Nasopharyngeal undifferentiated carcinoma with sarcomatoid features: Pitfalls in the immunohistochemistry.

We present a case of an undifferentiated subtype of non-keratinizing squamous cell carcinoma (NK-SCC) with sarcomatoid features in the nasopharynx in a 69-year-old man who was difficult to diagnose due to spindle-shaped malignant cells. He was admitted because of a right nasal obstruction and right headache, and imaging revealed a heterogeneously enhanced irregularly shaped mass at the nasopharynx. Histopathologically, the tumour was partially organised, and the tumour cells were epithelioid or spindle-shaped. Initially, we erroneously diagnosed the tumour as an angiosarcoma owing to its false-negative immunoreaction for cytokeratins and a mistaken interpretation for CD31. After in situ hybridization for Epstein-Barr virus was positive, a consultation and additional immunostaining (including re-staining for cytokeratin with varying dilutions) were performed, and the diagnosis was revised to NK-SCC with sarcomatoid features. We believe that sarcomatoid features may be observed in nasopharyngeal carcinoma and in this case, immunostaining using various epithelial markers is necessary and careful attention should be paid to the interpretation of immunostaining.

Hypomethylation reduced the aggressive potential of human malignant mesothelioma cells.

Epigenetic modifications have been implicated in the development of therapeutic resistance responsible for the poor prognosis of human malignant mesothelioma (HMM). To find a potential way to overcome this therapeutic resistance, this study investigated the anticancer effects of a DNA demethylating agent, 5-Aza-2'-Deoxycytidine (5-aza-dC), on HMM cells. The 5-aza-dC exhibited minimal detrimental effects on cell survival. However, treatment with 5-aza-dC significantly altered the biological characteristics associated with malignancy, such as cell migration and cell interaction, colony-forming capacity, and invasiveness. Moreover, it significantly reduced the fraction of side population (SP) cells, which are reportedly enriched for more aggressive cells. Large-scale methylation analysis based on methylated DNA immunoprecipitation revealed a more than two fold increase in the methylation level of major tumor suppressor genes in the SP fraction. The data indicated that SP cells might acquire malignancy by hypermethylation of tumor suppressor genes. Treatment with 5-aza-dC could attack more malignant cells through the modification of their methylation status. The results indicate that the modulation of DNA methylation might be a valuable strategy to overcome the therapeutic resistance of HMM. Moreover, ensuing changes in the biological characteristics provide a basis for further analysis of the role of methylation in HMM carcinogenesis.

Inhibition of hedgehog signaling reduces the side population in human malignant mesothelioma cell lines.

Deregulation of crucial embryonic pathways, including hedgehog signaling, has been frequently implicated in a variety of human cancers and is emerging as an important target for anticancer therapy. This study evaluated the potential anticancer effects of cyclopamine, a chemical inhibitor of hedgehog signaling, in human malignant mesothelioma (HMM) cell lines. Cyclopamine treatment significantly decreased the proliferation of HMM cells by promoting apoptosis and shifting the cell cycle toward dormant phase. The clonogenicity and mobility of HMM cells were significantly decreased by cyclopamine treatment. Treatment of HMM cells with cyclopamine significantly reduced the abundance of side population cells, which were measured using an assay composed of Hoechst 33342 dye staining and subsequent flow cytometry. Furthermore, the expression levels of stemness-related genes were significantly affected by cyclopamine treatment. Taken together, the present study showed that targeting hedgehog signaling could reduce a more aggressive subpopulation of the cancer cells, suggesting an alternative approach for HMM therapy.

A reusable robust radio frequency biosensor using microwave resonator by integrated passive device technology for quantitative detection of glucose level.

A reusable robust radio frequency (RF) biosensor with a rectangular meandered line (RML) resonator on a gallium arsenide substrate by integrated passive device (IPD) technology was designed, fabricated and tested to enable the real-time identification of the glucose level in human serum. The air-bridge structure fabricated by an IPD technology was applied to the RML resonator to improve its sensitivity by increasing the magnitude of the return loss (S21). The resonance behaviour, based on S21 characteristics of the biosensor, was analysed at 9.20 GHz with human serum containing different glucose concentration ranging from 148-268 mg dl(-1), 105-225 mg dl(-1) and at a deionised (D) water glucose concentration in the range of 25- 500 mg dl(-1) for seven different samples. A calibration analysis was performed for the human serum from two different subjects and for D-glucose at a response time of 60 s; the reproducibility, the minimum shift in resonance frequency and the long-term stability of the signal were investigated. The feature characteristics based on the resonance concept after the use of serum as an analyte are modelled as an inductor, capacitor and resistor. The findings support the development of resonance-based sensing with an excellent sensitivity of 1.08 MHz per 1 mg dl(-1), a detection limit of 8.01 mg dl(-1), and a limit of quantisation of 24.30 mg dl(-1).

Pancreatic adenocarcinoma upregulated factor, a novel endothelial activator, promotes angiogenesis and vascular permeability.

Pancreatic adenocarcinoma upregulated factor (PAUF) was recently reported to be a metastasis factor for pancreatic cancer cells. Here, we demonstrate a novel role for PAUF as a potent endothelial activator, promoting both angiogenesis and vascular permeability. Overexpression of PAUF in a mouse pancreatic cancer model resulted in increased tumor vascularity. Recombinant PAUF (rPAUF) enhanced proliferation, migration and capillary-like tube formation of human endothelial cells (ECs), consistently with increased neovascularization in vivo. rPAUF also increased endothelial permeability through the disruption of vascular endothelial-cadherin-facilitated cell-cell junctions in vitro and induced vascular leakage in mouse skin. These effects were attenuated upon treatment with an antibody against PAUF. Moreover, PAUF evoked a time- and dose-dependent activation of extracellular signal-regulated kinase (ERK)1/2, AKT and endothelial NO synthase (eNOS) in ECs, which are closely linked to rPAUF-induced angiogenesis. Finally, rPAUF upregulated the expression of C-X-C chemokine receptor 4 (CXCR4) in ECs and potentiated the in vitro and in vivo EC angiogenic responses to stromal cell-derived factor-1 (SDF-1), a ligand for CXCR4. Taken together, these data demonstrate that PAUF has a novel function in promoting angiogenesis and vascular permeability. Our findings suggest new possibilities for PAUF's role in the pathogenesis of angiogenesis-dependent diseases.

Effect of lifestyles on the blood mercury level in Korean adults.

Mercury (Hg) is widely distributed in the environment and oral exposure is a main route in the general population. In this study, we estimated the dietary intake of Hg and its relationship with blood Hg levels in Korean adults. The study subjects were recruited from three different districts (rural: 189, coastal: 208 and urban: 184). We used a general questionnaire to collect information about demographic factors, lifestyles and diet. Dietary habits were studied using the 24-h recall method. The estimation of Hg intake was performed using the database of Hg contents in 128 Korean foods based on the previous studies. Blood Hg was analyzed using Direct Mercury Analyzer with the gold-amalgam method. Daily intake of Hg by diet was estimated at 13.57 µg (0.22 µg/kg body weight). The geometric mean Hg concentration in whole blood was 3.92 µg/L. Blood Hg level and Hg intake by diet was higher in coastal areas than in urban or rural areas, respectively. Blood Hg level correlated with the intake of Hg consumed from diet. Seafood was highly responsible and account for 75.6% of total dietary Hg intake. In this study, blood Hg concentrations were found to be significantly affected by sex, age, individual lifestyles and especially the amount of seafood intake, which might play an important role in determining blood Hg levels in Korean adults.

Interleukin-8 and its receptor CXCR2 in the tumour microenvironment promote colon cancer growth, progression and metastasis.

BACKGROUND: Colorectal cancer (CRC) is a leading cause of death in the United States. Increased level of interleukin-8 (IL-8) and CXCR2 on tumours and in the tumour microenvironment has been associated with CRC growth, progression and recurrence in patients. Here, we aimed to evaluate the effects of tissue microenvironment-encoded IL-8 and CXCR2 on colon cancer progression and metastasis. METHODS: A novel immunodeficient, skin-specific IL-8-expressing transgenic model was generated to evaluate colon cancer growth and metastasis. Syngeneic mouse colon cancer cells were grafted in CXCR2 knockout (KO) mice to study the contribution of CXCR2 in the microenvironment to cancer growth. RESULTS: Elevated levels of IL-8 in the serum and tumour microenvironment profoundly enhanced the growth of human and mouse colon cancer cells with increased peri-tumoural angiogenesis, and also promoted the extravasation of the cancer cells into the lung and liver. The tumour growth was inhibited in CXCR2 KO mice with significantly reduced tumour angiogenesis and increased tumour necrosis. CONCLUSION: Increased expression of IL-8 in the tumour microenvironment enhanced colon cancer growth and metastasis. Moreover, the absence of its receptor CXCR2 in the tumour microenvironment prevented colon cancer cell growth. Together, our study demonstrates the critical roles of the tumour microenvironment-encoded IL-8/CXCR2 in colon cancer pathogenesis, validating the pathway as an important therapeutic target.

Effect of rebamipide in treatment of Helicobacter pylori-associated duodenal ulcer: attenuation of chemokine expression and nitrosative damage.

Production of cytokines along with increased activity of nitric oxide synthase has been implicated as one of the contributing mechanisms of Helicobacter pylori-mediated gastroduodenal diseases. We aimed to evaluate the effect of rebamipide in treating Helicobacter pylori-associated duodenal ulcers in terms of cytokine production and nitrosative damage of the gastric mucosa. In patients with duodenal ulcers, rebamipide or placebo were given randomly after eradication. Mucosal cytokine production was measured by enzyme linked immunoassay, and nitrotyrosine immunoexpression was measured by immunohistochemistry. The inflammatory activity and degree of neutrophil infiltration were graded accordingly. The mucosal production of RANTES, interleukin-8, and TNF-alpha showed a significant decrease after eradication in patients with rebamipide after-treatment. The nitrotyrosine immunoreactivity of gastric epithelium was significantly decreased in the rebamipide group. Rebamipide treatment after eradication resulted in a significant reduction in chemokine production along with nitrotyrosine immunoexpression in Helicobacter pylori-associated duodenal ulcers.

Diarylheptanoids with free radical scavenging and hepatoprotective activity in vitro from Curcuma longa.

Assay-guided fractionation of the EtOAc soluble fraction of the rhizomes of Curcuma longa furnished three DPPH free radical scavenging diarylheptanoids, curcumin (1), demethoxycurcumin (2), and bisdemethoxycurcumin (3). Compounds 1-3 showed the DPPH radical scavenging effects with IC(50) values of 2.8, 39.2, 308.7 microM, respectively. L-Ascorbic acid and resveratrol as positive controls exhibited IC(50) values of 22.5 and 25.0 microM, respectively. Compounds 1-3 showed significant hepatoprotective effects on tacrine-induced cytotoxicity in human liver-derived Hep G2 cells. The EC(50) values of 1-3 are 86.9, 70.7, and 50.2 microM, respectively. Silybin (EC(50) = 69.0 microM) and silychristin (EC(50) = 82.7 microM) were used as positive controls.

Roles of extracellular signal-regulated kinase and p38 mitogen-activated protein kinase in apoptosis of human monoblastic leukemia U937 cells by lectin-II isolated from Korean mistletoe.

The mitogen-activated protein kinase (MAPK) family members have been implicated in cell survival. We have previously demonstrated that cytotoxic lectin-II isolated from Korean mistletoe induces apoptotic cell death in the human monoblastic leukemia cell line, U937, via the activation of the stress-activated protein kinases/c-Jun N-terminal kinase (SAPK/JNK). In the present study, the roles of extracellular signal-regulated kinases (ERK1/2) and p38 MAPK in lectin-II-induced apoptosis have been investigated. Treatment of U937 cells with lectin-II resulted in apoptotic DNA fragmentation, which was preceded by the activation of ERK1/2, p38 MAPK and SAPK/JNK. This lectin-II-induced DNA fragmentation was significantly enhanced when ERK1/2 activation was selectively inhibited by PD098059. 12-O-tetradecanoylphorbol-13-acetate, which stimulates ERK activity in U937 cells, markedly reduced lectin-II-induced DNA fragmentation. Inhibition of p38 MAPK activity with p38-specific inhibitor, SB203580, partially inhibited lectin-II-induced DNA fragmentation. These results suggest that ERK1/2 and p38 MAPK may have opposite effects on cell survival in response to cytotoxic mistletoe lectin-II, which may contribute to the modulation of lectin-II-mediated cytotoxic activity.

4-Acetyl-12,13-epoxyl-9-trichothecene-3,15-diol isolated from the fruiting bodies of Isariajaponica Yasuda induces apoptosis of human leukemia cells (HL-60).

The fruiting bodies of Isaria fungi have been traditionally used in Korea to treat cancer. An apoptosis-inducing compound, 4-acetyl-12,13-epoxyl-9-trichothecene-3,15-diol, was isolated from the methanol extract of fruiting bodies of Isaria japonica Yasuda by bioassay-guided fractionation. The apoptosis of the human leukemia cells (HL-60) by the compound was accessed by propidium iodide-staining flow cytometric analysis, and apoptosis-inducing activity at IC50 concentration (10 nmol/l) was further confirmed by a nuclear morphological change, a ladder pattern of internucleosomal DNA fragmentation, and an activation of caspase-3.

Butein, a plant polyphenol, induces apoptosis concomitant with increased caspase-3 activity, decreased Bcl-2 expression and increased Bax expression in HL-60 cells.

In the present study we have investigated whether butein could induce apoptosis in human leukaemic HL-60 cells. The treatment of HL-60 cells with butein induced apoptotic cell death as determined by morphological and biochemical changes. Apoptotic DNA fragments in the butein-treated HL-60 cells were increased gradually as determined by flow cytometric analysis. The caspase-3 activity was increased during butein-induced apoptosis. However, caspase-3 inhibitor abrogated the butein-induced DNA fragmentation. Furthermore, the treatment of HL-60 cells with butein decreased the expression of Bcl-2 protein, but increased the expression of Bax protein. These results suggest that butein-induced apoptosis is mediated through the activation of caspase-3 and it is associated with changed expression of Bcl-2 and Bax proteins.

Capsazepine, a vanilloid receptor antagonist, inhibits the expression of inducible nitric oxide synthase gene in lipopolysaccharide-stimulated RAW264.7 macrophages through the inactivation of nuclear transcription factor-kappa B.

High amounts of nitric oxide (NO) production following the induction of inducible NO synthase (iNOS) gene expression has been implicated in the pathogenesis of inflammatory diseases. Capsaicin, a vanilloid receptor agonist, is known to have an inhibitory effect on NO production in macrophages. In the present study, we have found that capsazepine (CAPZ), a vanilloid receptor antagonist, also inhibited NO and iNOS protein syntheses induced by lipopolysaccharide in RAW264.7 macrophages via the suppression of iNOS mRNA. The mechanistic studies showed that CAPZ inhibited the expression of iNOS mRNA through the inactivation of nuclear transcription factor-kappa B (NF-kappa B). Thus, capsazepine may be a useful candidate for the development of a drug to treat inflammatory diseases related to iNOS gene overexpression.

Inhibitory effects of methanol extract of Cyperus rotundus rhizomes on nitric oxide and superoxide productions by murine macrophage cell line, RAW 264.7 cells.

The rhizomes of Cyperus rotundus (C. rotundus) have been used in oriental traditional medicines for the treatment of stomach and bowel disorders, and inflammatory diseases. Nitric oxide (NO) and superoxide (O2-) are important mediators in the pathogenesis of inflammatory diseases. This study was undertaken to address whether the metanol (MeOH) extract of rhizomes of C. rotundus could modulate NO and O2- productions by murine macrophage cell line, RAW 264.7 cells. The MeOH extract of rhizomes of C. rotundus showed the inhibition of NO production in a dose-dependent manner by RAW 264.7 cells stimulated with interferon-gamma plus lipopolysaccharide. The inhibition of NO production by the extract was due to the suppression of iNOS protein, as well as iNOS mRNA expression, determined by Western and Northern blotting analyses, respectively. In addition, the MeOH extract suppressed the production of O2- by phorbol ester-stimulated RAW 264.7 cells in dose- and time-dependent manners. Collectively, these results suggest that the MeOH extract of rhizomes of C. rotundus could be developed as anti-inflammatory candidate for the treatment of inflammatory diseases mediated by overproduction of NO and O2-.

The aqueous extract of Rhodiola sachalinensis root enhances the expression of inducible nitric oxide synthase gene in RAW264.7 macrophages.

In the present study, we examined the effects of the aqueous extract of Rhodiola sachalinensis root (RSE) on the expression of inducible nitric oxide (NO) synthase (iNOS) gene in RAW264.7 macrophages. RSE synergistically increased NO synthesis in interferon-gamma-primed macrophages. Reverse transcriptase polymerase chain reaction and Northern blotting analysis revealed that RSE may provide a second triggering signal for the synergistic induction of iNOS mRNA expression. Thus, iNOS-mediated NO synthesis in response to RSE may be one mechanism whereby this herbal medicine elicits its therapeutic effects.

Induction of granulocytic differentiation in acute promyelocytic leukemia cells (HL-60) by water-soluble chitosan oligomer.

Water-soluble chitosan oligomer (WSCO) has been reported to have anticancer activity, immuno-enhancing effect and antimicrobial activity. However, other biological activities are unknown. Herein, we have shown that WSCO is able to inhibit proliferation of human leukemia HL-60 cells and induce these cells to differentiate. Treatment with WSCO for 4 days resulted in a concentration-dependent reduction in HL-60 cell growth as measured by cell counting and MTT assay. This effect was accompanied by a marked increase in the proportion of G(0)/G(1) cells as measured by flow cytometry. WSCO also induced differentiation of the cells as measured by phorbol ester-dependent reduction of NBT, morphological changes as examined by Wright-Giemsa staining and expression of CD11b but not of CD14 as analysed by flow cytometry, indicating differentiation of HL-60 cells toward granulocyte-like cells. A combination of low dose of WSCO with all-trans retinoic acid, a differentiating agent toward granulocyte-like cells, exhibited a synergistic effect on the differentiation. In addition, treatment of HL-60 cells with WSCO for 6 or 8 days resulted in the induction of apoptosis as assayed qualitatively by agarose gel electrophoresis and quantitatively by Annexin V technique using flow cytometry. Collectively, there is a potential for WSCO in the treatment of myeloid leukemia.

Inhibitory effects of butanol fraction of the aqueous extract of Forsythia koreana on the nitric oxide production by murine macrophage-like RAW 264.7 cells.

The aim of this study was to investigate the effect of butanol fraction of the aqueous extract of Forsythia koreana fruits on the nitric oxide (NO) production and inducible nitric oxide synthesis (iNOS) gene expression in murine macrophage-like RAW 264.7 cells. Butanol fraction alone affected neither NO production nor iNOS gene expression in macrophage-like RAW 264.7 cells. However, the butanol fraction inhibited NO production and iNOS gene expression in RAW 264. 7 cells stimulated with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS). These findings suggest that inhibition of NO production by this butanol fraction in RAW 264.7 cells stimulated with IFN-gamma plus LPS was due to the suppression of iNOS gene expression.

Polymethoxyflavonoids from Vitex rotundifolia inhibit proliferation by inducing apoptosis in human myeloid leukemia cells.

Three polymethoxyflavonoids from the fruit of Vitex rotundifolia, namely 2',3',5-trihydroxy-3,6,7-trimethoxyflavone (Vx-1), vitexicarpin (Vx-5) and artemetin (Vx-6), were tested for their antiproliferative activity in human myeloid leukemia HL-60 cells. They showed a dose-dependent decrease in the growth of HL-60 cells. The concentrations required for 50% inhibition of the growth (IC50) after 96 h were 4.03 microM, 0.12 microM and 30.98 microM for Vx-1, Vx-5 and Vx-6, respectively. Treatment of HL-60 cells with the flavonoids induced morphological changes that are characteristic of apoptosis. We judged the induction of apoptosis by the detection of DNA fragmentation in agarose gel electrophoresis and the degree of apoptosis was quantified by a double-antibody sandwich ELISA and by flow cytometric analysis. The C-3 hydroxyl and C-8 methoxyl groups were found not to be essential for the activity, but the C-3' methoxyl instead of hydroxyl group lowered the antiproliferative and apoptosis inducing activity. These results suggest that the polymethoxyflavonoids isolated from V. rotundifolia may be used as potential chemopreventive and chemotherapeutic agents.

Synergistic cooperation between water-soluble chitosan oligomers and interferon-gamma for induction of nitric oxide synthesis and tumoricidal activity in murine peritoneal macrophages.

The effects of water-soluble chitosan oligomers (WSCO) on the synthesis of nitric oxide (NO) by murine peritoneal macrophages and on macrophage-mediated cytotoxicity towards murine fibrosarcoma Meth A cells were investigated. WSCO alone had no effect on NO synthesis and killing of tumor cells. However, treatment of macrophages with a combination of WSCO and interferon-gamma (IFN-gamma) synergically increased NO synthesis and enhanced killing of tumor cells. The synergism between IFN-gamma and WSCO in NO synthesis and tumoricidal activity was mainly dependent on increased secretion of tumor necrosis factor-alpha by WSCO.