THE USE OF MANDAMUS TO COMPEL EDUCATIONAL INSTITUTIONS TO CONFER DEGREES

Hispolon is an active phenolic compound of <i>Phellinus igniarius</i>, a mushroom that was recently shown to have antioxidant and anticancer activities in various solid tumors. Here, the molecular mechanisms by which hispolon exerts anticancer effects in acute myeloid leukemia (AML) cells was investigated. The results showed that hispolon suppressed cell proliferation in the various AML cell lines. Furthermore, hispolon effectively induced apoptosis of HL-60 AML cells through caspases-8, -9, and -3 activations and PARP cleavage. Moreover, treatment of HL-60 cells with hispolon induced sustained activation of JNK1/2, and inhibition of JNK by JNK1/2 inhibitor or JNK1/2-specific siRNA significantly abolished the hispolon-induced activation of the caspase-8/-9/-3. In vivo, hispolon significantly reduced tumor growth in mice with HL-60 tumor xenografts. In hispolon-treated tumors, activation of caspase-3 and a decrease in Ki67-positive cells were observed. Our results indicated that hispolon may have the potential to serve as a therapeutic tool to treat AML

Macrophage Infiltration Induces Gastric Cancer Invasiveness by Activating the β-Catenin Pathway - Fig 4

<p>(A) Migration activity of THP-1 monocyte co-cultured with different gastric cancer cell lines (AGS, N87, MKN-45 and TSGH). THP-1 monocyte will be recruited by different gastric cancer cell lines and the recruiting ability is dependent on the degree of malignancy. (B) Invasion ability of GC cells treated by macrophage CM. Four different types of GC cells (AGS, MKN45, N87 and TSGH) were seeded in a Boyden chamber and co-cultured with or without macrophage cells or macrophage CM for 24 hours. Invasion abilities of each cell line were measured <i>in vitro</i> for 24 hours. All data represent the arithmetic mean ± SEM. * p < 0.05, ** p < 0.01. (C) Effect of macrophage CM co-cultured with cells. N87 and AGS cells were co-cultured with macrophage CM respectively for 24 hours and cell viability was determined by MTT assay.</p

Immunohistochemistry analysis.

<p>Immunostaining against CD68 was performed on tissue slides from gastric cancer patients. CD68+ macrophage staining was sparse in the normal gastric mucosa (A). CD68+ cells were detected in both the stroma and tumor nest (B & C). The density of CD68+ macrophages in pathological tumor staging (pT) 1 (D) and pT4 (E) tumor lesions.</p

Immunohistochemistry analysis.

<p>β-catenin was highly expressed in tumors. (A) In most cases, β-catenin expression was primarily located in the cytoplasm. However, (B) strong nuclear staining of β-catenin in GC cells was positively associated with CD68+ macrophages in the tumor lesions.</p

Effect of macrophage CM on β-catenin pathway.

<p>(A) β-catenin accumulates in nucleus after treatment with macrophage CM for 30 minutes in N87 cells. (B) Immunofiuorescent images were observed with a confocal microscope. β-catenin positive cells were analyzed by using an anti-β-catenin antibody, which is recognized by secondary rabbit antibody conjugated with FITC, depicted by green fluorescence; Nuclear staining was detected by counterstaining cells with 4', 6-Diamidino-2-phenylindole (DAPI), represented as blue fluorescence. Co-culturing with macrophage CM, immunofiuorescence staining showed β-catenin-FITC complexes translocated into nucleus. (C) Invasion ability of GC cells treated by macrophage CM.</p

Kaplan-Meier overall survival curves for gastric cancer patients as TAM density (Low: TAM<671, blue line; High: TAM>671, brown line; <i>p</i> = 0.0073).

<p>Kaplan-Meier overall survival curves for gastric cancer patients as TAM density (Low: TAM<671, blue line; High: TAM>671, brown line; <i>p</i> = 0.0073).</p

Macrophage Infiltration Induces Gastric Cancer Invasiveness by Activating the β-Catenin Pathway - Fig 6

<p>(A) Effect of macrophage CM on mRNA and protein expression of β-catenin down-stream genes. N87 cells were cultured in the presence or absence of macrophage CM. For RNA and protein level, cells were collected after 6 hours and 24 hour treatment respectively. (B) N87 cells with or without knock-down of β-catenin utilizing shRNA-2 were co-treated in the presence or absence of macrophage CM. N87 cells were collected after 24 hours treatment and analyzed with western blot. (C) Effect of MMP7 neutralized antibody on GC cells invasion. N87 cells in the presence of macrophage CM for 24 hours were pre-treated with various doses MMP7 neutralized antibody (0, 0.625, 1.25 and 2.5μg/ml) for 1 hour Isotype IgG was used as a blocking control. (D) Effect of CD44 neutralized antibody on GC cell invasion. N87 cells in the presence of macrophage CM for 24 hours were pre-treated with various doses CD44 neutralized antibody (0, 1.25, 2.5, 5 and 10 μg/ml) for 1 hour. Isotype IgG was used as a blocking control. All data represent the arithmetic mean ± SEM. * <i>p</i> < 0.05.</p

Soluble AXL: A Possible Circulating Biomarker for Neurofibromatosis Type 1 Related Tumor Burden

<div><p>Neurofibromatosis type 1 (NF1) is the most common tumor predisposition disorder affecting 1/3500 worldwide. Patients are at risk of developing benign (neurofibromas) and malignant peripheral nerve sheath tumors (MPNST). The AXL receptor tyrosine kinase has been implicated in several kinds of cancers, but so far no studies have investigated the role of AXL in NF1 related tumorigenesis. Recently, the soluble fraction from the extracellular domain of AXL (sAXL) has been found in human plasma, and its level was correlated to poor prognosis in patients with renal cancer. Compared to normal human Schwann cells, a significantly high expression level of AXL was found in three of the four MPNST cell lines and two of the three primary MPNST tissues. Similarly, the level of sAXL in conditioned media corresponded to the protein and mRNA levels of AXL in the MPNST cell lines. Furthermore, in two different human MPNST xenograft models, the human sAXL could be detected in the mouse plasma. Its level was proportionate to the size of the xenograft tumors, while no human sAXL was detect prior to the formation of the tumors. Treatment with a newly developed photodynamic therapy, prevented further tumor growth and resulted in drastically reduced the levels of sAXL compared to that of the control group. Finally, the level of sAXL was significantly increased in patients with plexiform tumors compared to patients with only dermal neurofibromas, further supporting the role of sAXL as a marker for NF1 related tumor burden.</p></div

Release of sAXL into the conditioned media of MPNST cell lines.

<p><b>A</b>, all four MPNST cell lines (S462, STS26T, ST8814 and T265) and NHSC released sAXL into the conditioned media, at levels that roughly comparable to the protein levels of full length AXL in the cells (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115916#pone-0115916-g002" target="_blank">Fig. 2A</a>). A highly-matched correlation was found between the time in culture and the sAXL concentration in all four cell lines and NHSC (symbol cross for S462 cells with a coefficient of determination, R² = 0.97; diamonds for STS26T with R² = 0.99, squares for ST8814 with R² = 0.97, triangles for T265 with R² = 0.99 and star for NHSC R² = 0.94). To further highlight that all cell lines release sAXL at a constant rate, the data in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115916#pone-0115916-g003" target="_blank">Fig. 3A</a> is presented as 5 individual graphs in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115916#pone.0115916.s001" target="_blank">S1 Fig. <b>B–C</b></a>, Using specific lentiviral shRNA clones, the expression of AXL and TYRO3 has been knocked down in MPNST cell line T265, either alone or in combination to knockdown both genes (shDual). Successful silencing was verified with quantitative-PCR (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115916#pone-0115916-g003" target="_blank">Fig. 3B</a>) and Western blotting (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0115916#pone-0115916-g003" target="_blank">Fig. 3C</a>). <b>D</b>, the levels of sAXL in the cell medium were drastically reduced in AXL-knockdown cells. In contrast, knockdown of TYRO3 did not affected the release of sAXL compared to the empty vector (GIPZ). In each case the level of sAXL in the conditioned media corresponded to the mRNA levels in the cells.</p

Mice harboring human MPNST (STS26T) xenograft tumors release human-soluble AXL into the blood.

<p>There was a trend of size dependent increasing of sAXL level until the tumors reached 2000 mm³. The coefficient of determination, R² was 0.884.</p