field B. In studying multilayer epitaxial graphene, top gating is difficult since depositing a dielectric layer is difficult and the top layers would screen the electric fields. Back gating is impractical because it would require SiC substrate thinning. Therefore, in order to further study the observed direct I-QH transition, Doramapimod cell line we choose to study various samples with different classical mobilities (see Additional file 1). In all cases, an approximately T-independent point in ρ xx is observed. The approximated T-independent Hall results suggest that Dirac

fermion-Dirac fermion interactions are not significant in all our devices [35–38]. The crossing point and some other physical quantities are listed in Table 1. We note that for the same numbers of layer, the crossing field B MK-8931 datasheet c is lower when the mobility μ is higher, consistent with the results obtained in conventional GaAs-based 2D systems [39, 40]. Moreover, the spin degree of freedom does not play an important role in the observed direct I-QH transition [41–45]. The dependence of the crossing magnetic field on the number of layers and sample does not seem to show a trend and thus requires further studies. Table 1 Sample parameters   Sample 1 Sample 2 Sample 3 Sample 4 ρ (Ω) 583 520 443 367 n (1013 cm−2) 2.08 1.98 2.16 2.44 μ (cm2/V.S) 511 605 651 694 B c (T) 9.2 4.2 6.0 5.7 v c 94 194 148 178 ρ xx/ρ xy at B c 2.1 3.7 2.5 2.8 μB c 0.47 0.25 0.39 0.40 Samples 1 and 2 were from the same chip, processed at 1,850°C

for 45 min; the former is close to the edge, and the later is near the center. Samples 3 and 4 were also from the same chip, processed at 1,950°C for 30 min; the former is close to the center, and the latter is near the edge. Lower resistivity near the edge is expected in the FTG ZD1839 molecular weight process; near the center the graphene growth is suppressed because of the higher concentration of Si vapor. At the crossing fields, the corresponding Landau filling factors are much larger than 2. Therefore, we have observed direct I-QH transition in all our devices [17–20]. It was argued that for direct I-QH transition in conventional semiconductor-based 2D systems, near the crossing field, ρ xx is approximately ρ xy, and the product of μB c is close to 1 [46]. However, in all our devices, ρ xx/ρ xy is much greater than 1, and μB c is always smaller than 1.

D: Immunoreactivity for fluorescein labelled albumin. E: Same section as ‘D’, but ultraviolet optics reveal DAPI labelled nuclei. F: Merger of ‘D’ and ‘E’ demonstrating that albumin positive cells contain large round nuclei. Calibration bar in F = 50 μm for all images. Figure 6D presents images from the adjacent section, processed for albumin immunoreactivity to identify the parenchymal hepatocytes. When this image is merged with an ultraviolet image NU7441 ic50 showing the DAPI labelled nuclei (Figure 6E,F) it can be seen that the albumin positive cells contain the large round DAPI

labelled nuclei. Counts were made of F4/80 positive cells with clear DAPI labelled ovoid nuclei, and compared to counts from adjacent or neighboring liver sections of albumin positive cells with clear DAPI labelled large round nuclei; a ratio of hepatocytes to Kupffer cells was determined for each age. These metrics, summarized in Table 1 indicate no general trend in the number of F4/80 positive Kupffer cells, relative to the number of albumin positive cells, in the early postnatal period. Table 1 Ratios of numbers of hepatocytes (H: albumin positive cells) to Kupffer cells (K: F4/80 positive cells). Age

(n) Hn (d) H nr/area Kn (Lg d) Kn (St d) K nr/area Ratio H:K P3 (2) 10.3 (0.14) 29.7 (2.1) 9.5 (0.10) 4.3 (0.06) 6.3 (1.6) 4.7:1 (0.62) P6-8 (4) 9.9 (0.15) 30.2 (3.2) 8.2 (0.17) 4.0 (0.10) 9.1 (2.1) 3.3:1 (0.27) P10-11 (3) 9.6 (0.22) 28.6 (5.4) 8.6 (0.20) 4.0 (0.11) 9.1 (2.0) 3.6:1 (0.29) P15-16 (3) Selleckchem Etoposide 9.6 (0.19) 29.9 (2.9) 8.0 (0.25) 4.1 (0.10) 8.5 (1.4) 3.5:1 (0.29) P20-21 (2) 9.4 (0.20) 31.7 (3.4) 8.0 selleck products (0.25) 4.1 (0.15) 8.0 (1.5) 3.9:1 (0.32) Data include: Ages and numbers (n) of animals in each age;

Diameter (d, in μm) of hepatocyte nuclei (Hn) and numbers of positive cells (H) in an area (nr/area) of 46,800 μm2 (260 μm × 180 μm); Diameter (d, in μm) of Kupffer cell nuclei (Kn), both long axis (Lg d) and short axis (St d) and numbers of positive cells (K) in an area of 46,800 μm2; Ratios of numbers of hepatocytes (H)/numbers of Kupffer cells (K). Data are given as: mean (standard error). Discussion Technical considerations Two techniques were employed to identify Kupffer cells in developing mice. Immunoreactivity for F4/80 was used in early studies to identify macrophages in mice [22] and since that time has been demonstrated to provide a valid marker of macrophages throughout the body and in a variety of species. In addition, administration of fluorescently labelled latex microspheres took advantage of the phagocytic activity of the Kupffer cells, and demonstrated the Kupffer cells engulfed the microspheres and led to the co-localization of microsphere labeling and F4/80 immunoreactivity. While this approach works well in adults, the small size of developing mouse pups clearly poses a challenge to GDC-0994 making reliable tail vein injections.

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In the cluster that focuses on the future, two articles draw our attention to different approaches to visioning in sustainability science. The first, by Wiek and Iwaniec, posit that since sustainability science is about transformative change, visioning is a key method. As the authors point out, sustainability visions are “specific types

of visions that provide guidance to achieve sustainability and, therefore, adhere to value-laden or normative principles including that of intergenerational equity” (WCED 1987:43). As they note, sustainability criteria can help to avoid visions that violate important values

of justice, integrity and viability. The authors review the literature in this domain SCH727965 and synthesize their findings to provide scholars with a tool to enhance sustainability-visioning practices. Ten criteria Pictilisib price for sustainability visions are laid out in a triple axis model of a quality vision: normative, constructive and transformational. The authors present design guidelines that include applying a meaningful sequence to visioning methodologies from framing through analyses, revision and recomposition of the vision. They agree with the findings of Schneider that Hydroxychloroquine visioning whether through the use of scenarios or other approaches is an iterative procedure that is conducted in participatory setting to create a shared and plausible (one could say implementable) vision. Finally, Takeuchi et al. explore the significance of the transdisciplinary sustainability science approach to analyze social and ecological restoration in NE Japan following the devastating effects of the 2011 earthquake

and tsunami. This case study of the processes for restoration in the Tohoku region argues that building resilience in the affected area requires a transformation to sustainable LY2874455 concentration agriculture, forestry and fisheries and describes how the links between satoyama and satoumi, traditional rural territorial and coastal landscapes in Japan, can contribute to this revitalization and to strengthening the relationship between local residents and the landscape in the affected communities. Decision makers at local, regional and national levels need to take a holistic approach based on sustainability science to understand the inter-relationships between these landscapes and ecosystems to develop a robust rebuilding plan for the affected communities.