Improvement of Entrepreneurship Education Program using AR : An Actual Practice of Local Revitalization at Nishichiba Kodomo Kigyo juku

Surgical procedures. (A) Pneumoperitoneum was simulated with Surgineedle™ for 10 min after xenograft procedure and pressure setting was 4 mmHg. (B) Procedure of ovariectomy (OVX) of SCID mice. (TIF 470 kb

Additional file 1 of Enhanced tumor control activities of anti-mPD-L1 antibody and antigen-presenting cell-like natural killer cell in an allograft model

Supplementary Material 1: Supplementary Table 1. Reagents applied in this study. Supplementary Table 2. Antibodies applied in this study. Supplementary Figure 1. Schematic illustration of NKDC gating strategy. Supplementary Figure 2. Schematic illustration of gating strategy of NKs and DCs. Supplementary Figure 3. Schematic illustration of gating strategy of regulatory cells. Supplementary Figure 4. Schematic illustration of gating strategy of activated CD8 T cell

Additional file 2: Figure S1. of Oestrogen-induced angiogenesis and implantation contribute to the development of parasitic myomas after laparoscopic morcellation

Surgical procedures. (A) Pneumoperitoneum was simulated with Surgineedle™ for 10 min after xenograft procedure and pressure setting was 4 mmHg. (B) Procedure of ovariectomy (OVX) of SCID mice. (TIF 470 kb

Identification of a noncanonical function for ribose-5-phosphate isomerase A promotes colorectal cancer formation by stabilizing and activating β-catenin via a novel C-terminal domain

<div><p>Altered metabolism is one of the hallmarks of cancers. Deregulation of ribose-5-phosphate isomerase A (RPIA) in the pentose phosphate pathway (PPP) is known to promote tumorigenesis in liver, lung, and breast tissues. Yet, the molecular mechanism of RPIA-mediated colorectal cancer (CRC) is unknown. Our study demonstrates a noncanonical function of RPIA in CRC. Data from the mRNAs of 80 patients’ CRC tissues and paired nontumor tissues and protein levels, as well as a CRC tissue array, indicate RPIA is significantly elevated in CRC. RPIA modulates cell proliferation and oncogenicity via activation of β-catenin in colon cancer cell lines. Unlike its role in PPP in which RPIA functions within the cytosol, RPIA enters the nucleus to form a complex with the adenomatous polyposis coli (APC) and β-catenin. This association protects β-catenin by preventing its phosphorylation, ubiquitination, and subsequent degradation. The C-terminus of RPIA (amino acids 290 to 311), a region distinct from its enzymatic domain, is necessary for RPIA-mediated tumorigenesis. Consistent with results in vitro, RPIA increases the expression of β-catenin and its target genes, and induces tumorigenesis in gut-specific promotor-carrying RPIA transgenic zebrafish. Together, we demonstrate a novel function of RPIA in CRC formation in which RPIA enters the nucleus and stabilizes β-catenin activity and suggests that RPIA might be a biomarker for targeted therapy and prognosis.</p></div

Additional file 4: Figure S2. of Oestrogen-induced angiogenesis and implantation contribute to the development of parasitic myomas after laparoscopic morcellation

Cell density and IHS of ERα, PR, SMA, Ki67, vimentin, VEGF, and CD34 in samples of in situ uterine myoma (UM) and parasitic myoma (PM) in patient No. 1. The bars show the mean value ± standard deviation. (TIF 83 kb

RPIA expression is positively correlated with β-catenin protein levels and stability in HCT116 cells.

<p>(A) Knockdown of RPIA reduced β-catenin protein levels and overexpression of RPIA increased β-catenin protein levels in both the cytoplasmic and nuclear fractions of HCT116 cells. (B) Knockdown of RPIA did not decrease ERK and pERK protein levels, which were measured by western blotting in total protein analysis (up panel) in HCT116. Conversely, overexpression of RPIA did not increase ERK and pERK protein levels (up panel). In the lower panel, both cytoplasmic and nuclear fraction showed that ERK and pERK protein levels did not up-regulate in HCT116. (C) Scatter plots show a positive correlation between RPIA and β-catenin expression in the colon tissue or nucleus. (D) To determine the half-life of β-catenin protein, western blots were used to measure the abundance of β-catenin at different time points following the addition of 10 μg/ml of the protein synthesis inhibitor CHX to HCT116 cells transfected with either control siRNA or RPIA-siRNA. The lower panels show plots of the relative β-catenin protein level, expressed as a percentage as a function of time after CHX treatment. (E) RPIA-ΔD lost the ability to stabilize β-catenin. Relative β-catenin protein levels as measured by quantification of western blot are shown in HCT116 cells. (F) The reduced β-catenin levels by RPIA knockdown were rescued by 5 μM of MG132 treatment (left panel). Inhibition of RPIA stimulated ubiquitination of β-catenin (right panel). β-Catenin was precipitated by specific antibody. Coprecipitated ubiquitin levels were examined via western blot with antiubiquitin antibody. (G) Phosphorylated β-catenin (at Ser33/Ser37) versus total β-catenin was elevated upon RPIA knockdown. Gel images are shown in the up panel. (H) Overexpression of nondegradable β-catenin can overcome the growth inhibition induced by RPIA knockdown in HCT116 cells. The proliferation fold is compared to pMCV6 transfected control cell at first day. (I) The elevated viability induced by expression of RPIA was decreased upon ICRT14 (β-catenin inhibitor) treatment. Dose-dependent effects were revealed in HCT116 cells. (J) pGSK3β^Ser9 protein expression levels were up-regulated in the cytoplasmic extract upon overexpression of RPIA-WT but not upon RPIA-ΔD in HCT116 cells. (K) Cell proliferation was measured in RPIA knockdown HCT116 cells combined with 2.5 mM LiCl or 5 μM CHIR99021, respectively. The statistical significance was calculated with the Student <i>t</i> test (*** <i>P</i> < 0.001). Data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003714#pbio.2003714.s011" target="_blank">S3 Data</a>. CHX, cycloheximide; ERK, extracellular signal-regulated kinase; LiCl, lithium chloride; MG132, proteasome inhibitor; pcDNA, pcDNA3 vector control; pERK, phosphorylated-ERK; Rel, relative; RPIA-ΔD, RPIA deletion domain D mutant; RPIA, ribose-5-phosphate isomerase A; RPIA-WT, RPIA wild type; si-NC; negative control siRNA; siRNA, small interfering RNA; si-RPIA, RPIA small interfering RNA.</p

Brain-derived neurotrophic factor (BDNF) -TrKB signaling modulates cancer-endothelial cells interaction and affects the outcomes of triple negative breast cancer

<div><p>Aims</p><p>There is good evidence that the tumor microenvironment plays an important role in cancer metastasis and progression. Our previous studies have shown that brain-derived neurotrophic factor (BDNF) participates in the process of metastasis and in the migration of cancer cells. The aim of this study was to investigate the role of BDNF on the tumor cell microenvironment, namely, the cancer cell-endothelial cell interaction of TNBC cells.</p><p>Methods</p><p>We conducted oligoneucleotide microarray analysis of potential biomarkers that are able to differentiate recurrent TNBC from non-recurrent TNBC. The MDA-MB-231 and human endothelial HUVEC lines were used for this study and our approaches included functional studies, such as migration assay, as well as Western blot and real-time PCR analysis of migration and angiogenic signaling. In addition, we analyzed the survival outcome of TNBC breast cancer patients according to their expression level of BDNF using clinical samples.</p><p>Results</p><p>The results demonstrated that BDNF was able to bring about autocrinal (MDA-MB-231) and paracrinal (HUVECs) regulation of BDNF-TrkB gene expression and this affected cell migratory activity. The BDNF-induced migratory activity was blocked by inhibitors of ERK, PI3K and TrkB when MDA-MB-231 cells were examined, but only an inhibitor of ERK blocked this activity when HUVEC cells were used. Furthermore, decreased migratory activity was found for △BDNF and △TrkB cell lines. Ingenuity pathway analysis (IPA) of MDA-MB-231 cells showed that BDNF is a key factor that is able to regulate a network made up of metalloproteases and calmodulin. Protein expression levels in a tissue array of tumor slices were found to be correlated with patient prognosis and the results showed that there was significant correlation of TrkB expression, but not of BDNF. expressionwith patient DFS and OS.</p><p>Conclusion</p><p>Our study demonstrates that up-regulation of the BDNF signaling pathway seems tobe involved in the mechanism associated with early recurrence in triple negative breast cancer cell. In addition, BDNF can function in either an autocrine or a paracrine manner to increase the migration ability of both MDA-MB-231 cells and HUVEC cells. Finally, overexpression of TrkB, but not of BDNF, is significantly associated with a poor survival outcome for TNBC patients.</p></div

RPIA regulates colon cell proliferation through β-catenin expression in HCT116 cells.

<p>(A) Knockdown of RPIA significantly reduced cell proliferation and RPIA overexpression enhanced cell proliferation in HCT116 cells. Co-treatment of si-RPIA and pcDNA-RPIA rescued the reduction of cellular proliferation upon knockdown of RPIA in HCT116. Cell viability assays were performed by measuring the cells at the second, third, fourth, and fifth days and the proliferation fold is compared to control cell at the first day. Control: Co-transfect with scramble RNA and pcDNA empty vector as negative control. (B) RPIA knockdown significantly reduced colony formation ability, and RPIA overexpression enhanced colony formation ability in HCT116 cells. si-NC: Transfect with scramble siRNA as negative control. Representative images of colonies were shown on top of the quantification result. (C) Knockdown of RPIA reduced β-catenin protein levels as measured by western blotting (left panel) and quantified using Image J (middle panel) but did not significantly alter mRNA levels of β-catenin as measured by qPCR (right panel) in HCT116 cells. (D) RPIA overexpression increased β-catenin protein levels (left and middle panels) but did not affect β-catenin mRNA levels (right panel) in HCT116 cells. (E) Knock down of RPIA reduced the β-catenin/TCF luciferase reporter activity in HCT116 cells. (F) Overexpression of RPIA induced the β-catenin/TCF luciferase reporter activity in HCT116 cells. (G) Knockdown of RPIA decreased the mRNA levels of the β-catenin target genes <i>CCND1</i>, <i>CCNE2</i>, and <i>AXIN2</i> in HCT116 cells. (H) Overexpression of RPIA increased the mRNA levels of the β-catenin target genes <i>CCND1</i>, <i>CCNE2</i>, and <i>AXIN2</i> in HCT116 cells. The statistical significance was calculated with Student <i>t</i> test (* 0.01 < <i>P</i> < 0.05, ** 0.001 < <i>P</i> < 0.01, and *** <i>P</i> < 0.001). Data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003714#pbio.2003714.s010" target="_blank">S2 Data</a>. <i>AXIN2</i>, <i>Axis inhibition protein 2</i>; <i>CCND1</i>, <i>Cyclin D1</i>; <i>CCNE2</i>, <i>Cyclin E2</i>; <i>CTNNB1</i>, <i>CATENIN BETA 1</i>; pcDNA, pcDNA3 vector control; qPCR, quantitative PCR; RPIA, ribose-5-phosphate isomerase A; si-NC, negative control small interfering RNA; si-RPIA, RPIA small interfering RNA; TCF, T-cell transcription factor.</p

RPIA is highly expressed in different stages of CRC.

<p>(A) Representative RPIA IHC staining at different stages of colon cancer is shown. Scale bar: 500 μm. (B) The average IRS for RPIA staining showed significantly increased RPIA expression from stage I to IVB and the M. Stage III was divided into IIIB and IIIC, and stage IV was divided into IVA and IVB based on their subcategories. The statistical significance was calculated with Student <i>t</i> test (*** <i>P</i> < 0.001). (C) The average RPIA mRNA fold change in paired tissues (tumor tissue versus the surrounding normal tissue) from CRC patients at stages I to IV. The box plot indicates the median (central horizontal line), 75th percentile (the top of box), 25th percentile (the bottom of box), maximum value (the top end), minimum value (the bottom end), and the outlier (the point). Data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2003714#pbio.2003714.s009" target="_blank">S1 Data</a>. CRC, colorectal cancer; IHC, immunohistochemistry; IRS, immunoreactive score; M, metastasis stage; N, normal colorectal tissue; RPIA, ribose-5-phosphate isomerase A.</p

RPIA localizes in the nucleus and interacts with APC and β-catenin in HCT116 cells.

<p>(A) Nuclear localization of RPIA was detected by immunostaining with an anti-RPIA antibody (green) in HCT116 cells with and without overexpression of RPIA. DAPI was used to counterstain nuclei (blue). Scale bar: 50 μm. (B) Left panels: The cell lysates were precipitated by anti-APC, anti-β-catenin, and anti-RPIA antibodies in HCT116 cells. The APC, β-catenin, and RPIA interaction can be increased by RPIA-WT but not by RPIA-ΔD. Right panels: Protein loading input for IP assay of HCT116 cells. The orange boxes indicate the signals were enhanced by RPIA-WT but not in RPIA-ΔD. (C) Model of RPIA-β-catenin-APC interaction in HCT116 cell line. APC, adenomatous polyposis coli; Cyt, cytoplasm; IgA, immunoglobulin A; IP, immunoprecipitation; pcDNA, pcDNA3 vector control; Nu, nucleus; RPIA-ΔD, RPIA deletion domain D mutant; RPIA, ribose-5-phosphate isomerase A; RPIA-WT, RPIA wild type.</p